se 02glhs and reyli,~xes through the abdoj.!inal
TRANSCRIPT
SElSE 02GLHS AND REYLI~XES THROUGH THE ABDOJINAL
NERVB CORD O THE COCKROACH
BIAlnmQsect CRAIn IPR
A Ihesjn
Submitted to
the Department of BioloCY
Kansas State Teachers ColleGe Emporia Kansas
In Partial Fulfillment
of the Requirements for the Degree
Master of Science
by
J Grant Kehler
August 1968
Approved for Major Department
) j=J cL --lv__
for Graduate ~l
-r_ ~
( Jv-o-(
4
r
Approved Cj
y ----1I -1 tV ( 1 _t_I~ 1
I
11
ACKWILEDGErrENT
I thenk Dr Edllard C Rowe for his invaluable
assistance and advice during the course of this work
thank Dr Katherine N Smalley for helpful discussions
and for the use of her diagram showing the relationship
of the lateral chordotonal organ to the cuticle I
thank Mrs Cynthia Brolm for her technical assistance
with the histological studies and my wife Patti for
typing the manuscript
iii
TABLE OF CONTBNTS
PAGE
INTRODUCTIon bullbull HATERIALS AND HETHODS 4
A Alimals bullbull middot 4
B Ana tomical bull 4
C Physiological 6
RESULTS 10
I Anatomical and Physiological Description of
the Abdominal Nerves and Sense Organs 10
A Ventral nerves and sensory elements 14
VN-1 14
VN-2 middot 14
VN-3 middot 15
VN-3A and AN-3Aa 15
Anatomy and physiology of the mid-
sternal rec eptor bull 16
B Dorsal nerve and sensory elements 27
DN-1 middot 27
DN-2 middot 28
DN-3 bull middot 28
DN-3A middot 30
DN-3B middot 30
iv
PAGE
Physiology of the D~-3A and DN-3B
receptors including the lateral
chordotonal organ 33
DN-4A middot 39
DN-4B inclUding the cardiac segmental
nerve 41
Relationship of DN-4 to the heart rate bullbull 43
Lateral cardiac nerve 47
II Responses Triggered by the Ventral Receptors 47
A Ventral nerve to homolateral dorsal
nerve pathvray 49
B Ventral nerve to heterolateral dorsal
nerve pathiay 53
C Ventral nerve rebound pathway bullbullbullbullbull 53
D Repetitive response in the homolateral
connective 57
E Responses of the cardiac lateral nerve bull 57
F Experiments to establish function of
ventral nerve-dorsal nerve reflex bullbullbull 59
1 Ventral nerve-dorsal nerve reflex
initiated by the C02 sensitive
elements of the mid-sternal receptor bull 61
2 Effects of respiratory movement on
dorsal and ventral nerve afferent
activitybull 62
v
PAGE
3 Reflex responses to pressure
which activates the mid-sternal
receptor 64
4 Reflex responses to groundborne
vilIra tlons 64
III Responses Triggered by the DN-3B Receptors bullbullbullbull 66
A Dorsal nerve to homolateral ventral
nerve pa thrray 66
B Dorsal nerve rebound 68
C Dorsal nerve to heterolateral dorsal nerve
pa th-ray bull bull bull bull bull bull bull bull 71
D Experiments to establish function of
dorsal nerve to ventral nerve reflex bullbullbull 74
IV Stability of Reflexes 80
V Further Interganglionic Pa thrays bullbullbullbullbullbull 82
A Spread of ventral nerve input bullbullbullbull 84
B Spread of dorsal nerve input bullbullbull 84
C Several generalizations about the
dissemination of sensory information bullbull 85
VI EVidence for Proprioceptive Reflex System
Behavioral Observations bullbullbullbullbull 97
DISCUSSION 99
I Nid-sternal Receptor 99
II Lateral Chordotonal Organs 100
III Functions of the Mid-sternal Receptor and
Lateral Choraotonal Organ bullbullbullbullbullbullbullbullbull 101
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
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VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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ure
1
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Rhy
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N-2
[1J
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
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-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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1---middot_--c ---J)rVo~
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
~)( Cf~r~lt (1 lmiddotmiddotmiddot p r -Itoshy ~) i tmiddott~r~~J _~ - ~~ V~~k-gt n l-
to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
I $Fi C shy
rebound Rnd simulteneouG neXv~e stimulat1on
VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
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tor
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rsal
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ower
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ure
47
D
ors
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
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B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
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Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
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Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
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tera
l (u
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er tr
ace)
and
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late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
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stim
ula
te
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ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
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ACKWILEDGErrENT
I thenk Dr Edllard C Rowe for his invaluable
assistance and advice during the course of this work
thank Dr Katherine N Smalley for helpful discussions
and for the use of her diagram showing the relationship
of the lateral chordotonal organ to the cuticle I
thank Mrs Cynthia Brolm for her technical assistance
with the histological studies and my wife Patti for
typing the manuscript
iii
TABLE OF CONTBNTS
PAGE
INTRODUCTIon bullbull HATERIALS AND HETHODS 4
A Alimals bullbull middot 4
B Ana tomical bull 4
C Physiological 6
RESULTS 10
I Anatomical and Physiological Description of
the Abdominal Nerves and Sense Organs 10
A Ventral nerves and sensory elements 14
VN-1 14
VN-2 middot 14
VN-3 middot 15
VN-3A and AN-3Aa 15
Anatomy and physiology of the mid-
sternal rec eptor bull 16
B Dorsal nerve and sensory elements 27
DN-1 middot 27
DN-2 middot 28
DN-3 bull middot 28
DN-3A middot 30
DN-3B middot 30
iv
PAGE
Physiology of the D~-3A and DN-3B
receptors including the lateral
chordotonal organ 33
DN-4A middot 39
DN-4B inclUding the cardiac segmental
nerve 41
Relationship of DN-4 to the heart rate bullbull 43
Lateral cardiac nerve 47
II Responses Triggered by the Ventral Receptors 47
A Ventral nerve to homolateral dorsal
nerve pathvray 49
B Ventral nerve to heterolateral dorsal
nerve pathiay 53
C Ventral nerve rebound pathway bullbullbullbullbull 53
D Repetitive response in the homolateral
connective 57
E Responses of the cardiac lateral nerve bull 57
F Experiments to establish function of
ventral nerve-dorsal nerve reflex bullbullbull 59
1 Ventral nerve-dorsal nerve reflex
initiated by the C02 sensitive
elements of the mid-sternal receptor bull 61
2 Effects of respiratory movement on
dorsal and ventral nerve afferent
activitybull 62
v
PAGE
3 Reflex responses to pressure
which activates the mid-sternal
receptor 64
4 Reflex responses to groundborne
vilIra tlons 64
III Responses Triggered by the DN-3B Receptors bullbullbullbull 66
A Dorsal nerve to homolateral ventral
nerve pa thrray 66
B Dorsal nerve rebound 68
C Dorsal nerve to heterolateral dorsal nerve
pa th-ray bull bull bull bull bull bull bull bull 71
D Experiments to establish function of
dorsal nerve to ventral nerve reflex bullbullbull 74
IV Stability of Reflexes 80
V Further Interganglionic Pa thrays bullbullbullbullbullbull 82
A Spread of ventral nerve input bullbullbullbull 84
B Spread of dorsal nerve input bullbullbull 84
C Several generalizations about the
dissemination of sensory information bullbull 85
VI EVidence for Proprioceptive Reflex System
Behavioral Observations bullbullbullbullbull 97
DISCUSSION 99
I Nid-sternal Receptor 99
II Lateral Chordotonal Organs 100
III Functions of the Mid-sternal Receptor and
Lateral Choraotonal Organ bullbullbullbullbullbullbullbullbull 101
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
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VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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Fig
ure
1
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Rhy
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ic acti
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on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
~ i
-5 i
-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
bullt ~ r - Cmiddotlt r ~- --~ ~ - -J 8UOCGJ rO+OIi -Jl~lvtG0P lJli (i~-G~-c ~~IOl~) f~t[+~ L~UOO
81 Ul HUOQSGJ ~U2l2jJj3 ~j(1pi)GYJ- middotX81Jl1I [IltgtU TT3JIOP
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bull uo ~~t T~TnurIgt~s aAIBU T~3Jl-TTGA 0+ 2AX2U 11G~rOp 311+ r~o ()J1IOd~~QX KrGU11~r(i ~~~ GJnC1~
1---middot_--c ---J)rVo~
o~
51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
s 1J
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
~i-u~~ h0 tc~4r1t~r ormiddot ~r~~ vOl+r-1 ~-~v~ cnv1~~ ~+~J if 2-r -~ J~t~) 0 1t -bullbull c-_ 1-1 lt_ -~ bull il -C~
stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
~)( Cf~r~lt (1 lmiddotmiddotmiddot p r -Itoshy ~) i tmiddott~r~~J _~ - ~~ V~~k-gt n l-
to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
I $Fi C shy
rebound Rnd simulteneouG neXv~e stimulat1on
VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
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tor
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rsal
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icate
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tor
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ower
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ure
47
D
ors
al a
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ion
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
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middot~middotmiddot1middotmiddotmiddot
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B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
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I
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Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
ho
mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
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stim
ula
te
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ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
I
11
ACKWILEDGErrENT
I thenk Dr Edllard C Rowe for his invaluable
assistance and advice during the course of this work
thank Dr Katherine N Smalley for helpful discussions
and for the use of her diagram showing the relationship
of the lateral chordotonal organ to the cuticle I
thank Mrs Cynthia Brolm for her technical assistance
with the histological studies and my wife Patti for
typing the manuscript
iii
TABLE OF CONTBNTS
PAGE
INTRODUCTIon bullbull HATERIALS AND HETHODS 4
A Alimals bullbull middot 4
B Ana tomical bull 4
C Physiological 6
RESULTS 10
I Anatomical and Physiological Description of
the Abdominal Nerves and Sense Organs 10
A Ventral nerves and sensory elements 14
VN-1 14
VN-2 middot 14
VN-3 middot 15
VN-3A and AN-3Aa 15
Anatomy and physiology of the mid-
sternal rec eptor bull 16
B Dorsal nerve and sensory elements 27
DN-1 middot 27
DN-2 middot 28
DN-3 bull middot 28
DN-3A middot 30
DN-3B middot 30
iv
PAGE
Physiology of the D~-3A and DN-3B
receptors including the lateral
chordotonal organ 33
DN-4A middot 39
DN-4B inclUding the cardiac segmental
nerve 41
Relationship of DN-4 to the heart rate bullbull 43
Lateral cardiac nerve 47
II Responses Triggered by the Ventral Receptors 47
A Ventral nerve to homolateral dorsal
nerve pathvray 49
B Ventral nerve to heterolateral dorsal
nerve pathiay 53
C Ventral nerve rebound pathway bullbullbullbullbull 53
D Repetitive response in the homolateral
connective 57
E Responses of the cardiac lateral nerve bull 57
F Experiments to establish function of
ventral nerve-dorsal nerve reflex bullbullbull 59
1 Ventral nerve-dorsal nerve reflex
initiated by the C02 sensitive
elements of the mid-sternal receptor bull 61
2 Effects of respiratory movement on
dorsal and ventral nerve afferent
activitybull 62
v
PAGE
3 Reflex responses to pressure
which activates the mid-sternal
receptor 64
4 Reflex responses to groundborne
vilIra tlons 64
III Responses Triggered by the DN-3B Receptors bullbullbullbull 66
A Dorsal nerve to homolateral ventral
nerve pa thrray 66
B Dorsal nerve rebound 68
C Dorsal nerve to heterolateral dorsal nerve
pa th-ray bull bull bull bull bull bull bull bull 71
D Experiments to establish function of
dorsal nerve to ventral nerve reflex bullbullbull 74
IV Stability of Reflexes 80
V Further Interganglionic Pa thrays bullbullbullbullbullbull 82
A Spread of ventral nerve input bullbullbullbull 84
B Spread of dorsal nerve input bullbullbull 84
C Several generalizations about the
dissemination of sensory information bullbull 85
VI EVidence for Proprioceptive Reflex System
Behavioral Observations bullbullbullbullbull 97
DISCUSSION 99
I Nid-sternal Receptor 99
II Lateral Chordotonal Organs 100
III Functions of the Mid-sternal Receptor and
Lateral Choraotonal Organ bullbullbullbullbullbullbullbullbull 101
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
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VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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ure
1
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Rhy
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N-2
[1J
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
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-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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1---middot_--c ---J)rVo~
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
~)( Cf~r~lt (1 lmiddotmiddotmiddot p r -Itoshy ~) i tmiddott~r~~J _~ - ~~ V~~k-gt n l-
to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
I $Fi C shy
rebound Rnd simulteneouG neXv~e stimulat1on
VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
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tor
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rsal
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ower
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ure
47
D
ors
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
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B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
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Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
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Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
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tera
l (u
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er tr
ace)
and
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late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
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stim
ula
te
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ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
iii
TABLE OF CONTBNTS
PAGE
INTRODUCTIon bullbull HATERIALS AND HETHODS 4
A Alimals bullbull middot 4
B Ana tomical bull 4
C Physiological 6
RESULTS 10
I Anatomical and Physiological Description of
the Abdominal Nerves and Sense Organs 10
A Ventral nerves and sensory elements 14
VN-1 14
VN-2 middot 14
VN-3 middot 15
VN-3A and AN-3Aa 15
Anatomy and physiology of the mid-
sternal rec eptor bull 16
B Dorsal nerve and sensory elements 27
DN-1 middot 27
DN-2 middot 28
DN-3 bull middot 28
DN-3A middot 30
DN-3B middot 30
iv
PAGE
Physiology of the D~-3A and DN-3B
receptors including the lateral
chordotonal organ 33
DN-4A middot 39
DN-4B inclUding the cardiac segmental
nerve 41
Relationship of DN-4 to the heart rate bullbull 43
Lateral cardiac nerve 47
II Responses Triggered by the Ventral Receptors 47
A Ventral nerve to homolateral dorsal
nerve pathvray 49
B Ventral nerve to heterolateral dorsal
nerve pathiay 53
C Ventral nerve rebound pathway bullbullbullbullbull 53
D Repetitive response in the homolateral
connective 57
E Responses of the cardiac lateral nerve bull 57
F Experiments to establish function of
ventral nerve-dorsal nerve reflex bullbullbull 59
1 Ventral nerve-dorsal nerve reflex
initiated by the C02 sensitive
elements of the mid-sternal receptor bull 61
2 Effects of respiratory movement on
dorsal and ventral nerve afferent
activitybull 62
v
PAGE
3 Reflex responses to pressure
which activates the mid-sternal
receptor 64
4 Reflex responses to groundborne
vilIra tlons 64
III Responses Triggered by the DN-3B Receptors bullbullbullbull 66
A Dorsal nerve to homolateral ventral
nerve pa thrray 66
B Dorsal nerve rebound 68
C Dorsal nerve to heterolateral dorsal nerve
pa th-ray bull bull bull bull bull bull bull bull 71
D Experiments to establish function of
dorsal nerve to ventral nerve reflex bullbullbull 74
IV Stability of Reflexes 80
V Further Interganglionic Pa thrays bullbullbullbullbullbull 82
A Spread of ventral nerve input bullbullbullbull 84
B Spread of dorsal nerve input bullbullbull 84
C Several generalizations about the
dissemination of sensory information bullbull 85
VI EVidence for Proprioceptive Reflex System
Behavioral Observations bullbullbullbullbull 97
DISCUSSION 99
I Nid-sternal Receptor 99
II Lateral Chordotonal Organs 100
III Functions of the Mid-sternal Receptor and
Lateral Choraotonal Organ bullbullbullbullbullbullbullbullbull 101
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
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VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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ure
1
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Rhy
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N-2
[1J
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
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shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
bullt ~ r - Cmiddotlt r ~- --~ ~ - -J 8UOCGJ rO+OIi -Jl~lvtG0P lJli (i~-G~-c ~~IOl~) f~t[+~ L~UOO
81 Ul HUOQSGJ ~U2l2jJj3 ~j(1pi)GYJ- middotX81Jl1I [IltgtU TT3JIOP
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bull uo ~~t T~TnurIgt~s aAIBU T~3Jl-TTGA 0+ 2AX2U 11G~rOp 311+ r~o ()J1IOd~~QX KrGU11~r(i ~~~ GJnC1~
1---middot_--c ---J)rVo~
o~
51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
s 1J
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
~i-u~~ h0 tc~4r1t~r ormiddot ~r~~ vOl+r-1 ~-~v~ cnv1~~ ~+~J if 2-r -~ J~t~) 0 1t -bullbull c-_ 1-1 lt_ -~ bull il -C~
stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
~)( Cf~r~lt (1 lmiddotmiddotmiddot p r -Itoshy ~) i tmiddott~r~~J _~ - ~~ V~~k-gt n l-
to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
I $Fi C shy
rebound Rnd simulteneouG neXv~e stimulat1on
VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
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tor
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rsal
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ower
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ure
47
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ors
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
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oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
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Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
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70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
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l (u
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er tr
ace)
and
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late
ral
do
rsal
ner
ve
resp
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se
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tr
ace)
to n
atu
ral
stim
ula
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n o
f th
e D
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re
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ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
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tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
iv
PAGE
Physiology of the D~-3A and DN-3B
receptors including the lateral
chordotonal organ 33
DN-4A middot 39
DN-4B inclUding the cardiac segmental
nerve 41
Relationship of DN-4 to the heart rate bullbull 43
Lateral cardiac nerve 47
II Responses Triggered by the Ventral Receptors 47
A Ventral nerve to homolateral dorsal
nerve pathvray 49
B Ventral nerve to heterolateral dorsal
nerve pathiay 53
C Ventral nerve rebound pathway bullbullbullbullbull 53
D Repetitive response in the homolateral
connective 57
E Responses of the cardiac lateral nerve bull 57
F Experiments to establish function of
ventral nerve-dorsal nerve reflex bullbullbull 59
1 Ventral nerve-dorsal nerve reflex
initiated by the C02 sensitive
elements of the mid-sternal receptor bull 61
2 Effects of respiratory movement on
dorsal and ventral nerve afferent
activitybull 62
v
PAGE
3 Reflex responses to pressure
which activates the mid-sternal
receptor 64
4 Reflex responses to groundborne
vilIra tlons 64
III Responses Triggered by the DN-3B Receptors bullbullbullbull 66
A Dorsal nerve to homolateral ventral
nerve pa thrray 66
B Dorsal nerve rebound 68
C Dorsal nerve to heterolateral dorsal nerve
pa th-ray bull bull bull bull bull bull bull bull 71
D Experiments to establish function of
dorsal nerve to ventral nerve reflex bullbullbull 74
IV Stability of Reflexes 80
V Further Interganglionic Pa thrays bullbullbullbullbullbull 82
A Spread of ventral nerve input bullbullbullbull 84
B Spread of dorsal nerve input bullbullbull 84
C Several generalizations about the
dissemination of sensory information bullbull 85
VI EVidence for Proprioceptive Reflex System
Behavioral Observations bullbullbullbullbull 97
DISCUSSION 99
I Nid-sternal Receptor 99
II Lateral Chordotonal Organs 100
III Functions of the Mid-sternal Receptor and
Lateral Choraotonal Organ bullbullbullbullbullbullbullbullbull 101
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
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Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
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Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
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to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
4
6
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uced
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spo
nse
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om
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e m
id-s
tern
al
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tor
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ng
th
e
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rsal
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e
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ow i
nd
icate
s th
e
tim
e o
f C
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cati
on
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pp
er tr
ace
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tral
ner
ve
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h in
tact
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-ste
rnal
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tor
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ower
tr
ace
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tor
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y
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the
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rsal
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e
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tral
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ve
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ere
nt
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rd
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er tr
ace
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ure
47
D
ors
al a
nd
ven
tral
sen
sory
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emen
ts
resp
on
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g
to v
en
tila
tory
pum
ping
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emen
ts
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ts
ind
icate
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mp
ress
ion
0
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l
64shy
3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
0 ~--14~ ~~I~ I~t-yenNImiddoty~iJr bull j I l i I r 1
middot~middotmiddot1middotmiddotmiddot
I 1 It I f 1I bull l 1
A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
1 1 1 1 _ - I l II 1 1 I t t ~~ ~~~l~-iYfwIb~(I~IgtL1p~1
bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
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mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
cep
tors
-J
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stim
ula
te
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rfi~~l
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ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
IOHer trace
------- ----__--__--__------~---_-- -- -
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77
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r tr
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Fig
ure
6
4
Res
po
nse
s on
th
e ~omolateral
ven
tral
and
d
ors
al
nerv
es
to
pre
ssu
re
stim
ula
tio
n o
f th
e
do
rsal
recep
tors
A
rrow
in
dic
ate
s ap
pro
xim
ate
tim
e st
imu
lati
on
was
b
egu
n
-J
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79
I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
v
PAGE
3 Reflex responses to pressure
which activates the mid-sternal
receptor 64
4 Reflex responses to groundborne
vilIra tlons 64
III Responses Triggered by the DN-3B Receptors bullbullbullbull 66
A Dorsal nerve to homolateral ventral
nerve pa thrray 66
B Dorsal nerve rebound 68
C Dorsal nerve to heterolateral dorsal nerve
pa th-ray bull bull bull bull bull bull bull bull 71
D Experiments to establish function of
dorsal nerve to ventral nerve reflex bullbullbull 74
IV Stability of Reflexes 80
V Further Interganglionic Pa thrays bullbullbullbullbullbull 82
A Spread of ventral nerve input bullbullbullbull 84
B Spread of dorsal nerve input bullbullbull 84
C Several generalizations about the
dissemination of sensory information bullbull 85
VI EVidence for Proprioceptive Reflex System
Behavioral Observations bullbullbullbullbull 97
DISCUSSION 99
I Nid-sternal Receptor 99
II Lateral Chordotonal Organs 100
III Functions of the Mid-sternal Receptor and
Lateral Choraotonal Organ bullbullbullbullbullbullbullbullbull 101
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
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gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
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Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
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Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
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Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
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Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
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lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
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to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
4
6
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uced
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nse
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om
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tern
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tor
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ng
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e
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rsal
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e
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ow i
nd
icate
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e
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e o
f C
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on
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pp
er tr
ace
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tral
ner
ve
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h in
tact
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tor
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ower
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ace
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tor
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y
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rsal
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e
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tral
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ve
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ere
nt
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rd
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pp
er tr
ace
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ure
47
D
ors
al a
nd
ven
tral
sen
sory
el
emen
ts
resp
on
din
g
to v
en
tila
tory
pum
ping
mov
emen
ts
Do
ts
ind
icate
co
mp
ress
ion
0
V
l
64shy
3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
0 ~--14~ ~~I~ I~t-yenNImiddoty~iJr bull j I l i I r 1
middot~middotmiddot1middotmiddotmiddot
I 1 It I f 1I bull l 1
A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
1 1 1 1 _ - I l II 1 1 I t t ~~ ~~~l~-iYfwIb~(I~IgtL1p~1
bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
~~J I III ~ lIJtmiddotAil1rfmiddotUII~fitgtrOI~-r7middotfii(V~ilJ~IfjyenA~I~~I~MLli~~l1 r Ir Ii I 1 11 I r i-I IIi r rmiddot-~r-I~--r
Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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~1t11CC d
Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
~UL
gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
~rromiddotT r1~A dlt-l~-gt-(nf~ -r--gtlrn~~ + ~rnrCgtn Ion - -0)rJ -11 v j~tll Llt -J Jlt-- 11 l-JJ _l _ - c J ~
I I
I1lt01 ~ VolN- ~ ~ t l i tgtgt~ LtI~I 1 I IN I ~f -- jlI-middotf-lt~L~middotL_ laMh ~
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A B c
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A Response O~ ~~- B ResDonse CYl ni-2 C R ~~ c~
u v~C~ r~_llbull e)lJol-l-~J T
71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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ulate
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re
59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
ho
mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
cep
tors
-J
OL
stim
ula
te
~~I
I II
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rfi~~l
re
cor~
~
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Fig
ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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nse
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ors
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ner
ve
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ula
tio
n i
n t
he h
ete
rola
tera
l d
ors
al
n
erv
e
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teri
or
co
nn
ecti
ves
cu
t
so th
at
the
maj
or
resp
on
se is
lo
st
---
J U
1
76
Q
(2)
VN
(3)
Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
IOHer trace
------- ----__--__--__------~---_-- -- -
o
~ ~ 4(1~~) t 1 J ~~lj~~middotJ~~~~I~ Jl~a ~bullaJ bullbull~ ~~~ ~I -~~~ plt1C~tr) r~~__ ~iIl~_oli~C~ ~1~
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77
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wer
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ace
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pre
ssu
re
stim
ulu
s
() ~
roo
ord
-
up
po
r tr
aco
Fig
ure
6
4
Res
po
nse
s on
th
e ~omolateral
ven
tral
and
d
ors
al
nerv
es
to
pre
ssu
re
stim
ula
tio
n o
f th
e
do
rsal
recep
tors
A
rrow
in
dic
ate
s ap
pro
xim
ate
tim
e st
imu
lati
on
was
b
egu
n
-J
CD
79
I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
vi
PAGE
A Rhythmometer function 101
B Proprioceptor function bullbullbullbullbull 102
IV Sponatneously Active Fiber Responding to
Hultiple Sensory Input bullbullbullbullbullbull 103 V Neural Control of the Heart bullbullbull 104
SUnmiddot1ARY bullbullbullbullbullbullbullbullbullbullbull It bullbullbullbullbull It bullbullbullbullbullbull 106It bullbull It
LITERATURE CITED 109
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
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gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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ill
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ure
1
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Rhy
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on D
N-2
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
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-5 i
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shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
bullt ~ r - Cmiddotlt r ~- --~ ~ - -J 8UOCGJ rO+OIi -Jl~lvtG0P lJli (i~-G~-c ~~IOl~) f~t[+~ L~UOO
81 Ul HUOQSGJ ~U2l2jJj3 ~j(1pi)GYJ- middotX81Jl1I [IltgtU TT3JIOP
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bull uo ~~t T~TnurIgt~s aAIBU T~3Jl-TTGA 0+ 2AX2U 11G~rOp 311+ r~o ()J1IOd~~QX KrGU11~r(i ~~~ GJnC1~
1---middot_--c ---J)rVo~
o~
51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
~i-u~~ h0 tc~4r1t~r ormiddot ~r~~ vOl+r-1 ~-~v~ cnv1~~ ~+~J if 2-r -~ J~t~) 0 1t -bullbull c-_ 1-1 lt_ -~ bull il -C~
stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
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to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
4
6
CO2-
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tor
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th
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rsal
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Arr
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nd
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f C
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ve
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ven
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Fig
ure
47
D
ors
al a
nd
ven
tral
sen
sory
el
emen
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resp
on
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to v
en
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tory
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Do
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ion
0
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
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middot~middotmiddot1middotmiddotmiddot
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A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
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oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
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I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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Jco
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74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
vii
LIST OF FIGURES
FIGURE PAGE
1 (L) Diagram of the major branches of the dorsal
bullganglion of Blaberus 12 nerve and ventral nerve of the third abdominal
(B) Diagram shoHing the rela tlonship of mT-4 to the heart and the inner tergal muscles bullbullbullbull 13
2 Whole mount of the mid-st3rnal receptor in Blab erus 17
3 Whole mount of the mid-sternal receptor in Blaberus at a higher magnification than Fig 2 bullbullbull 17
4 Mid-sternal receptor in Periplaneta bullbullbullbullbullbullbullbull 18
5 Mid-sternal receptor in Periplaneta at a higher magnification than Fig 4 18
6 Cross section of the mid-sternal receptor bullbullbullbullbullbullbull 19
7 Oross section of the mid-sternal receptor at a higher magnification than Fig 6 showing the sensory and supporting cells nuclei 20
8 Cuticular attachment of the mid-sternal ree eptor 20
9 Cross section of VN-3Aa 22
10 Response of the mid-sternal receptor to groundborn e vibrations 22
11 Response of the mid-sternal receptor to compression of the ventral musculature bullbullbullbullbullbullbullbull
12 Mid-sternal receptor responding to groundborne vibrations and C02 24
13 Response of the mid-sternallt receptor to 100 CO2 applied as a gas bull 24
14 Spontaneous efferent activity on VN-3Aa bullbullbullbullbullbullbullbullbullbullbull 25
15 (A B and 0) Experiments designed to show that ventral nerve efferents activate a sensory response from the mid-sternal receptor bullbullbullbull 26
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
l1
WI
j rll~
I
i
~
II
1 I
ill
II
I
I
I
r
I I
11
r
~JI
~~~
~~~
~~
~
~~~w
~~
~--1
--~~~r~~ln~~~~l~rr-~~~hr1~~
Y1~
J~~~
~1
1I
Jr
--
-~IIIIiliillIl
II
I li
-~-
~l
I~
J
~
t 1
11
11
II
lJlI
IJJ
I
I I
L
I
I I
I
L I
11
11
lilj1
II
I
loo
Mfc
c
~_~
---
~--------
--_-----_-------~-
Fig
ure
1
6
Rhy
thm
ic acti
vit
y
on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
~ i
-5 i
-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
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JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
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Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
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Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
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t-1M15j
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59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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47
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
~UL
gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
~rromiddotT r1~A dlt-l~-gt-(nf~ -r--gtlrn~~ + ~rnrCgtn Ion - -0)rJ -11 v j~tll Llt -J Jlt-- 11 l-JJ _l _ - c J ~
I I
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A B c
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A Response O~ ~~- B ResDonse CYl ni-2 C R ~~ c~
u v~C~ r~_llbull e)lJol-l-~J T
71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
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tera
l (u
pp
er tr
ace)
and
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mo
late
ral
do
rsal
ner
ve
resp
on
se
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wer
tr
ace)
to n
atu
ral
stim
ula
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n o
f th
e D
N-3
re
cep
tors
-J
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stim
ula
te
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cor~
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Fig
ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
IOHer trace
------- ----__--__--__------~---_-- -- -
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77
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ors
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79
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1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
viii
FIGURE PAGE
16 Rhytlunlc activl ty on DN -2 29
17 Methylene blue whole mount showing the hairs on Blah~ in the vicinity innervated by DN-3 bull 31
18 Methylene blue whole mount of the pleural region in Blab~ showing the two major branches of DN-3 31
19 Methylene blue stained whole mount of the posterior pleural region of Blabet~~ showing the many fine branches of DN-3A and DN-3B bullbullbullbullbull 32
20 Sagittal section of the lateral chordotonal organ showing attachment one in the posterior inter-segmental membrane 34
21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment one bull 34
22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells bullbullbullbullbullbull 35
23 Lateral chordotonal receptor in relationship to the dorsal musculature 35
24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the splracl e 36
25 Afferent responSe on DN-3 to groundborne vibrations 38I
26 Response in the dorsal nerve to rubbing the sterni tes 38
27 Response in the dorsal nerve to light pressure over the lateral juncture between two tergites 38
28 Response in the dorsal nerve to an air stream directed at the lateral fold bullbull 38
29 Simultaneous recording from deafferented and deefferented dorsal nerve~ ventilatory rhythm initia ted by CO2 bullbullbullbullbull 40
30 Methylene blue whole mount of the longitudinal and vertical receptors bullbullbullbullbull 42
31 Afferent actiVity fron DIl-4A 42
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
l1
WI
j rll~
I
i
~
II
1 I
ill
II
I
I
I
r
I I
11
r
~JI
~~~
~~~
~~
~
~~~w
~~
~--1
--~~~r~~ln~~~~l~rr-~~~hr1~~
Y1~
J~~~
~1
1I
Jr
--
-~IIIIiliillIl
II
I li
-~-
~l
I~
J
~
t 1
11
11
II
lJlI
IJJ
I
I I
L
I
I I
I
L I
11
11
lilj1
II
I
loo
Mfc
c
~_~
---
~--------
--_-----_-------~-
Fig
ure
1
6
Rhy
thm
ic acti
vit
y
on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
~ i
-5 i
-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
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homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
47
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ors
al a
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sory
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emen
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on
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to v
en
tila
tory
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icate
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mp
ress
ion
0
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l
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
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middot~middotmiddot1middotmiddotmiddot
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bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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~1t11CC d
Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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A B c
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
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Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
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2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
iX
FIGURE PAGE
32 Response on DN-4B to stimulation of the Iateral nerve 44
33 Activity recorded from the lateral nerve shoving a spike l-ri th polarity opposite to most in the record 48
34 Long duration spikes on the lateral nerve bullbull 48
35 Primary response on the dorsal nerve to ventral nerve stimulation ~ 50
36 Data showing the delay in the ventral nerve to dorsal nerve reflex 50
37 Response on DN-l to ventral nerve stimulation 52
38 Response on DN-4 to ventral nerve stimulation bull 52
39 Response on the dorsal nerve to stimulation of the heterolateral dorsal nerve of the same gang1ion 55
40 Same as Fig 39 only the anterior connectives are cut 56
41 Ventral rebound and simultaneous dorsal nerve response to ventral nerve stimulation bullbullbull 58
42 Ventral rebound reflex showing in-going afferent and out-going efferent spikes bullbullbullbull 58
43 Repetitive response in the homolateral connective to ventral nerve stimulation bullbullbullbullbullbull 58
44 Lateral nerve response to ventral nerve stimulation 60
45 Changes in the heart rate during stimulation of two dorsal nerves 60
46 CO2 induced response from the mid-sternal receptor activating the dorsal nerve bull bullbullbullbullbullbullbull 63
47 Dorsal and ventral sensory elements responding to ventilatory elements 63
48 (A and E) Two examples of the ventral nerve to homolateral dorsal reflex induced by compreEsion in the ventral musculature bullbullbullbullbullbullbullbullbull 65
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
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VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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Fig
ure
1
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Rhy
thm
ic acti
vit
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on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
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-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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81 Ul HUOQSGJ ~U2l2jJj3 ~j(1pi)GYJ- middotX81Jl1I [IltgtU TT3JIOP
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1---middot_--c ---J)rVo~
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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ure
4
0
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Fig
3
9
exce
pt
co
nn
ecti
ves
cu
t an
teri
or
to g
an
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re
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ed
fro
m
(see
dia
gra
m)
~pward-directed
arr
ow
in
dic
ate
s b
egin
nin
g o
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tor
resp
on
se a
nd
dow
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d d
irecte
d a
rro
w i
nd
icate
s b
egin
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g
of
sin
gle
fi
ber
resp
on
se
in
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rsal
nerv
e
S =
sen
sory
in
pu
t M
=m
oto
r re
spo
nse
J
l ff
i
57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
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j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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09
61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
4
6
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uced
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nse
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om
th
e m
id-s
tern
al
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tor
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vati
ng
th
e
do
rsal
nerv
e
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ow i
nd
icate
s th
e
tim
e o
f C
02
ap
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cati
on
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pp
er tr
ace
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tral
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ve
wit
h in
tact
mid
-ste
rnal
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tor
L
ower
tr
ace
mo
tor
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vit
y
on
the
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rsal
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e
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tral
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ve
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ere
nt
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rd
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ace
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ure
47
D
ors
al a
nd
ven
tral
sen
sory
el
emen
ts
resp
on
din
g
to v
en
tila
tory
pum
ping
mov
emen
ts
Do
ts
ind
icate
co
mp
ress
ion
0
V
l
64shy
3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
0 ~--14~ ~~I~ I~t-yenNImiddoty~iJr bull j I l i I r 1
middot~middotmiddot1middotmiddotmiddot
I 1 It I f 1I bull l 1
A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
1 1 1 1 _ - I l II 1 1 I t t ~~ ~~~l~-iYfwIb~(I~IgtL1p~1
bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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~1t11CC d
Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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(reco
rd
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race
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er tr
ace
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stim
ulate
~O~
F
igu
re
59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
ho
mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
cep
tors
-J
OL
stim
ula
te
~~I
I II
I(J
rfi~~l
re
cor~
~
It j
11 I
IrI
1 0
l
Fig
ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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77
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Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
x
FIGURE PAGE
49 Response in the thoracic connectives between T-2 and T-3 to natural stimulation of the mid-eternal receptor of A-1 bullbullbullbullbullbull 65
50 Simultaneous recording of dorsal nerve afferent activity and ventral nerve efferent activity during stimulation by groundborne vibrations 67
51 Simultaneous records of sensory input on an intact ventral nerve and response on the posterior homolateral connective bullbullbullbullbullbull 67
52 Primary response on the main trunk of the ventral nerve to dorsal nerve stimulation bullbullbull 67
53 (A B and 0) Responses on branchee of the ventral nerve to dorsal nerve stimulation bullbullbullbullbull 69
54 Dorsal rebound and ventral nerve response to stimulation of the dorsal nerve bullbullbullbullbullbullbull 69
55 Dorsal rebound reflex recorded in the anterior connectie 70
56 (A B and 0) Dorsal rebound reflex recorded on the peripheral branches receiving efferent informa tion 70
57 Two records showing response of the lateral nerve to stimulation of the dorsal nerve bullbullbull 72
58 Same as Fig 57 but connection to ganelion intact 72
59 lieterolateral and homolateral dorsal nerve response to natural stimulation of the DN-3 ree eptors 73
60 Response to dorsal nerve stimulation on the cut heterolateral connective 73
61 Response to dorsal nerve stimulation in the heterolateral dorsal nerve with the anterior connective cut 75
62 Diagram emphasizing the dorsal nerve fiber which responds to multiple sensory inputs bullbullbullbullbullbull 76
63 (A and B) Two examples of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve bullbullbullbullbullbull77
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
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Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
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Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
~)( Cf~r~lt (1 lmiddotmiddotmiddot p r -Itoshy ~) i tmiddott~r~~J _~ - ~~ V~~k-gt n l-
to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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rebound Rnd simulteneouG neXv~e stimulat1on
VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
bull -~
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ure
4
6
CO2-
ind
uced
re
spo
nse
fr
om
th
e m
id-s
tern
al
recep
tor
acti
vati
ng
th
e
do
rsal
nerv
e
Arr
ow i
nd
icate
s th
e
tim
e o
f C
02
ap
pli
cati
on
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pp
er tr
ace
ven
tral
ner
ve
wit
h in
tact
mid
-ste
rnal
recep
tor
L
ower
tr
ace
mo
tor
acti
vit
y
on
the
do
rsal
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e
ven
tral
ner
ve
aff
ere
nt
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ors
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ere
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er
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rd
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co
rd
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pp
er tr
ace
Fig
ure
47
D
ors
al a
nd
ven
tral
sen
sory
el
emen
ts
resp
on
din
g
to v
en
tila
tory
pum
ping
mov
emen
ts
Do
ts
ind
icate
co
mp
ress
ion
0
V
l
64shy
3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
0 ~--14~ ~~I~ I~t-yenNImiddoty~iJr bull j I l i I r 1
middot~middotmiddot1middotmiddotmiddot
I 1 It I f 1I bull l 1
A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
1 1 1 1 _ - I l II 1 1 I t t ~~ ~~~l~-iYfwIb~(I~IgtL1p~1
bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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~1t11CC d
Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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ulate
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re
59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
ho
mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
cep
tors
-J
OL
stim
ula
te
~~I
I II
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rfi~~l
re
cor~
~
It j
11 I
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1 0
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Fig
ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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rd
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rd
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re 6
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po
nse
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ors
al
ner
ve
stim
ula
tio
n i
n t
he h
ete
rola
tera
l d
ors
al
n
erv
e
An
teri
or
co
nn
ecti
ves
cu
t
so th
at
the
maj
or
resp
on
se is
lo
st
---
J U
1
76
Q
(2)
VN
(3)
Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
IOHer trace
------- ----__--__--__------~---_-- -- -
o
~ ~ 4(1~~) t 1 J ~~lj~~middotJ~~~~I~ Jl~a ~bullaJ bullbull~ ~~~ ~I -~~~ plt1C~tr) r~~__ ~iIl~_oli~C~ ~1~
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77
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rd
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wer
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ace
~~
pre
ssu
re
stim
ulu
s
() ~
roo
ord
-
up
po
r tr
aco
Fig
ure
6
4
Res
po
nse
s on
th
e ~omolateral
ven
tral
and
d
ors
al
nerv
es
to
pre
ssu
re
stim
ula
tio
n o
f th
e
do
rsal
recep
tors
A
rrow
in
dic
ate
s ap
pro
xim
ate
tim
e st
imu
lati
on
was
b
egu
n
-J
CD
79
I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
xi
FIGURE PAGE
6f Response nerves to
on p
the homolateral ventrressure Etimulation of
al and dorsal the dorsal
receptors - 78
65 Afferent response to groundborne vibrations on the dorsal nerve activating an efferent response on the ventral nerve 79
66 A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex 81
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
l1
WI
j rll~
I
i
~
II
1 I
ill
II
I
I
I
r
I I
11
r
~JI
~~~
~~~
~~
~
~~~w
~~
~--1
--~~~r~~ln~~~~l~rr-~~~hr1~~
Y1~
J~~~
~1
1I
Jr
--
-~IIIIiliillIl
II
I li
-~-
~l
I~
J
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t 1
11
11
II
lJlI
IJJ
I
I I
L
I
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11
11
lilj1
II
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loo
Mfc
c
~_~
---
~--------
--_-----_-------~-
Fig
ure
1
6
Rhy
thm
ic acti
vit
y
on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
~ i
-5 i
-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
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JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
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Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
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lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
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t-1M15j
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59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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47
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
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I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
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ve
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60
R
esp
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se
to
DN st
imu
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on
on
the
cu
t h
ete
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tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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77
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79
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lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
xii
LIST OF TABLES
I
II
III
Typical experiments showing the effect of C02 on the cockroach heart rate
Interganglionic pathways Responses to ventral nerve stimulation
Interganglionic pathways Responses to dorsal nerve stimulation
PAGES
45-46
86-89
90-g4
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
l1
WI
j rll~
I
i
~
II
1 I
ill
II
I
I
I
r
I I
11
r
~JI
~~~
~~~
~~
~
~~~w
~~
~--1
--~~~r~~ln~~~~l~rr-~~~hr1~~
Y1~
J~~~
~1
1I
Jr
--
-~IIIIiliillIl
II
I li
-~-
~l
I~
J
~
t 1
11
11
II
lJlI
IJJ
I
I I
L
I
I I
I
L I
11
11
lilj1
II
I
loo
Mfc
c
~_~
---
~--------
--_-----_-------~-
Fig
ure
1
6
Rhy
thm
ic acti
vit
y
on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
~ i
-5 i
-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
bullt ~ r - Cmiddotlt r ~- --~ ~ - -J 8UOCGJ rO+OIi -Jl~lvtG0P lJli (i~-G~-c ~~IOl~) f~t[+~ L~UOO
81 Ul HUOQSGJ ~U2l2jJj3 ~j(1pi)GYJ- middotX81Jl1I [IltgtU TT3JIOP
0+ Gil-lGU TBJ+ UGA 2l1 Tr~ j~q~p h~T ~Ull)Oi~S Tq B bull lt( 3Trl1pound
bull uo ~~t T~TnurIgt~s aAIBU T~3Jl-TTGA 0+ 2AX2U 11G~rOp 311+ r~o ()J1IOd~~QX KrGU11~r(i ~~~ GJnC1~
1---middot_--c ---J)rVo~
o~
51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
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t-1M15j
~ 5 volts
25 volts
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Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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47
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
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record upper trace - motor~ 0~
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
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Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
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70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
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74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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77
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Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
INTRODUCTION
Sherringtons studies of mammalian spinal reflexes
~Sherrington 1906) provided a detailed basis for Eccles r tudies of integration at the single cell leve~ (Eccles
Granit 1967) There 1s not yet a corresponding
body of background information about insect reflexes
in spite of the recognition (Roeder 1967) that insects
carry out quite complex coordinations with compared to
vertebrates surprisingly few nerve cells This may mean
that neural integrative processes are fundamentally
different in insects or that the processes are simpler and
should be easiar to analyze In either case insect central
nervous systems are worthy objects of comparative study
Hughes (1965) has warned that an understanding of central
integration in cellular terms may be delayed by the
present shortage of information about reflex pathways
The present study contributes to knowledge about
insect reflex pathways specifically those mediated by the
six abdominal ganglia of the cockroach Blaberus craniifer
The research is based on the following questions which
are directed towards a more complete understanding of the
physiology of the abdominal nerve cord
A Where are the abdominal receptors located and what are
their functions
B What are the intraganglionic reflex pathways and
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
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Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
bullt ~ r - Cmiddotlt r ~- --~ ~ - -J 8UOCGJ rO+OIi -Jl~lvtG0P lJli (i~-G~-c ~~IOl~) f~t[+~ L~UOO
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1---middot_--c ---J)rVo~
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
~)( Cf~r~lt (1 lmiddotmiddotmiddot p r -Itoshy ~) i tmiddott~r~~J _~ - ~~ V~~k-gt n l-
to v2ntrsl nerve Gtlm~JJ_Etion
~iv_ _ ventra~ nerve
j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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rebound Rnd simulteneouG neXv~e stimulat1on
VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
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om
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tor
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rsal
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ow i
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icate
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e
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h in
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tor
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ower
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ace
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rsal
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ure
47
D
ors
al a
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en
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ion
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
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middot~middotmiddot1middotmiddotmiddot
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B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
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Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
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I
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Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
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)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
ete
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l (u
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er tr
ace)
and
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late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
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stim
ula
te
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ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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rsal
recep
tors
A
rrow
in
dic
ate
s ap
pro
xim
ate
tim
e st
imu
lati
on
was
b
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-J
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79
I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
2
effector organs served by these sensory receptors
C What are the interganglionic reflex pathways which
could possibly serve to relay sensory information to
distant abdominal effectors
In cockroaches several abdominal receptrs have
been reported which have a bearing on the present study
Finlayson and Lowenstein (1958) found a single bipolar cell with
stretch receptor function located longitudinally in the
dorsal musculature of Periplaneta A later study by Osborne
and Lowenstein (1962) described a similar single celled
stretch receptor vertically arranged in the dorsal
musculature Shankland (1966) recorded a stretch receptor
response in a small nerve near the mid-ventral line in
Periplaneta A peripheral receptor in the vicinity of the
pleural fold has been found by Florentine (1967) to respond
to airborne sound Farley Case and Roeder (1967) have
reported unidentified abdominal receptors which fire
during inspiration and expiration in Periplaneta moreover
Farley and Case (1968) have shown that this sensory
information is capable of altering the frequency of the
respiratory rhythm
Relatively little work has been done on either
intraganglionic or interganglionic reflex pathways in
cockroach abdomens with the exception of the following
two studies on Blaberus Hughes and Wilson (1965) found
sensory fibers which enter one abdominal ganglion and make
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
l1
WI
j rll~
I
i
~
II
1 I
ill
II
I
I
I
r
I I
11
r
~JI
~~~
~~~
~~
~
~~~w
~~
~--1
--~~~r~~ln~~~~l~rr-~~~hr1~~
Y1~
J~~~
~1
1I
Jr
--
-~IIIIiliillIl
II
I li
-~-
~l
I~
J
~
t 1
11
11
II
lJlI
IJJ
I
I I
L
I
I I
I
L I
11
11
lilj1
II
I
loo
Mfc
c
~_~
---
~--------
--_-----_-------~-
Fig
ure
1
6
Rhy
thm
ic acti
vit
y
on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
I i
l
~ i
-5 i
-
shy
Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
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11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
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Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
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Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
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Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
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58
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t-1M15j
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59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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47
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3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
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Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
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f i I I ~I I I 1 1~vf- ~~V1 0 ~rh_-~Io ilo-~l1 -I~IVii I j I P 11 1 1 I 111 I l ~ll I J rflll I I j I i I ) I
~1t11CC d
Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
~UL
gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
~rromiddotT r1~A dlt-l~-gt-(nf~ -r--gtlrn~~ + ~rnrCgtn Ion - -0)rJ -11 v j~tll Llt -J Jlt-- 11 l-JJ _l _ - c J ~
I I
I1lt01 ~ VolN- ~ ~ t l i tgtgt~ LtI~I 1 I IN I ~f -- jlI-middotf-lt~L~middotL_ laMh ~
I i -[ II ~-rlfimiddot ~--~
A B c
- - 1Figure ) n01321 netve rC~gtYUH1 ri~ILf)X on tHeperlpoera branchcB rccsivi-~~ Effer()nt Infor~~tton-l-
A Response O~ ~~- B ResDonse CYl ni-2 C R ~~ c~
u v~C~ r~_llbull e)lJol-l-~J T
71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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ulate
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59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
ho
mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
cep
tors
-J
OL
stim
ula
te
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rfi~~l
re
cor~
~
It j
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Fig
ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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---
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76
Q
(2)
VN
(3)
Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
IOHer trace
------- ----__--__--__------~---_-- -- -
o
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77
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e ~omolateral
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and
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ors
al
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es
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79
I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
3
synaptic contacts in the next anterior ganglion The
resulting motor response returns to the nerve of the
ganglion stimulated Smalley (1963) in a study designed to
map respiratory reflex pathways established a number of the
basic reflexes studied in more detail here
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
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i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
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J II 1Abullbull fVIl i ( I i Ii I
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VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
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ure
1
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Rhy
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N-2
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30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
~ q til
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Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
_Jr ~---- bull
- iI -
---+
Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
I ~I 1111 I I I
~ rrlrtr(ri IAr~rtvV---lilt ~ fI ~
f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
~~ry-Jlr~~It+-w--w(~~~~~~~~~~-
J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
--kLaMsc Co
Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
e-J ~JJ ~~ )-~~~~~~
lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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1---middot_--c ---J)rVo~
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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ure
4
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Fig
3
9
exce
pt
co
nn
ecti
ves
cu
t an
teri
or
to g
an
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re
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ed
fro
m
(see
dia
gra
m)
~pward-directed
arr
ow
in
dic
ate
s b
egin
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g o
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tor
resp
on
se a
nd
dow
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d d
irecte
d a
rro
w i
nd
icate
s b
egin
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g
of
sin
gle
fi
ber
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on
se
in
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rsal
nerv
e
S =
sen
sory
in
pu
t M
=m
oto
r re
spo
nse
J
l ff
i
57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
~~~~r~~C~~ ~~~~ ~ ~~~ ~ ~ ~ C~~ ~ ~~~~r1~~ ~ l~~ ~~ 1~~~~~ 1 Er~~~middot~~~~te Sf)Ons e ~
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
1 1 dOl ~dJ-Pl __~
II I if
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j ~ j dOl~f(- 1 nr-Tve~ (-s r~ - t- ) ~
11f ~ I I
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VentlBl len tral
~i~11lmiddot~ 4~ ~~0~1~1~ra c i c J bull I~ - LoJ-- v ~~
homolatersl con~ective
Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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09
61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ure
4
6
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uced
re
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nse
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om
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e m
id-s
tern
al
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tor
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vati
ng
th
e
do
rsal
nerv
e
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ow i
nd
icate
s th
e
tim
e o
f C
02
ap
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cati
on
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pp
er tr
ace
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tral
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ve
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h in
tact
mid
-ste
rnal
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tor
L
ower
tr
ace
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tor
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y
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the
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rsal
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e
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tral
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ve
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ere
nt
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rd
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ace
Fig
ure
47
D
ors
al a
nd
ven
tral
sen
sory
el
emen
ts
resp
on
din
g
to v
en
tila
tory
pum
ping
mov
emen
ts
Do
ts
ind
icate
co
mp
ress
ion
0
V
l
64shy
3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
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0 ~--14~ ~~I~ I~t-yenNImiddoty~iJr bull j I l i I r 1
middot~middotmiddot1middotmiddotmiddot
I 1 It I f 1I bull l 1
A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
1 1 1 1 _ - I l II 1 1 I t t ~~ ~~~l~-iYfwIb~(I~IgtL1p~1
bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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~1t11CC d
Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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(reco
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er tr
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stim
ulate
~O~
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igu
re
59
H
ete
rola
tera
l (u
pp
er tr
ace)
and
ho
mo
late
ral
do
rsal
ner
ve
resp
on
se
(lo
wer
tr
ace)
to n
atu
ral
stim
ula
tio
n o
f th
e D
N-3
re
cep
tors
-J
OL
stim
ula
te
~~I
I II
I(J
rfi~~l
re
cor~
~
It j
11 I
IrI
1 0
l
Fig
ure
60
R
esp
on
se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
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77
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Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
LITERATURE CITED
Alexandroricz JS 1926 The innervation of the heart of the cockroach J Compo Neurol 41 291-310
Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
Haskell PT 1956 Hearing in certain Orthoptera I physiology of sound receptors J Exp Biology 33 756-766
Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
Roeder KD 1967 Nerve cells and Insect Behavior Harvard University Press Cambridge Massachusetts
Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
Shankland DL 1965 Nerves and muscles of the pregenital abdominal segments of the American cockroach Periplaneta J Morphol 117 353-385
Sherrington CS 1906 The Integrative Action of the
in the cockroach Blaberus craniifer PhD Thesis
nervous system Yale University Press New Haven
Smalley KN 1968 Personal commun~cation
Smalley KN 1963 The neural regulation of respiration
State University of Iowa University Microfilms Ann Arbor Mich (Diss Abstr 638036)
Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
Weevers RDeG 1966 The physiology of a lepidopteran muscle receptor III The stretch reflex J Exp BioI 45 229-249
Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
MATERIALS AND METHODS
I ANIMALS
Adult male cockroaches Blaberus craniifer
Burmeister were used for most of this investigation The
larger size of the females made them useful in heart lateral
nerve studies but the fat bodies and egg cases interfered
with ventral nerve cord experiments Male adult
Periplaneta americana were used in comparative histological
studies of abdominal sense organs Male and female
Periplaneta americana nymphs were utilized in heart rate
experiments because the heart is visible through the
dorsal cuticle
The cockroaches were kept in 8 by 20 by 10 steel
cages Each cage contained several hundred male and female
cockroaches of various ages The animals were fed apples
Gaines dog food and oatmeal
II ANATOlHeAL
To trace fine nerves to their sensory endings
02 ml of reduced methylene blue was injected into the
abdominal cavity For temporary preparations a saturated
ammonium molybdate solution was applied to stop the staining
process after thirty minutes If permanent whole mounts
of sense organs were required methylene blue was injected
5
for twenty to thirty minutes followed by ammonium picrate
for five minutes and ammonium molybdate for twelve hours
The preparation was then washed in distilled water given
two changes of tertiary butyl alcohol in two hours cleared
in xylene for thirty minutes and mounted on aslide
(see Stark Smalley and Rowe 1968)
For histological studies of the peripheral sense
organs serial sections were reQuired Newly moltGd
Blaberus craniifer were fixed in alcoholic Bouins A
sink aspirator was used to remove air in the tracheal
system Each successive stage of the standard alcoholic
dehydration was also done under reduced pressure Segments
from the periphery of the abdomen were removed and embedded
in paraffin Sagittal frontal and cross sections were
cut at five to ten microns Several series of slides
were stained with Luxol fast blue and phosphotungstic acid
while the best results were with Mallorys triple stain
Photomicrographs of interesting whole mounts were
taken with an American Optical Microscope with a Polaroid
attachment A Wratten 80A filter was used with outdoor
color Polaroid film A Wild camera and dissecting
microscope were used with Polaroid black and white film to
take pictures of the distribution of the dorsal nerve
branches to the pleural fold
6
III PHYSIOLOGICAL
Prior to dissection all experimental animals were
anesthetized with CO2 Most animals were opened dorsally
pinned to a balsa block and the gut and tergites were
removed to expose the abdominal nerve cord When it
was necessary to eliminate extraneous activity two adjacent
ganglia their peripheral nerves and the connectives
joining them were isolated from the rest of the nerve cord
be severing the connectives anterior and posteior to the
pair of ganglia
In experiments on the lateral nerves of the heart an
animal was pinned to a waxed petri dish one lateral
incision was made the gut was removed and the tergites were
folded to the side and pinned down exposing the abdominal
portion of the heart Once the ventral cord or the heart
was exposed it was perfused periodically with saline
(Yamasaki and Narahashi 1959)
Fine silver wires (0005 in) served as recording and
stimulating electrod2s and were positioned by manipulators
The recording electrodes were connected through a
Tektronix 122 low level preamplifier to a two-channel
Tektronix 502 oscilloscope Nerve activity was photographed
with Grass C4 and Polaroid cameras A Grass AM5 audioshy
amplifier was used to supplement the oscilloscope in
monitoring nerve activity Stimulus pUlses were delivered
from a Grass s4 stimulator through a Stimulus Isolation
7
Unit
In several instances when stimulating or recording
from very small nerves it was necessary to use even smaller
wire electrodes The wire was bent into the shape desired
and sharpened electrolytically Occasionally ~he nerve to
be stimulated or recorded from was coated with Vaseline to
prevent it from drying
In the usual recording procedure a nerve was lifted
out of the saline with a small glass hook onto one recording
electrode and the second recordin~ electrode was placed in a
drop of saline in contact with the animal A nerve to be
stimulated was placed on a pair of stimulating electrodes
and lifted into the air In most experiments the stimulus
intensity varied between 15 and 60 volts and the duration
was held constant at 10 msec In many experiments the
thresholds increased as the preparations aged This was
compensated for by increasing the stimulus intensity When
working with rebound reflexes (in which sensory impulses
enter and motor impulses leave the same nerve) the duration
had to be reduced to 01 msec to reduce the stimulus
artifact In reflexes involving more than one ganglion
connectives were cut systematically to further define
interesting pathways
Inherent in any electrical stimulating study is the
possibility that non-physiological pathways may be
activated This was compensated for by using low stimulus
8
intensities
Saline saturated with 100 CO2 was usually used in
experiments which called for initiation of the ventilatory
rhythm Less frequently 100 or 5 CO2 was appled directly
as a gas
In experiments designed to determine whether or not
a nerve terminates in a sensory structure two procedures
were used In many instances it was possible to cut the
branch supplying the receptor and to record from a point
distal to the cut In nerve branches too small to be
recorded from directly records were taken from the
deefferented main trunk
In order to eliminate the effects of antifromic
conduction resulting from electrical stimulation various
natural stimuli were applied to activate suspected but
quiescent sensory elements Groundborne vibrations
probing in the musculature pressure on the sternites
tergites and CO 2 were all capable of initiating a response
in one receptor or another
Heart rates were monitored with a transducer
described by Miller (1968) The main components of this
transducer are a battery-operated FM transmitter module
and an F~ tuner A light wire lever is placed in contact
with the heart The movable end of the lever is placed
near the antenna of the transmitter As the lever moves the
effective capacitance of the transmitter antenna is varied
9
and this causes the output poundrequency opound the transmitter to
vary Within the ~l tuner the change in poundrequency is
converted into a change in dc voltage proportional to
the movement opound the heart and this voltage is displayed on
an oscilloscope
RESULTS
I ANATOMICAL AND PHYSIOLOGICAL DESCRIPTION OF THE
ABDOMINAL NERVES AND S~NSE ORGANS
This section reports the structure funption and
locations of the abdominal sensory elements and provides a
physiological description of the peripheral nerves described
anatomically by Smalley (1963)
Fig 1 (A and B) sho~s the anatomical arrangement of
nerves in the abdomen of BlaberuG cranilfer The
designation of nerves and ganglia follows that of Smalley
(1963) modified only where necessary to include finer
branches than Tere described in that study The six
abdominal ganglia are designated A-1 through A-6 The more
anterior paired nerve in each ganglion is designated th~
dorsal nerve (DN) and the more posterior paired nerve is
designated the ventral nerve (VN) Branches of each of
these nerves are numbered in proximal to distal order The
smaller rami of these branches are designated by capital
letters and further divisions (only occasionally
necessary) are designated by lower case letters Thus
VN-3Aa is a small division off the first ramus of the third
branch of the ventral nerve
The numbers of fibers given in the succeeding sections
are based on the number of different spike heights recorded
11
DN-l
DN-2
DN-3
DN-4
VN-1
VN-2
VN-3
Destinations of the abdominal nerve branches
Dorsa Nerve
inner sternal muscle
spiracular interpleural and inner tergal muscles
DN-2A branch connecting DN-2 with VN-3 of the anterior segment
DN-3A and DN-3B several receptors associated with the lateral fold including the lateral chordotonal organ
DN-4A primarily sensory vertical and longitudinal single celled receptors
DN-4B inner tergal muscle lateral nerve of the heart
Ventral Nerve
terminates at the ventral body wall near the base of the tergo-sternal muscle
tergo-sternal muscle
VN-2A joins with DN-2 of the same segment
outer sternal muscle
VN-3A outer sternal muscle
VN-3Aa sensory mid-sternal receptor
Q~k ~e
12
ltI Nbull z
c
~
w
~ bullbull
0114 - ltI
c11 -shy A
~Z~ gtJ -YN -2 _ ~ B
VN-3Aa ----YN-3
A-3
Figure l-A Diagram of major branches of dorsal nerve (DN) and ventral nerve (VN) of the third abdominal ganglion (A-3) of Blaberus Minor variations in branching pattern and relative lengths of nerves exist on other ganglia Destinations of these nerves given on opposite page
1
13
heart
ON-41 segmen tal nerve
lateral nerve
Figure 1-B Diagram showing the relationship of DN-4 to the heart and to the inner tergal muscles
14
from nerves firing spontaneously or under the influence of
0deg2
A VENTRAL NERVES AND SENSORY ELE11ENTS
VN-1
VN-1 branches from ~~-2 and terminates in the
vicinity of the lateral body wal~ Two motor fibers are
present To test for sensory elements in VN-l recordings
were made from an isolated segment of VN-2 with only VN-l
functionally intact and the area surrounding the nerve
was sUbjected to sensory activating stimuli No sensory
elements could be demonstrated Because of this and the
difficulty of obtaining records from VN-l due to its small
size its responses to stimulation were not tested in the
ensuing reflex work
VN-2
VN-2 innervates the tergo-stfrnal muscle and
contains three large and one small motor fibers
Possibly one receptor cell contributes to this nerve
On one occasion a single afferent spike was found firing
spontaneously A careful histological study of the tergoshy
sternal muscle revealed no sensory elements among the
muscle fibers A small ramus branches from VN-2 to curve
around the tergo-sternal muscle Its terminations could
not be follored and may possibly include a sensory
15
structure
There is a small neural connection VN-2A between
VN-2 and DN-2 of the same ganglion Its anatomical
designation as a branch of ~1-2 is based on the following
physiological observations recordings from tqis nerve
detached from DN-2 showed spontaneously firing motor fibers
and when recordings were made from the same connection
detached froQ VN-2 the spontaneous activity stops In the
sUbsequent reflex study VN-2A was always cut to insure that
dorsal nerve stimulation did not directly activate these
ventral nerve fibers
bull VN-3
VN-3 innervates the outer sternal muscle and the
lateral intersegmental connective tissue Three motor
fibers are present and the nerve participates in the
ventilatory rhythm No sensory elements have been detected
histologically or electrophysiologically
VN-3A and VN-3Aa
VN-3A innervates the outer sternal muscle Distal
to the small sensory branch VN-3Aa which passes midshy
ventrally the nerve is entirely motor Three motor
fibers are found which participate in a respiratory rhythm
Methylene blue preparations show VN-3Aa passes
under the ventral nerve cord and terminates in five bipolar
16
sensory neurons (Fig 2 3) The presence of this receptor
was demonstrated electrophysiologically in ventral nerves
of ganglia A-l through A-6 in males and A-1 through A-5
in females It is also present in Periplaneta (Fig 4 5)
presumably contributing to the same ganglia asthe receptors
in Blaberu~ This receptor will be referred to as the midshy
sternal receptor
Simultaneous recordings from VN-2 VN-3 and VN-3A
indicated there are no bifurcating fibers
ANATOtIT AND PHYSIOLOGY OF THE MID-STEfu~AL RECEPTOR
Sections of this receptor stained with Mallorys
triple stain show that the distal ends of the paired
receptors of a single segment converge upon approximately
the same spot on the cuticle (Fig 6)
A thick layer of connective and fatty tissues
overlies the cell bodies These cell bodies are encapsulated
by a blue-staining tissue which appears to be connective
tissue or Schwann cell investment (Fig 7) The large
round nuclei of the receptor stained black with methylene
blue and red with Mallorys stain The distal ends of the
bipolar cell bodies are enmeshed in a thick strand of
connective tissue which in turn is attached to the cuticle
at the mid-ventral line (Fig 8) The nuclei of 20-30
non-neural supporting cells are found in close association
with the receptor cell bodies The connective tissue
17
Pigure 2 Whole mount of the mid-sternal receptor (arrow)in Blaberus VN-3Aa passes horizontally to join VN-3A larger nerve along left side of photomicrograph (50X)
Pigure 3 Whole mount of the mid-sternal receptor at a higher magnification One nucleus visible along lower border (225X)
18
Figure 4 Mid-sternal receptor (lo~g arrow) in Periplaneta Note the inner sternal muscle is found on either side of the receptor The distal end of the receptor is visible (short arrow) (50X)
Figure 5 Mid-sternal receptor in Periplaneta showing four of the five nuclei of suspected receptor cells One nucleus 11es over another at the ventral (lower) end of the receptor (225X)
19
Figure 6 Cross section of the mid-sternal receptor Both the cell bodies and point of attachment of the one receptor (short arrow) can be seen but only the distal point of attachment (blue stainin~ area long arrow) of the other receptor is visible (50X)
20
Figure 7 Cross section of the mid-sternal receptor Note the supporting cells (short arrow) and surrounding fatty tissue (long arrow) (225X) bull
bull 1_ r
~-
r
bull
Figure 8 Cuticular attachment of the mid-sternal receptor Note the large supporting cells in the cuticular epidermis (short arrow) and the connective tissue strands ~assing through the epidermis to the cuticle (long arrow) Fig 7 and 8 are higher magnificztion photographs of the same section as shom in Fig 6 (225X)
21
strands stain red with Mallorys stain and contrast with the
blue-stsined nerve cells
The majority or the connective tissue strands
terminate at the epidermal layer of the cuticle Several
central strands pass through the epidermal cel~ layer to
fuse with what appears to be the laminated endocuticle
In the epidermis tuo large (trichogen tormosen) cells are
located around the strands embedded in the cuticle
The cuticle in the vicinity of the receptor is
stained blue in contrast with the rest of the cuticle
which is stained red with Mallorys Florentine (1967)
noticed similar staining differences in the vicinity of
peripheral sensory structures and attributed them to local
differences in cuticular permeabilities
Cross sections of VN-3Aa show that at least seven
axons are present (Fig 9) Electrophysiologically five
of these axons can be sho~ to be sensory and two middotmore can
be shown to be efferent
The five sensory cells were surprising in the range
of stimuli to which the group responded and the degree or
overlap in function between units Two fibers responded
with a discrete burst or activity to groundborne
vibrations set up by tapping on the floor of the recording
cage (Fig 10) The smaller fiber responded to a single
stimulus with a burst of 10-25 spikes while the larger
fiber fired once or twice Two to five ribers fired
phasically in response to compression in the ventral
22
cr~
i ~
i
1
fV
~ ~ ii1 - t ~
Figure 9 Cross section of VN-3Aa (arrow) At least seven axons are present (500X)
(00 tv ~er I
j111 Ii I f I I ~ I l I ~~~llrlr~r-T~t-~~rtl~middotbullbull~tbullJr_v~middot~
1
Figure 10 Response of mid-sternal receptor to groundborne vibrations
23
musculature (Fig 11) One fiber firad spontaneously in 90
of the preparations CO2 saturated solution evoked a
response from three fibers Tapping stimulation under these
conditions also initiated a response from two fibers
(Fig 12) Exposure to CO2 gas stimulated allof the fibers
(Fig 13) In a receptor stimulated by 100 CO2 gas it
was impossible to initiate a further response to groundborne
vibrations Insect saline adjusted to pH 4 or 45 with HCl
simulated the effects of CO2 gas on the receptor
Two efferent fibers were occasionally found firing
spontaneously on VN-3Aa (Fig 14) These efferents were
shown to activate the receptor by the following experiment
(see diagram in Fig 15) The ventral nerve was cut near
the ganglion and a pair of stimulating electrodes was
placed distal to the cut All peripheral connections
except VN-3Aa were cut A recording electrode was placed
between the mid-sternal receptor and the stimulating
electrodes On stimulation two fibers (A in Fig 15)
responded at a latency of 1 msec This was followed by a
highly repeatable burst of one or two fibers responding
eight to thirty times per stimulus (B in Fig 15) Two
facts strongly suggest that response A is efferent and
B afferent response A remains but B drops out after
cutting VN-3Aa at the receptor The polarity of response
An is opposite to that of response liB (C in Fig 15)
Obviously muscle tissue intermeshed with the
24
JoILMkL
O-___gt-~--I~ll~I~ ~ l iI ~ 1i~flil~ itr1fI~t ii I IiiI bull -----~~-
Figure 11 Response of mid-sternal receptor to compression of the ventral musculature
bull I
I I
~~-~-JJ_JWV ~~j~J~rtWtJ)Jll roo I
100 11fecI
Figure 12 Smaller spikes responding to CO2 in solution larger responding to groundborne Vibrations
bull 1 I ~ ~ ~ 1 1 bull ~ I Jl I I I bull I I I I I I I 1 ~ bull-I~ tI ~ r-~ ~Iol
I Ii j I I I II Iii I I i I I I I
COMftc
Figure 13 Response of mid-sternal receptor to 100 CO2applied as a gas
25
I I ~~~~
I (Of) amp1 r
Figure 14 Spontaneous efferent activity on VN-3Aa Nerve destroyed distal to recording electrode
26
i
I 115 _ ~
A
ilJ 1~J1 0101-0 I ~I rI
J II 1Abullbull fVIl i ( I i Ii I
J) (1-_-Irvi$~~ - - -~
~G~
gt Ji _11 t) ~ II ti -fti Stilliula te 1Aj~1 j~r~llt-f I ~ 1iltltV~I IN -shy11 1 1 i I Record
VNmiddotmiddot3Aa B
mid-sternal receptor t-Ce- f_ _0
-~ shy----~
cmiddot
Figure 15 Expar12ents dzslz~el to show thfl t 7n1 tt2 1 nerve D=-o middotAi~~rl
1~)efferents acmiddottiv~t2 th0 sensory re8ponse~ ~t_ -- J- condi tion3 FtS shO~trn il diflrJ
A 3fferent r28po~lse Oiy
B rrwo examp183 of the Dffer~nt reizponse C Records tsiell at t-ro diffr~(lt 3G(~2d~) to erJphf3ize th3
efr~1t middotrn~--middoton~o t~I ~ ~ +h(~ middot-fltfmiddot~-ltl+middot l~lt-lnmiddot-middotmiddotmiddotgt 11~l~- ~~- _ ~ _~~lt _ -~ _ ---_ ~J~ ~ -L- ~gt ~dL ~
respectively Not~ the rsversal of polarity
27
connective tissue strands would explain the observed
response to efferent activity but cross sections as thin
as 5-7 microns fail to confirm the presence of nmscle cells
A more definite statement on the question of whether or
not there are muscle fibers in this receptor must await
an electron microscope study
Because the distal ends of both ventral receptors
from a single ganglion lie in close proximity to one
another it was important to determine whether the activity
in one receptor affected the other Simultaneous recordings
from both receptors during stimulation by tapping showed
each receptor responded independently in terms of latencies
number of fibers participating and the intensity of the
tapping required to initiate a response Slight compression
in the ventral musculature of one side of the animal caused
only the homolateral receptor to respond Electrical
stimulation of the efferent input of one ventral receptor
produced no response in the opposite receptor
B DORSAL NERVES AND SENSORY ELEMENTS
DN-l
DN-l innervates the inner sternal muscle and consists
of three large and one small motor fibers which participate
in the respiratory rhythm Stimulation of DN-l results in a
visible contraction of the muscle There are no sensory
elements associated with this nerve as determined by
28
electrophysiological analysis and confirmed histologically
DN-2
DN-2 innervates the inner pleural inner tergal and
spiracular muscles It contains at least five small motor
fibers which fire spontaneously and one large fiber Which
fires in bursts of 10-25 spikes per burst The frequency of
bursts is highly variable The bursts may occur every few
milliseconds or may be separated by intervals up to 60
seconds (Fig 16) Occasionally the bursting does not
occur When present the spontaneous activity can be
eliminated by cutting any connective anterior to the
ganglion from which the recording is being made This
rhythmic activity appears to coincide with the normal
respiratory rhythm Which involves the rest of the motor
fibers of DN-2 when the animal is perfused with CO2 in
solution
No sensory elements could be detected on this nerve
by electrophysiological or ~istological techniques
DN-3
Smalley (1963) described DN-3 as innervating the
dorsal body wall in the region of the pleural membrane
Electrophysiological analysis indicates this nerve is
primarily sensory and the histological analysis concurs
In Periplaneta and Blaberus many hairs are found on
Il~
l1
WI
j rll~
I
i
~
II
1 I
ill
II
I
I
I
r
I I
11
r
~JI
~~~
~~~
~~
~
~~~w
~~
~--1
--~~~r~~ln~~~~l~rr-~~~hr1~~
Y1~
J~~~
~1
1I
Jr
--
-~IIIIiliillIl
II
I li
-~-
~l
I~
J
~
t 1
11
11
II
lJlI
IJJ
I
I I
L
I
I I
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L I
11
11
lilj1
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I
loo
Mfc
c
~_~
---
~--------
--_-----_-------~-
Fig
ure
1
6
Rhy
thm
ic acti
vit
y
on D
N-2
[1J
()
30
the abdomen in regions innervated by DN-3 Wigglesworth
(1950) states Nearly 811 the spines and hairs on the body
surface (of an insect) are sensory end organs This is
certainly not true of all hairs on the abdomen in ~laberus
and Periplaneta In a series of methylene blu~ whole
mounts (Fig 17) and serial cross sections no neural
elements were found associated with the hairs on the
dorsolateral cuticle
Several millimeters from the pleural fold the nerve
divides and the A branch receives contributions from
20 to 30 small neurons originating in the pleural fold
The B branch originates from the wall of the cuticle
adjacent to the spiracle (Fig 18)
DN-3A
It is difficult to ascribe a function to the small
nerves contributing to DN-3A although electrophysiological
analysis of DN-3 indicates most of them are sensory Whole
mounts of the pleural fold region show small darkened areas
possibly nuclei of receptor neurons in several of the
branches (Fig 19) Unfortunately serial sections have
proven useless in attempts to define further these small
nerve branches
DN-3B
DN-3B has similar small nerve cells associated with
31
i ~l bull I ~
Figure 17 Methylene blue whole mount showing hairs on the abdomen of Elaberus in the vicinity innervated by DN-3 (50X)
Figure 18 Methylene blue whole mount of the pleural reGion in Blaberus showing the two major branches of DN-3 (225XJ A - DN-3A B DN-3B
32
~ IIf t~t
bull - bull
Figure 19 Methylene blue stained whole mount of the posterior pleural region of Blaberus showing the many fine branches of DN-3A and DN-3B (SOX)
33
the cuticle and ~n important dorsal receptor a heretofore
undescribed chordotonal org~n rhich was best seen in
sagittal sections
The receptor which will be referred to as the
lateral chordotonal org~n h~s two cuticular attachments
The first (to be called attachment 1) attaches to the
posterior margin of the segment (Fig 20 21) It is
located dorsal and medial to the line of fusion between the
sternite and tergite This attachment appears to be fanshy
shaped in three dimensions and to consist of connective
tissue fibers which stain blue with Luxol Fast Blue The
fibers pass anteriorly to the cell bodies which stain red
with Mallorys stain (Fig 22 23) The sensory axons leave
the cell bodies pass anteriorly and dorsally and
eventually contribute to DN-3B The cell bodies are
encapsulated with blue-staining connective tissue Several
nuclei from supporting cells are associated with the
sensory cell bodies The second more anterior attachment
is an extension of the line formed by attachment 1 and cell
bodies and is located at the lateral fusion line
Fig 24 shows the relationship of the receptor to
the dorsal and ventral cuticle
PHYSIOLOGY OF THE DN-3 RECEPTORS INCLUDING
THE LATERAL CHORDOTONAL ORGAN
Since a minimum of ~O sensory cells contribute to
34
Figure 20 Sagittal section of the lateral chordotonal organ showing attachment 1 in the posterior intersegmental membrane (arrow) just medial to the lateral fold (50X)
J -~~~ gt~ If
_ bull ltt ~ bull amiddot ~Ill7lt- -c 1amp fIT bull ~
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Figure 21 Same as Fig 20 but at a greater magnification to show fan-shaped attachment 1 (225X)
35
bull - shy- -
shybull I iii ~ ~
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- iI -
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Figure 22 Encapsulated lateral chordotonal receptor showing nuclei of five sensory cells (225X)
Figure 23 Lateral chordotonal receptor (arrow) in relationship to the dorsal musculature (50X)
Ventral
36
Posterior
spiracle
Dorsal
lateral chordotonal organ
attachment
Anterior attachment
Figure 24 Diagram showing the relationship of the lateral chordotonal organ to the cuticle around the spiracle
37
DN-~ and different stimuli elicit characteristic and
repeatable response~ it is obvious that the lateral
chordotonal organ is not the only source of sensory
information on this nerve
Three and occasionally four fibers responded to
groundborne vibrations (Fig 25) Coating the lateral fold
with Vaseline a method which has been used to inhibit
sensory hair responses (Haskell 1956) failed to halt the
sensory response to tapping
Gentle rubbing of the ventral surface in the
Vicinity of the lateral fold initiated a sensory response
of 4-6 fibers (Fig 26) Several of these same fibers
seemed to respond to compression of the tergites
Uniform light pressure over the lateral junction
between two tergites initiated a pronollilced response of five
to seven fibers (Fig 27) This is the area which overlies
the lateral chordotonal organ Stronger compression on the
tergites stimulated up to twelve fibers
An air stream directed at the pleural fold initiated
a response in up to 12-15 fibers (Fig 28) Vaseline
applied to the pleural fold and vicinity did not affect
these responses The air stream presumably exerts its
influence through mechanical deformation and comes as close
to activating all of the sensory elements as any stimulus
Receptors in DN-3 responded to ventilatory movements
Three to five fibers respond 30 to 60 msec after the
inspiratory motor bursts and one during expiration
38
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f IP tASe (
Figure 25 Afferent response on DN-3 to groundborne vibrations
T I I bull 1 I r
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J---l~~
Figure 26 Response in the dorsal nerve to rubbing sternites
~~I J I I
I~OMS~~I
Figure 27 Response in the dorsal nerve to light pressure over the lateral junction between two tergites
I I I j I I I bull 1 t I ~ J I I IIll~ ~l ~_ I ~ 1 hJI 1 11 o~ bullbull ~ I -LI_~
MI~i~ Inmiddotmiddotl1Y-lwmiddot rlt (jlrrI uttJi II (II t fi~1 bull1n1t~ -(yyl ~l r1 ~JIf I I I I I II I r L bull ( I I I I II II
bull I
~~
Figure 28 Response in the dorsal nerve to an air stream directed at the lateral fold
39
(Fig 29) A similar cell firing during expiration in
Periplaneisect was reported by Farley Ca se and Roeder (1967)
although its anatomical location was not knom
The sensory elements of DN-3 were tested for
susceptibility to various chemicals in an atte~pt to
activate a quiescent chemo-receptor such as the possible
CO2 receptor which Case (1957) suggested as a logical means
for an insect to monitor its internal environment The
nerve cord was removed and the deefferented dorsal nerves
were exposed to C02 No responses were noted
The area of the pleural fold was then exposed to
1 5 10 and 15 solutions of acetic acid ethyl
alcohol sodium hydroxide and sodium chloride in saline
Again no responses were noted No afferent activity was
initiated when distilled water was placed on the fold in an
attempt to detect an osmoreceptor The possibility of
thermoreceptors was ruled out by applying saline heated
between 30 and 50 degrees centigrade
Stimulating dorsal nerve efferents while recording
from DN-3 with sensory connections intact failed to initiate
an afferent response On the basis of this information the
possibility of a muscle receptor organ contributing to this
nerve was ruled out
DN-4A
Finlayson and Lowenstein (1958) reported a singleshy
40
Sensory
~~rvJNfmiddot+r((Jr4VJiu+)rJJ~rvI~r-frWJ)~II~~rJ~fl+j~~L pound90 M5elt-I
~~~II)-~~rlr)hl~I~I~I~~~IIl+~~_hl~II~~1 I I I I I I I I 1- middot~1r I I I~
I
Motor
JO record dorsal nerve - motor - lower trace
11 G Lrecord dorsal nerve - sensory - upperJ Ittt trace
) 0 r
Figure 29 Simultaneous recordings from deafferented and deefferented dorsal nerv~ ventilatory rhythm initiated by CO2
41
celled longitudinal stretch receptor under the fourth and
fifth bands of the inner tergal muscle in Periplaneta A
later study by Finlayson and Osborne (1962) reported a
similar vertical receptor near the longitudinal receptor
Both receptors are found in Blaberus (Fig 30)and each
contributes a fiber to DN-l+A
These receptors were destroyed during the usual
dorsal dissections used in these experiments and were
usually not included in the electrophysiological
experiments However it was determined that electrical
stimulation of DN-4 does not elicit a response in the
homolateral ventral nerve one of the major intraganglionic
pathways under study
Prelimary electrophysiological analysis of this nerve
indicates several sensory cells in addition to the vertical
and longitudinal stretch receptors (Fig 31)
DN-4B INCLUDING CARDIAC SEGMENTAL NERVE
DN-4B is of considerable importance to this study
because the distal end terminates in the lateral nerve of
the heart In heart studies the dozen pairs of these distal
branches have been referred to as the segmental nerves
(Alexandrowicz 1926 McIndoo 1945) More proximally
DN-4B which contained four motor fibers divides into
small branches to innervate the inner tergal muscle It was
possible on several occasions to record spontaneous activity
which indicated at least two fibers were present in the
42
Figure 30 Methylene blue whole mount of the longitudinal receptor (long arrow) and vertical receptor (short arrow) in the dorsal body wall of Blaberus (225X)
I I ~~middot~VJJ-~1t4Ji~~~)~~t~~~ t 18)(N__ U J difl
Figure 31 Afferent actiVity from DN-4A Recording made from DN-4 Nerve cut proximal to recording electrode DN-4B destroyed Note more than two spike heights are discernable
43
cardiac segmental nerve
There arc no sensory elements in DN-4B including the
cardiac segmental nerve Spontaneous activity on the
cardiac lateral nerve does not contribute to or affect the
activity on DN-4B This has been determined by recording
from a deefferented DN-4B connected to the lateral nerve
Electrical stimulation of the lateral nerve resulted
in a response of three fibers on DN-4B (Fig 32) In view
of the fact that lateral nerve activity is not conducted
to the segmental nerve these fibers are evidently
antidromically stimulated dorsal nerve efferent fibers The
three responding fibers followed up to 50 stimuli per
second
Relationship of DN-4B Fibers to the Heart Rate
DN-4B participates in the respiratory rhythm The
following experiments (Table I) were designed to determine
whether or not the respiratory rhythm affects the heart
The heart rate in the continous presence of high
concentrations of CO2 slows and eventually stops To
determine whether this slowing of the heart is a result of
central neural inhibition or an intrinsic property of the
heart itself heart rates were counted in intact animals
(Table I-A) and in isolated heart preparations (Table I-B)
in the presence of CO2 _ The heart rate slowed appreciably
in both instances indicating an effect of CO2 independent
middot3AJBU
45
TABLE I Typical experiments showing the effect of CO2 on the cockroach heart rate
A Decapitated Otherwise Intact Cockroach 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
107 104 105
Minutes 1 2 3 4 5
Heart rate (beats per minute) during application of CO2
90 74 64 31 Narcosis
B Isolated Heart 100 CO2 Administered In A Gas Chamber
Normal heart rate (beats per minute)
100 102 90 94
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
51 24 28 Narcosis
All hearts were from adult male Blaberus
46
TABLE I Typical experiments showing the effect of C02 on the cockroach heart rate
C Isolated Heart - Lateral Nerve Intact CO2 Dissolved in Saline
Normal heart rate (beats per minute)
60 62 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
54 52 46 27
D Isolated Heart - Lateral Nerve Cut - In Six Places CO2 Dissolved In Saline
Normal heart rate (beats per minute)
60 68 66
Minutes 1 2 3 4
Heart rate (beats per minute) during application of CO2
44 52 40 34
All hearts were from adult male Blaberus
47
of central neural control This experiment does not exclude
the possibility that the cardiac lateral nerve could mediate
the CO2 effects To test this isolated hearts were exposed
to CO2 before and after the lateral nerve was cut in six
places (Tables I-C and I-D) In both instances the heart
slowed under the influence of C02 This result shows that
the heart muscle is independently sensitive to CO2 but it
does not rule out the possibility that the lateral nerve
may enhance the CO2 sensitivity of the heart
LATERAL CARDIAC NERVE
Six to ten fibers fire spontaneously on the lateral
nerve Several fibers exhibit opposite polarities to the
majority of the fibers indicating that the spikes travel
both anteriorly and posteriorly (Fig 33) At least two
fibers which travel from posterior to anterior have a
spike duration of about 4 msec (Fig 34)
II RESPONSES TRIGGERED BY VENTRAL RECEPTORS
Stimulation of the ventral nerve resulted in a
large number of responses which are described in this
section Since after an exhaustive search the mid-sternal
receptor was the only significant receptor found on the
ventral nerve it is probable that this receptor organ
normally triggers all of the reflexes described here
48
~ Ill I I I I~~Vr1vJI v-W-V 1)fYi1-Iv11 I~v 1- I (I I I J bullbull bull
bull I I r
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Figure 33 Activity recorded from lateral nerve Arrow points to spike with polarity opposite to most spikes in record
1 I I ~ I ~r---r-~vv--~--vv- i
1
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lOQMf(C
Figure 34 Long duration spikes on the lateral nerve
49
A VENiHAL NERVE TO II010LATEHAL DOHSAL
NEHVE PATmlAY
Stimulation of a ventral nerve initiated a response
in the dorsal nerve of the same ganglion Typically five to
eight dorsal nerve fibers responded at a singie threshold
which varied between 25 and 6 volts depending on the age of
the preparation Occasionally the response could be divided
into a lower threshold group (Fig 35) of two or three
fibers and a higher threshold group of three or four fibers
The typical response followed up to 7 or 8 stimuli per
second in freshly dissected animals The stimulus strengths
for the ensuing reflex studies were frequently adjusted to
insure that the maximum number of fibers were activated
This response to ventral nerve stimulation travels
up the connective on the side stimulated to the next anterior
ganglion where it sj~apses then returns via the same
connective to the dorsal nerve This pathway was established by
monitoring the response as it traveled through the reflex
arc Afferent spikes traveled at 2~ to 3 meters per
second in the ventral nerve Conduction time through the
first ganglion normally required 1 to 2 msec too fast for
complicated synaptic events to occur The response was
conducted anteriorly through the connective at the slightly
lower velocity of 2 meters per second and followed trains
of stimuli up to 40 per second There was a 4 to 5 msec
synaptic delay in the anterior ganglion (Fig 36) where it
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51
appears soue integration occurs Unfortunately
anticromically stimulated efferent fibers partially masked
the afferent response and made an accurate evaluation of
integration impossible For the efferent half of the
pathway the conduction rate was 2 meters per ~econd in the
connectives and 2 meters per second in the dorsal nerves
The conduction time in the posterior ganglion was 1 msec
When the anterior connective was stimulated the response on
the dorsal nerve followed up to 100 stimuli per second
which indicates there are no synaptic events in this half of
the reflex The latency for the entire response was 12 to
15 msec representing an average conduction velocity of 125
to 15 meters per second
The dorsal nerve responses to ventral nerve
stimulation could be traced to DN-1 and DN-4 On DN-1 two
to three fibers were activated (Fig 37) and it was possible
to observe slight contractions of the inner sternal muscle
innervated by this nerve Three to four fibers of DN-4 were
also found to respond to ventral nerve stimulation (Fig 38)
DN-2 and DN-3 did not respond to ventral nerve
stimuli as strong as 8 volts
When the mid-sternal receptor was SUbjected to
natural or electrical stimulation the afferent response was
conducted centrally only No responses were found on VN-2
VN-3 or distally on VN-3A ~ no evidence for a peripheral
synapse
53
B VENTRAL NERVE TO HETEROLATERAL
DORSAL NERVE PATHWAY
In addition to the homolateral pathway just
described stimulation of a ventral nerve also leads to
excitation of fibers in the heterolateral dorsal nerve
The heterolateral dorsal nerve responds with a
latency of 10 to 14 msec Most of the response travels up
the homolateral connective to the next anterior ganglion
where it crosses over and returns to the heterolateral dorsal
nerve via the heterolateral connective (Fig 39) However
the response of one fiber requires only the ganglion
stimulated with the anterior and posterior connectives cut
the fiber continues to respond to natural stimulation
(Fig 40)
The heterolateral ventral nerve does not respond to
ventral nerve stimulation
C VENTRU NERVE REBOUND PATlmAY
Stimulation of the main trunk of the ventral nerve
results in a rebound reflex The response consists of one
VN-2 fiber firing several times in response to each
stimulus Unlike the dorsal nerve response to ventral nerve
stimulation the synaptic activity only occurs in the
stimulated ganglion The rebound portion of the response is
lost when the nerve is cut close to the ganglion
Simultaneous records from the dorsal and ventral
54
record upper trace
record lower trace
stimulate - mid-sternal receptor
Figure 39 Response on the heterolateral dorsal nerve to pressure in the ventral musculature which activated the mid-sternal receptor The response utilizes the interganglionic and intraganglionic pathways described in text Diagram indicates electrode placement Arrow indicates approximate time of stimulation
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57
nerves during stimulation of the ventral nerve indicate that
the ventral rebound response is synchronized with the dorsal
response (Fig 41) The long latency of the rebound reflex
10 to 12 maec could be the result of either slow-conducting
fibers or a long synaptic delay By arranginamp the
stimulating and recording electrodes to observe the input as
well as the output a ganglionic delay of 5 to 6 msec is
observed (Fig If2) If it is assumed that the efferents
conduct at the same rate as the afferents 2 meters per
second the synaptic delay could account for the
synchronization of responses
D REPETITIVE RESPONSE IN HOMOLATERAL CONNECTIVE
An additional response to ventral nerve stimulation
was found in the anterior homolateral connective With
stimuli of 1 to 2 volts above the threshold for the normal
ventral nerve to dorsal nerve response an additional fiber
in the connective began to fire at 8 to 10 spikes per
stimulus (Fig 43) This may be the activity of a higher
threshold sensory fiber with an axon which passes through the
ganglion or it may be indicative of the occurrence of
ganglionic synaptic events In either case this fiber
does not participate in the ventral nerve - dorsal nerve reflex
E RESPO~SES OF THE CARDIAC LATERAL NERVE
Electrical stimulation of the ventral nerve
stimulates one to three fibers on the lateral nerve (Fig 44)
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stii1llaticn at 8 point dtstal to the recording electrodes The first burst of spikes represents tl18 afferent act1vlty~
The second burst 1s efferent~
58
Upper
dorsal nerve
JILgtiJIt
llGrVe nerve
t-1M15j
~ 5 volts
25 volts
~ I
11~tJfV~~--tt~ ~ n t ra1 ~ j) l L ~ -~~~ I
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Plgure 1~1 response to
59
The latencies depend on the distance the recording electrode
is from the segmental nerve-lateral nerve junction In the
experiment described by Fig 44 they vary between 14 and
16 msec In the record described the geometry of the
stimulating and recording electrodes was suchthat the
spikes had to travel from posterior to anterior
Natural stimulation of the mid-sternal receptor was
also observed to evoke a response in the homolateral lateral
nerve The duration of the response appeared to be
proportional to the strength of the input
Utilizing the FM wireless transducer to monitor
heart contractions attempts were made to show that
segmental nerve input can modify not only lateral nerve
activity but also heart rate There were no observable
changes in the heart rate when a single dorsal nerve was
stimulated at intensities up to six volts and at frequencies
of one to fifty per second However heart rate increases
were obtained when two dorsal nerves were stimulated as
shown in Fig 45 In this example the rate before and after
stimulation was 80 per minute and during stimulation it was
90 per minute or 12 increase during stimulation
F EXPERIMENTS TO ESTABLISH FilllCTION OF VENTRAL
NERVE-DOaSAL NERVE REFLEX
Of the responses to ventral nerve stimulation
described above the responses of the dorsal nerve involve
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61
the largest number of efferent fibers and have received the
most attention For the detailed mapping which was the
purpose of that part of the study precisely controllable
electrical stimuli were necessary but to determine the
function of this reflex natural stimuli were more
appropria te
Of the three forms of natural stimulation found to
stimulate the mid-sternal receptor only exposure to carbon
dioxide and co~pression of the ventral sternites elicit a
full dorsal nerve response Groundborne vibrations fail to
elicit a response The effects of each are described in
this section
1 VENTRAL NERVE-DORSAL NERVE REFLEX INITIATED BY
THE CO2 SENSITIVE EL~lENTS OF THE
MID-STEIDfAL RECEPTOR
The following experiments on carbon dioxide effects
were designed to determine whether the mid-sternal receptor
initiates or modifies ventilatory activity Two ganglia
were isolated by cutting all their connections except for
one ventral nerve Large sections of the ventral cuticle
adjacent to the receptor were removed and the animal was
pinned over a longitudinal hole in a wax-filled petri dish
A mound of clay was inserted under the two ganglia to ensure
isolation from the CO2-saturated saline The sensory
connections of the ventral nerve were checked to be sure
62
that they were intact A small drop of C02-saturated saline
was placed on the receptor with a small pipette Excess
saline drained through the holes in the cuticle Under
these conditions it was possible to observe a gradual
increase in the CO 2-induced afferent activity and a
concomitant increase in dorsal nerve motor activity (Fig 46)
Once an induced increase in spike frequency was obtained
additional CO2 placed on the receptor had no further effect
on the dorsal nerve activity Since great care was taken to
insure the drop of saline touched only the receptor it
appears that the afferent activity initiated the dorsal
nerve spike frequency change However two other
explanations are possible The C02 may antidromically
stimulate ventral nerve motor fibers which activate the
dorsal response or enough CO2 may come out of solution to
affect the ganglion or dorsal nerve directly No
evidence on these points is presently available
2 EFFECTS OF RESPIRATORY MOVEMENT ON
DORSAL AND VENTRAL NERVE
AFFERENT ACTIVITY
Compression of the ventral musculature occurs
during a normal respiratory movement During the rhythmic
respiratory pumping movements motor bursts correspond to
bursts of ventral afferent activity Simultaneous records
of afferent activity from dorsal and ventral nerves indicate
they both fire in synchr~ny to the abdominal displacement (Fig 47)
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ors
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ner
ve
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-
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rd
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rd
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pp
er tr
ace
Fig
ure
47
D
ors
al a
nd
ven
tral
sen
sory
el
emen
ts
resp
on
din
g
to v
en
tila
tory
pum
ping
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emen
ts
Do
ts
ind
icate
co
mp
ress
ion
0
V
l
64shy
3 REFLEX RESPONSES TO PRESSURE WHICH ACTIVA~ES
THE MID-STEmlAL RECEPTOR
In order to determine whether abdominal compression
is capable of initiating a response on the dorsal nerve the
following experiment was carried out Two ganglion were
isolated and deafferented with the exception of a ventral
nerve Recording electrodes were placed under the ventral
nerve and its homolateral dorsal nerve By gently
compressing tlle ventral musculature with a small glass rod
it was possible to activate the receptor The dorsal nerve
was found to respond under these conditions (Fig 4-8 A and B)
The efferent response varied with the intensity of the input
In addition to the dorsal nerve response compression
stimulated a response in the thoracic connectives When
the mid-sternal receptor of A-I ~~s stimulated naturally a
response was found between the connectives of the second
and third thoracic ganglia (Fig 4-9) but the
heterolateral thoracic connectives did not respond
Afferent activity from A-3 did not reach the thoracic
connectives
4 REFLEX RESPONSES TO GROUNDBORNE VIBRATIONS
The third possibility that t~e mid-sternal receptor
serves to relay information about groundborne vibrations to
65 I bull I 1 l I 1 I
0 ~--14~ ~~I~ I~t-yenNImiddoty~iJr bull j I l i I r 1
middot~middotmiddot1middotmiddotmiddot
I 1 It I f 1I bull l 1
A-~~Ijlr~r ~r3f~rf(-tWfcJ~fl tffrr(~~~ tlrff(tf ~s(~
1 1 1 1 _ - I l II 1 1 I t t ~~ ~~~l~-iYfwIb~(I~IgtL1p~1
bull bull bull bull bull I I bull 1 bull II I bullbullbullbull I I
B 1 I bull ~~~~II1il~jIwowoI(~~tJ~r
oQM5fG r1 I J I I tmiddot
record upper trace - motor~ 0~
-record ~ lower trace - sensorygtJI 0
Figure 48 (A and B) Ventral nerve to homolateral dorsal nerve reflex induced by compression in the ventral musculature which activates the mid-sternal receptor
-- - -- cshy=_-o~ -~ _r~
f OOM5tC
olmiddot1i~~1lt-I~~~n ff~IWimiddotjII~iL~~it~~1~1~~ll)~~~~f~~flf1r1~~ftN~Ti~ I ~I I [I i 01 III I JIliJl J I [ J I I JI
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Figure 49 Response on the thoracic connectives upper trace between T-3 and T-2 to natural stimulation of the mid-sternal receptor of A-1 lower trace Arrow indicates approxima te time the stimulation lias begun
66
the dorsal and ventral musculature was tested in the
following way two abdominal ganglia were isolated and all
peripheral nerves were cut except one ventral nerve which
was to provide sensory input Recording electrodes were
placed on the ventral nerve and on the homola~eral dorsal
nerve to monitor both sensory input and motor output
Analysis of 12 records of such experiments failed to turn up
dorsal nerve responses to tapping stimulation (Fig 50) It
is possible that the sensory response to tapping is too weak
to activate the synapses necessary for the dorsal response
It did not seem entirely reasonable that the clearshy
cut sensory response to tapping would completely fail to
produce a motor response so responses were sought on other
nerves No responses were found on the other motor nerves
of the same ganglion but a response to tapping was recGorded
from the posterior homolateral connective (Fig 51)
III RESPONSES TIGGERED BY DORSAL NERVE RECEPTORS
A DORSAL NERVE TO HOMOLATERAL VENTRAL NERVE PATHWAY
Stimulation of DN-3 results in a response on the
homolateral ventral nerve Two thresholds are present in
fresh preparations At a threshold of 25 volts one and
sometimes two fibers responded to single well-spaced stimuli
(Fig 52) At 3 to 35 volts three or four additional
fibers responded These responses followed trains of
stimuli up to 12 per second The central pathway of this
6(
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Pirsure 50 Smul trJneOf recording of dorBal nerve efferent +Jmiddot middot +~ (() bull --t8) p-(l flmiddotmiddot~llrQl n=1-~C nffltIr O lj at~1 t-shy
(uP1-~8r trace) dUTlnz stimulation bv ~roundborne vibrations ac ~ f L - _~ _ u ~ t -~ 1 lk I ~ ~ t v J - Y
-- ~~- -shy
I I r - yen ~ ~middot~~~_--_V
~--middotmiddotmiddot------~ryenl~~~---r--~r_-r---__lmiddot
~dLcJ-l
Figure 51 Simultaneous records of sensory input on an intact ventral nerve (upper trace) fLnd response on posterior homolateral connective
]ligure 52 Primary response on the m~in trunk of the v2ntral nerve to dGrsill 1~2rvc ~tl~~Jl t]~YJ
68
response is confined to the ganglion stimulated since the
response remains after anterior and posterior connectives
are cut The latency varied from eight to nine msec
representing an average conduction velocity of 225 to 250
meters per second
The response to dorsal nerve stimulation can be
traced to DN-2 (three to four fibers activated) VN-3
(two fibers) and VN-3A (one to three fibers) (Fig 53
A B C)
B DORSAL NERVE REBOUND PATHIIAY
The stimulation of DN-3 or the main dorsal nerve
trunk initiates a rebound reflex This reflex must travel
to the next ganglion to make synaptic contact It then
returns on the same side to activate the dorsal musculature
The response which lags 1-2 msec behind the ventral
response (Fig 54) has a latency of 10-12 msec representing
an average conduction velocity of 2 to 25 meters per
second The synaptic delay is on the order of 45 to 5
msec (Fig 55) These reflexes are not as closely
synchronized in time as the dorsal and ventral nerve
responses evoked by ventral nerve stimulation Six to
eight fibers constitute the total response
In terms of number of fibers activated the dorsal
nerve rebound reflex is of more importance to the animal
than the dorsal nerve to ventral nerve reflex In the
rebound reflex two fibers respond on DN-1 (Fig 56-A)
69
iMJc(A 0 t-fpoundS B
Ily~~ I ~ ttJM~=-~
c
Figure 53 Respons2s on branches of the ventral nerve to dorsal nerve stimulation A Response on VN 3 Response on VN-3 C Response on VN-3A
-
1 ~
I I Ifi~ ~rt~middot~JfI-t~t-~ 11 0 ~t
I
ventral nerve response i ~lt bullbull - i -I ~~scord I V ~-rlti l- 1 dorsal nerve rebound I~ NHr~-gt~OOd i
Figure 5l~ Dorsal TeDoL1nd and vsntr[-Ll t8spona to stlmulation of the dOYial nervmiddots Aff8T~nt res)()ne TIIBI1e1 bS 10~lg arro~r and effermiddot~nt reS00Iises m21~k2d by short arrOi~ ~J bull
70 1 Ib1Slt
t
t
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gt _f record
- - ~ stimulate
)1 n~ Flgl1re 55 Dorsal nerve rebound reflex recorded in the -nteTlor~ - co~r~0i~~r~ -~t~~0~ant ~_c ~-n~~A~n~ i
(qho~~__Jv _~_~ ~ --middot _ - ~ O ~~~yo~~ ~t -ltshy
~rromiddotT r1~A dlt-l~-gt-(nf~ -r--gtlrn~~ + ~rnrCgtn Ion - -0)rJ -11 v j~tll Llt -J Jlt-- 11 l-JJ _l _ - c J ~
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A B c
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71
four fibers on DN-2 (Fig 56-B) and three to four fibers on
D~-4 (Fig 56-C) When DN-1 or DN-4 was stimulated no
rebound reflex was initiated and this served as a control
on the above data
The rebound reflex also reaches the lateral nerve
When a dorsal nerve was cut then stimulated distal to the
cut a response of three to four fibers was found on the
lateral nerve (Fig 57) When the dorsal nerve was left
connected to the ganglion both short and long latency
responses were obtained (A and B respectively in Fig 58)
Response B was probably the rebound response
C DORSAL NERVE TO HETEROLATERAL DORSAL NERVE PATHWAY
Natural or electrical stimulation of a dorsal nerve
leads to a response of the heterolateral nerve of the same
ganglion (Fig 59) Cutting experiments show that the
main response travels up the homolateral connective to the
next anterior ganglion crosses over presumably makes
synaptic contact and returns via the heterolateral connective
to the heterolateral dorsal nerve When either right or
left anterior connective is cut the typical response is
lost Fig 60 shows the response within the heterolateral
connective The cut below the recording point eliminated
the possibility of recording from fibers which crossed
over in the posterior ganglion
A single efferent fiber receives synaptic input in
the stimulated ganglion since it continues to respond to
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59
H
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and
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ral
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ve
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to n
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ula
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ure
60
R
esp
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se
to
DN st
imu
lati
on
on
the
cu
t h
ete
rola
tera
l --
Jco
nn
ecti
ve
VJ
74
natural stimulation after the anterior connectives are cut
(Fig 61) This is the same fiber which receives input from
the heterolateral ventral nerve of the same ganglion under
the same recording conditions This fiber has also been shown
to respond to cereal nerve stimulation (see diagram Fig 62
which emphasizes the importance of this fiber)
In addition to these responses in the dorsal nerve a
response to dorsal receptors could be traced to the cardiac
lateral nerve Natural stimulation of one set of dorsal
receptors evoked a response on the heterolateral lateral
cardiac nerve (Fig 63)
There is no response to dorsal nerve stimulation on
the heterolateral ventral nerve
D EXPERIMENTS TO ESTABLISH FillCTION OF
DORSAL NERVE TO VENTRAL NERVE REFLEX
Mechanical pressure in the region of the dorsal
receptors initiated a motor change on the ventral nerve
(Fig 64) The response varied with the intensity of the
afferent input Stimuli of this nature appear to simulate
the normal pressure changes observed during the respiratory
rhythm
Tapping stimuli sets up groundborne vibrations which
initiate responses in three to four fibers of the
homolateral ventral nerve (Fig 65) This indicates that a
receptor contributing to DN-3 serves the physiological
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76
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Figure 62 Diagram emphasizing the dorsal nerve fiber which responded to mUltiple sensory inputs This cell which was spontaneously active also responded to stimulation of the heterolateral do~sal nerve (1) heterolateral ventral nerve (2) and homolateral cercus (3)
Figure 63 Two examples (A and B) of the response on the heterolateral lateral nerve to natural stimulation of the dorsal nerve Upper trace is the dorsal sensory input Lower trace is the response (between arrows) on the lateral nerve
reco~d
IOHer trace
------- ----__--__--__------~---_-- -- -
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77
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ure
6
4
Res
po
nse
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th
e ~omolateral
ven
tral
and
d
ors
al
nerv
es
to
pre
ssu
re
stim
ula
tio
n o
f th
e
do
rsal
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tors
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rrow
in
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ate
s ap
pro
xim
ate
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egu
n
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79
I I t1_~ l ~ ~ I ~ bull ii~ ~ r U r bullbullbull~_4 ieraquo [ di bullbull I I 10 I I J I I bull I I iii Iii I i I opA1lec t
1 11~1 1 111 r I II I bull
tZ4middotmiddotlr ~middot_Jmiddotmiddot-r -10 jr-r--r--~rp I ~
J G ~ecord - upper trace stimulate
lt record - lower trace11 0
Figure 65 Afferent response to groundborne vibrations (upper trace) on the dorsal nerve activating an efferent response on the ventral nerve (lower trace)
80
function of responding to vibrations
IV STABILITY OF REFLEXES
The dorsal nerve to ventral nerve and ventral nerve
to dorsal nerve reflexes differ not only in th~ number of
fibers participating but also in their stability A limited
study on their longevity was undertaken to aid in planning
experiments
Thresholds were measured every 15 minutes Using well
spaced stimuli The ventral nerve to dorsal nerve reflex
remained viable for up to an hour and half The less
sensitive dorsal nerve to ventral nerve reflex remained for
up to three hours sometimes longer The thresholds
progressively increased in both reflexes as the preparation
aged (Fig 66) When the ventral to dorsal nerve response
was lost in the dorsal nerve a response could still be
obtained in the connectives This indicates that it was the
synaptic activity in the next anterior ganglion which was
lost When the tracheae and ventral diaphragm supplying the
nerve cord were completely removed the longevity of both
reflexes was diminished This indicates that oxygen
deficiency is at least partly responsible for the loss of
synaptic activity
The difference in longevity of the two responses
implies there are no common interneurons between reflexes
o -------1---1------------1------
Figure 66 bull A typical experiment comparing the thresholds of the dorsal to ventral nerve reflex to the ventral to dorsal nerve reflex
ltA Dorsal nerve to ventral nerve reflex
120 150 180 210 24090
81
~ ven tral nerve
to dorsal nerve reflex
Time in minutes
6030
14 1 13
12
1 1
10
9
III 8 p r-l 0 7 ~
l1 6 d r-l 0 5
lt1 Dl OJ I-t 4 t1
3
2
82
V FURTHER INTERGANGLIO~IC PAT~~AYS
The reflexes described to this point involve activity
on the homolateral dorsal and ventral nerves belonging to a
single ganglion The following survey was undertaken to
determine possible broader distribution of sensory
information in the abdominal nerve cord The results are
summarized by Tables II and III Primarily abdominal
ganglia 2-5 have been used because of their accessibility
By studying pathways stimulated both electrically and by
natural stimulation it was possible to ascribe a
physiological function to pathways whose significance would
be otherwise undetermined This technique combined with the
cutting of connectives has made it possible to ascertain
gross regions of synaptic activity in several instances
Because high stimulus intensities may activate
non-physiological pathways only those reflexes have been
presented which have thresholds below 7 volts Since
repetitive stimulation has been noticed to have similar
undesirable effects only responses triggered with well
spaced stimuli were normally used Smalley (1963) reported
that stimulation of a dorsal or ventral nerve can produce a
response in any of the connectives leading from the
ganglion of the nerve stimulated depending on the stimulus
intensity Under the more conservative levels of stimulation
used here the dissemination of sensory information appears
to follow more restricted pathways
83
An approximate threshold is reported for each
reflex Since the thresholds vary with the age of the
preparation those reported are from freshly dissected
animals
Latencies for a single reflex are variable from animal
to animal but never by more than several milliseconds The
calculated conduction velocities were obtained by dividing
the length of the presumed pathway by the latency of the
earliest response Since the latency value includes
synaptic delays of unknown duration the average conduction
velocity will always underestimate the conduction velocities
of the axons carrying the response
The calculated conduction velocities are averages
for the fastest fiber(s) participating in the response
The maximum rate at which a reflex can follow
stimulation is also presented In conjunction with
latencies and average conduction velocities this value
may indicate the complexity of the synaptic activity
The connectives traveled are only reported for
reflexes of some interest The natural stimulation methods
described in this section are the same as described
previously A plus sign in the appropriate box of Tables II
and III indicates the reflex may be induced by natural
stimulation of one receptor or another A minus sign in the
box indicates the appropriate receptor was stimulated but did
not induce an efferent response A blank space indicates
the reflex was not tested
Tables II and III summarize the wider dist~ibution
84
of responses to dorsal and ventral nerve stimulation within
the abdominal cord
A SPREAD OF VENTRAL NERVE INPUT
Responses to ventral nerve input were ~raced
anteriorly two ganglia (Table II pathways 1-6) On the
homolateral side both ventral and dorsal nerve responses
were found on the first and second anterior ganglia
(Table II pathways 1-4) On the heterolateral side single
stimuli produced responses on the ventral and dorsal nerve
of the first anterior ganglion (Table II pathways 5 and 6)
The heterolateral ventral and dorsal nerves of the second
anterior ganglion could also be stimulated but only by
repetitive stimulation (Table II pathways 7 and 8)
The posterior spread of ventral nerve input was
limited to a response on the homolateral dorsal nerve of the
first posterior ganglion (Table II pathway 9) No
response was seen on the first posterior ventral nerve
(Table II pathway 10) and no responses were seen on
posterior ganglia on the heterolateral side (Table II
pathways 11-1 Jf)
B SPREAD OF DORSAL NERVE INPUT
Responses to dorsal nerve input spread anteriorly to
ventral nerves and dorsal nerves on both sides of the
next two anterior ganglia (Table III pathways 1-8)
Latencies for all of these responses were of the order of
85
10-15 msec The homolateral ventral and dorsal nerves of
the first anterior ganglion were able to follow stimulation
at a higher frequency (8-10 per second) than was true of the
other anterior pathways (which responded up to 3 per
second)
The posterior spread of responses to dorsal nerve
input was more restricted than the anterior spread (Table
III pathways 9-14) On the homolateral side dorsal nerve
responses were found on the first and second posterior
ganglia (Table III pathways 10 and 12) but ventral nerve
responses were found only on the first posterior ganglion
(Table III pathways 9 and 11) On the heterolateral side
single stimuli elicited no responses on posterior ganglia
but repetitive stimulation produced a response on the
ventral nerve of the first posterior ganglion (Table II~
pathways 13 and 14)
C SEVERAL GENERALIZATIONS ABOUT THE DISSEMINATION
OF SENSORY INFOffi1ATION
Several generalizations about the dissemination of
sensory information may be drawn from the results
summarized by Tables II and III
(1) The spread of responses tends to be stronger to
the homolateral side than to the heterolateral
side (11-3 4 vs 11-7 8 111-9 10 vs 111-13
14) The most complex response patterns (in terms
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
U~
86
12-15
1-2
+
15
35
5-6
+
10-12
3
15-17
7-8
3-4
25
+
20
8-10
8-10
3-4
(lL) 0 L (2)
1D ~
Latency (msec)
Responds to natural stimulation
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
Conduction rate (meters per second)
87
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Ascending Pathways
(4) ( (5)~ U recoM ~ (6) - U ~
n~ ~ r lor0n [ I n~--~npound ~1 ill~ I1nJf I [jFnu~ n 1 n
Number of fibers 3-4 1-2 4-5 participating in the reflex
Threshold in 35 4 6-8 volts
Conduction rate 15 1 1 (meters per second)
Latency (msec) 10-15 8-10 13-18
Maximum frequency 4-5 6 1-2 followed (stimuli per second)
Responds to + Easily natural activated by stimulation repetitive
stimulation
88
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Number of fibers participating in the reflex
Threshold in volts
Conduction rate (meters persecond)
Latency (msec)
Ascending Pathways
(7) Jiilshy~t~u l ~nl gt~~
3-1+
~
(~~J )JJpound
1ur[ )JsiITlU~T-e
3-4
Maximum frequency followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation 4 volts 20 per second
Responds only to high frequency repetitive stimulation 3-5 volts 20-30 per second
89
TABLE II Interganglionic pathways Responses to ventral nerve stimulation
Descending Pathways
(9) -J 0 L (10)~ (11 3I1~14)
7 ~u~ J1fmltMramp1 -~11 r a J D ~ ~ 1~
(II)
If(Il) rlfOd II shy11 n1 r~od (1) [
lt11) - Dr
Number of fibers 2-3 participating in the reflex
4Threshold in volts
Conduction rate 125 (meters persecond)
La tency (msec) 12-15
Maximum frequency 3 followed (stimuli per second)
Responds to + natural stimulation
No response to well-spaced stimuli
Ascending Pathways
90
2
3
10-13
5-7
3-4
+
1 5
12-15
8-10
4-5
3t
(2) ~D ~ (3) J u ~
1 r 1 reeord ~ U retolci J t1 ~
~ U 1 tl [Shu~ gt1111 ( gt1 a r
2
3
10
+
10
4-5
(1~OL
)u_ gtf1nl
Number of fibers participating in the reflex
Threshold in volts
Maximum frequency followed (stimuli per second)
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Responds to natural stimulation
La tency (msec)
Conduction rate (meters per second)
91
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways
1nll~
Number of fibers 3-4 2 3-5 participating in the reflex
Threshold in 4-5 5-6 5-6 vol ts
Conduction rate 1 to 15 1 1 to 15 (meters per second)
Latency (msec) 13-15 10-12 12-15
Maximum frequency 3-4 2-3 2-3 followed (stimuli per second)
Responds to + + natural stimulation
92
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Ascending Pathways JL (7)~ (8)
rtotord] f rttlllti ~[ ~U~ ~U~~u1~middotw lpoundI[~~~ )~ 1111 t
Number of fibers 1-2 4-5 participating in the reflex
Threshold in 6 6 vol ts
Conduction rate 1 1 (meters per second)
Latency (msec) 12-13 12-15
Maximum frequency 2-3 2-3 followed (stimuli per second)
Responds to natural stimulation
93
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
No response
=Ji~~Q
~ut ~ut -l~reeord
( 1-1 )
+
4
125-15
12-15
5-6
2-3
12-15
15
6-8
Descending Pathways
Responds to natural stimule tion
Latency (msec _)
Threshold in volts
Maximum frequency followed (stimuli per second)
Number of fibers participating in the reflex
Conduction rate (meters per second)
94
TABLE III Interganglionic pathways Responses to dorsal nerve stimulation
Descending Pathways
( 14) (12)) n (13)--J n 4
JJn~-lJu~ ( fA] f reeotl [J [J Ieto~ 0 r ~ gt1) 0 r ~
Number of fibers 3-4 No participating in 2-3 response the reflex
Threshold in 4 volts
Conduction rate 1-1 5 (meters per second)
Latency (msec) 15-18
Maximum frequency 5-6 followed (stimuli per second)
Responds to + natural stimulation
Responds only to repetitive stimulation
95
of ease of fatique variability of latency etc)
recorded during this part of the experiment were
recorded on heterolateral nerves (11-7 8)
(2) The spread of responses tends to be greater to
anterior than posterior nerves (III-~ 5 vs
III-11 14 11-1 6 vs II-10 12) bull
(3) The spread of responses tends to be stronger to
nearby ganglia than to more distant ganglia
(III-9 vs III-11 III-14 II-78)
(4) In any dorsal nerve response to ascending input it
can be seen that both the next anterior ganglion
and the connective homolateral to the responding
dorsal nerve are required (111-2 4 6 8 11-2
4 6) The probable anatomical basis for this
result is that the cell bodies and synapses of the
dorsal nerve cells lie in the next anterior
ganglion and their axons must pass through ~he
homolateral connective en route to the dorsal
nerve This is consistent wlth a conclusion
reached earlier in this paper (see section II) and
by Smalley (1963)
(5) In dorsal nerve responses to ascending input it was
shown that the connectives homolateral to the
sensory input must be intact (111-2 4 6 8
11-2 4 6) EVidently the afferent part of the
pathway in these responses passes up the homoshy
lateral connectives and no crossing-over of
96
sensory information occurs in ganglia posterior
to the ganglion in which synapses take place
However in pathways which lead to excitation of
ventral nerve fibers there appears to be some
crossing over posterior to the ganglion in which
the synapses are located (111-5)
(6) Ventral nerve responses do not require a ganglion
anterior to the responding nerve in ascending
reflexes (111-1 3 5 7 11-1 3 5 7) nor in
descending response do they require a ganglion
more posterior than the one on which the ventral
nerve responded (111-9) The probable anatomical
basis for this generalization is that the synapses
for the ventral nerve fibers all lie within the
ganglion of origin of a ventral nerve
(7) The dorsal nerve responses also tend to have
longer latencies and higher thresholds than do the
ventral nerve responses In general the dorsal
nerve responses have more labile properties
suggesting more complex synaptic properties and
more complex integ~ative phenomena This is
consistent with another indicator of more complex
synapses in an earlier section of this paper
(section IV Fig 66) it was shown that the
thresholds of dorsal nerve responses rise more
rapidly and are more sensitive to detracheation
than are dorsal nerve responses
97
VI EVIDENCE FOR PROPRIOCEPTIVE REFLEX SYSTEM
BEHAVIORAL OBSERVATIONS
One very likely function for reflexes studied here is
that of proprioception Although antigravity reflex systems
have been described in the abdomens of other itlsects
(Weevers 1966) and in cockroach thoracic legs (Pringle
1940) there appears to be no published evidence for a
proprioceptive reflex system in the cockroach abdomen and the
following experiments were accordingly carried out The
effects of adding weights to the abdomens of male
Periplaneta were observed Small drops of warm wax were
progressively layered on the posterior dorsal abdomen of
anesthetized roaches until the desired weights were reached
The normal roach walks with abdomen elevated and
parallel to the ground Four out of six roaches tested were
capable of carrying loads up to 110 per cent of their body
weight When the loads were placed well to one side of the
midline the animal continued to maintain a normal posture
with loads up to 20 or 30 per cent of total body weight
Animals with their connectives cut between A-3 and
A-4 continue to walk naturally and maintain a normal
posture however a load of 10 per cent of their total
body weight is enough to cause the abdomen to drop when
walking Similar results were found when the connectivesmiddot
were cut at two points between A-2 and A-3 and
between A-3 and A-4 When the connectives were cut between
98
A-l and the third thoracic ganglion the abdomen could not
be held up and was dragged as the animal walked
DISCUSSION
I MID-STERNAL RECEPTOR
Shankland (1966) recorded afferent imp~lses from
nerve B-1 of Periplaneta (which corresponds to a branch of
the ventral nerve of Blaberus) and inferred the presence of
a midventral stretch receptor On the basis of methylene blue
preparations he concluded that the efferent impulses
originate from a connective tissue strand suspended between
two points on adjacent sternel plates although he could not
find the cell bodies or ascertain the number cf fibers
constituting the stretch response A careful histological
study of the midventral region of Blaberu~ in this
laboratory indicates the mid-sternal receptor described here
is the only sensory element present Furthermore methylene
blue preparations in Periplanete co~firm the presence of
bipolar cell bodies in the same location as in Bla~~
It must be concluded in the light 0 this new evidence that
Shankland has attributed the physiological activity of the
mid-sternal receptor to the wrong anatomical structure
In addition to cells responsive to stretch this
study demonstrated some cells capable of responding to
both CO 2 and stretch These results raise two questions
are the responses to CO 2 part of the normal physiology of
these cells If so are there bimodal receptor cells
which respond to both CO2 and mechanical stimuli in the
100
normal functioning of the organism There are no previously
reported CO2 receptors in insects although Case (1957)
pointed out the reasonableness of such an organ On the
basis of the evidence presented here it would be premature
to assign a definite CO2 receptor function to the rnidshy
sternal receptor The argument that this is a CO2 receptor
could bc strengthened by data linking this receptor with the
respiratory center or data showing that the CO2
concentrations which stimulate this receptor are within
physiological limits Until such data become available
the following conservative conclusion must stand the
mid-sternal receptor serves a mechanoreceptor function and
three of its cells are unusually sensitive to C02
II LATE~~L CHORDOTONAL ORGANS
Florentine (1966) recorded from an abdominal receptor
sensitive to substrate and airborne vibrations in
Periplanet~ Based on his histological study which
utilized cross sections of the lateral fold Tegion it was
concluded that the anatomical structure responsible for
the physiological response was a fan-shaped multicellular
hair receptor The present histological study on ]Laberus
and a confirming study on Periplaneta (Smalley 1968
unnublished) indicate that what Florentine considered to be
sensory cell bodies were actually part of attachment
(Fig 20) of the lateral chordotonal organ described in
this study Light pressure on the tergite juncture the
101
approximate location of the lateral chordotonal organ
evokes afferent spikes of a different spike height than
those induced by groundborne vibrations In the light of
this evidence it must be concluded that Florentine attributed
a physiological response to the wrong anatomical structure
Although this study has ruled out the structure
postulated to be responsive to groundborne vibrations an
alternative has not been found Because Vaseline does not
inhibit the receptor activity and the histological study
does not show innervation of the hairs hair receptors
seem to be ruled out It is probable that several of the
small nerves contributing to DN-3A and DN-3B are the true
vibration receptors
III FUNCTIONS OF THE MID-STE~AL RECEPTOR
AND LATERAL CHORDOTONAL ORGAN
A JmYTH1~OMETER FUNCTION
The dorsal and ventral receptors probably serve
both as proprioceptors and as rhythmometers The idea of a
rhythmometer was suggested in 1928 by Eggers who
postulated sense organs coordinating respiration and
circulation Hughes (1956) has found afferent fibers from
unidentified receptors in the locust which respond to
inspiration others which respond to inspiration and
expiration and a third group which is inhibited during
102
inspiration He suggested this information is used to
modulate ventilatory movements Farley and Case (1968)
have verified Hughes hypothesis in the American cockroach
They have shoHn that the afferent input from unidentified
abdominal receptors is capable of altering the frequency of
the respiratory movements The nature of their stimulation
techniques was such that they probably activated the
mid-sternal receptor
For a receptor to modulate the activity of the
thoracic respiratory center it must be capable of activating
a pathway which reaches the respiratory center The midshy
sternal receptor meets this reqUirement since this study
shows that its response to natural stimulation can be
found as far anterior as the thoracic connectives The
CO2-sensitive property of this receptor could possiblYbe
a factor in initiating the entire ventilatory cycle
Both the lateral chordotonal organ and mid-sternal
receptor fire during abdominal pumping movements so in
actuality either or both receptors could serve as
rhythmometers The answer to the question which receptor
is more important in modUlating the respiratory rhythm
must await additional experiments designed to eliminate
one set of receptors While leaVing the other functionally
intact during induced respiratory movements
B PROPRIOCEPTOR FUNCTIOn
The reflexes under consideration in this stUdy
103
appear also to serve a proprioceptive flmction
The behavioral observations suggest that roaches
carrying a load in some manner compensate for the additional
weight The similar work of Planck (described by 1[ilson
1968) with stick insects carrying weights on their backs
concludes that proprioceptlve loops compensate for the
additional load
A function for the dorsal nerve to ventral nerve
reflex is suggested here In response to load the dorsal
nerve mechanoreceptors are activated The response on the
ventral nerve of the same segment activates the outer
sternal and tergo-sternal muscles The rebound response
on the dorsal nerve activates the inner sternal tergal
and spiracular muscles Since inner and outer sternal
muscles serve as retractors of thc sternum while the
tergal muscles Serve as retractors of the tergum (Shankland
1966) the predicted result of stimulating the dorsal
receptors is a shorting and general stiffening of the abdomen
Ifhen it is considered that similar reflex events arc
occurring on both sides of the animal as well as up and
down the nerve cord it is reasonable to conclude that the
dorsal receptors are involved in postural control
IV SPONTANEOUSLY ACTIVE FIBER RESPONDING TO
MULTIPLE SENSORY INPUT
Several cells have been shown to fire spontaneously
on the dorsal nerve of ~n isolated abdominal ganglion
104
Roeder (1955) reports similar cells present on isolated
thoracic ganglia Wiersma (1962) feels that an understanding
of the functional significance of such spontaneously active
cells would enhance our knowledGe of the transmission of
impulses in the central nervous system It c~n be
conclusively stated that one cell firing spontaneously
receives multiple sensory input from theheterolateral
mid-sternal and DN-3 receptors and the homolateral cercus
(see Fig 62) The synapses for this fiber unlike those of
other dorsal nerve cells are in the ganglion froln which
the dorsal nerve originates
V NEURAL CONTROL OF THE HEART
In 1926 Alexandrowicz described the lateral and
segmental nerves of the heart Since that time there has
been a great debate among physiologists as to the function of
these nerves The evidence presented in this study was
directed toward proof of functional neural pathways which
could convey cardiac regulatory information to the heart
Evidence for a regulatory function lvas sought by
stimulating cardiac segmental fibers Stimulation of a
single dorsal nerve did not alter the heart rate but
stimulation of two dorsal nerves did increase the heart
rate up to 12 per cent over the normal values This
evidence implies neural control over the heart All of
the heart data presented in this paper are consistent with
the conclusions of Miller and Metcalf (1968) who feel the
105
heart is myo~enic with a superimposed neural control
It is worth pointing out that Wigglesworth (19~0)
anticipated this conclusion
SUI1MARY
The purpose of this study was to investigate major
reflex pathways through the abdomen of the cockroach
Blaber~sect poundL~iifer Attention was also given to the
structure and function of sense organs which provide the
normal input to these reflexes
Two previously undescribed abdominal receptors were
found in this study and named the mid-sternal receptor
and the lateral chordotonal organs
The five bipolar sensory cells making up the midshy
sternal receptor are located at the midventral line and
contribute their axons to the ventral nerve The distal
ends of the receptors of a segment converge at
approximately the same spot on the cuticle giving the
receptors a V-shaped appearance in cross section
Electrophysiologically two motor fibers have been shown
to activate the receptor and their presence has been confirmed
histologically Three of the five cells are unusally
sensitive to CO2 two cells respond to groundborne
vibrations and all five cells respond to compression of
the ventral musculature
The lateral chordotonal organ located in the
lateral fold is a seven-celled mechanoreceptor which
contributes its axons to dorsal nerve branch three (DN-3)
At least 14 additional sensory cells contribute axons to
1~
DN-3 Three to four of these cells respond to groundborne
vibrations four to six to rubbing on the ventral surface
and the ~emainder to cuticular deformation
Twenty-nine reflexes initiated by stimulating either
the dorsal or ventral nerves have been studi~d Special
emphasis has been given to reflexes through a single
ganglion
One reflex studied in detail was the response of a
dorsal nerve to stimulation of the homolateral ventral
nerve of the same ganglion The latency of the response
is 12-15 msec representing a conduction velocity between
125 and 15 meters per second The response is found on
branches innervating the inner sternal and tergal muscles
and the lateral nerve of the heart Ventral nerve stimulation
also results in a rebound intraganglionic reflex
synchronized with the dorsal nerve response This response
returns to the tergo-sternal muscle
The heterolateral dorsal nerve responds to ventral
nerve stimulation with a latency varying between 10-14
msec The major response travels up the connectives on the
side stimulated to the next ganglion where it crosses over
and returns to the dorsal nerve via the heterolateral
connective One fiber utilizes an intraganglionic
pathway This cell which responds to heterolateral
ventral nerve input also receives input from the
heterolateral dorsal receptors and the homolateral cercus
108
All of the major branches of the homolateral ventral
nerve respond to dorsal nerve stimulation This reflex
utilizes an intraganglionic pathway Dorsal nerve
stimulation also initiates a rebound response on the same
nerve In this response synaptic contact is made in the
anterior ganglion before the response returns to activate
the dorsal musculature
There is also a major dorsal nerve to heterolateral
dorsal nerve pathway which makes synaptic contact in the
next anterior ganglion before returning to the dorsal
nerve via the heterolateral connective
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Case JF 1957 The median nerves and cockroach spiracular function J Ins Physiol 1 85-94
Eccles JC 1957 The physiology of nerve cells The John Hopkins Press Baltimore
Eggers F 1928 Die stiftflihrenden Sinnesorgane Zoologische Baustein Berlin 2
Farley RD and JF Case 1968 Sensory modulation of the ventilatory pacem~ker output in the cockroach ~ipaneta americana J Ins Physiol 14 591-601
Farley RD JF Case and KD Roeder 1967 Pacemaker for tracheal ventilation in the cockroach Peripla~eta
american~ (L) J Ins Physiol 13 1713-1728
Finlayson LB and O Lowenstein 1958 The structure and function of abdominal stretch receptors in insects Proc Roy Soc B 148 433-449
Florentine G 1967 An abdominal receptor of the American cockroach Periplaneta americana and its response to airborne sound bullJ Ins Physiol 13 215-218
Granit R 1967 Charles Scott Sherrington A biography of a neurophysiologist Doubleday and Company Inc New York
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Hughes GM 1952 Abdominal mechanoreceptors in Dytiscus and Locust~ Nature 170 531-532
Hughes Gll 1965 Neuronal pathways in the insect central nervous system p 179-112 In JE Treherne and JWL Beament The Physiology of the Insect Central Nervous System Academic Press New York
Mclndoo NE 1945 Innervation of insect heart J Comp Keurol 83 141-155
Miller T and R Metcalf 1968 Site of action of
110
Pharmacologically active compounds on the heart of Periplaneta americ8nad J Ins Physiol 14 383-394
Osborne MP and LH Finlayson 1962 The structure and topography of stretch receptors ill representatives of seven orders of insects Quart J Micr Sci 103 227-242
Pringle JWS 1940 The reflex mechanism of the insect leg J Exp BioI 17 8-17
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Roeder KD 1948 Organization of the ascending giant fiber system in the cockroach (Periplaneta americana) J Exp Zool 108 342-362
Roeder KD 1955 Spontaneous activity and behavior The Scientific Monthly 80 362-370
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Stark M KN Smalley and EO Rowe 1968 Staining of insect axons In preparation
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Wiersma OAG 1962 The organization of the arthropod nervous system American Zoologist 2 67-78
Wigglesworth VB 1950 The principles of insect physiology EP Dutton and Sons New York
Wilson DM 1968 The flight control system of the locust Scientific American 218(5) 83-90
111
Yamasaki T and T Narahashi 1959 The effects of potassium and sodium ions on the resting and action potentia~s of the cockroach giant axon J Ins Physiol 3 230-242
