J. Physiol. (1962), 164, pp. 189-199 189With 5 text-figuresPrinted in Great Britain

THE ISOMETRIC CONTRACTION CHARACTERISTICSOF CAT INTERCOSTAL MUSCLE

BY T. J. BISCOE*From the War Department Chemical Defence Experimental

Establishment, Porton Down, Salisbury

(Received 9 February 1962)

Modern techniques have not been applied to the study of the charac-teristics of intercostal muscle contraction in response to nerve stimulation.Martin & Hartwell (1879) and Scharpey-Schafer & MacDonald (1925)recorded spontaneous intercostal contraction, but provided no evidenceabout the speed of contraction. Douglas & Matthews (1952) observedthat spontaneous respiration in the cat sometimes continued after dia-phragmatic paralysis; this suggested that the accessory muscles ofrespiration, including the intercostal muscles, may differ in some way fromthe diaphragm. However, Paton & Zaimis (1951) studying the responsesto D-tubocurarine and to decamethonium considered that the intercostalmuscles resemble the diaphragm and 'slow' types of muscle such assoleus, rather than the 'fast' type such as tibialis anterior.

In order to define a muscle as 'fast' or 'slow' Buller, Eccles & Eccles(1960) used the time course of the single isometric contraction and thestimulus interval at which halfthemaximum tetanus: twitch ratio occurred.These parameters have now been measured for the intercostal musclecontraction in response to indirect stimulation in vivo both, in adult catsand in a few 4-week-old kittens. Furthermore, the sensitivity of themuscles to D-tubocurarine and to decamethonium was examined.A preliminary communication of these results has already been pre-

sented (Biscoe, 1961).METHODS

Cats were anaesthetized with sodium pentobarbitone 40 mg/kg, administered intra-peri-toneally; if necessary further doses were given intravenously during the experiment. Theanimals were artificially ventilated.The isometric contraction response to indirect stimulation was recorded from the inter-

costal, soleus and tibialis anterior muscles. Records were made with an electromyographand an RCA 5734 valve, as described by Talbot, Lilienthal, Beser & Reynolds (1951). Thevalve formed one arm of a bridge circuit of which the out-of-balance voltage was amplifiedand displayed on an oscilloscope. The records were photographed on 70 mm paper andmeasured with a graticule marked in 1mm squares. The temperature of the muscles wasrecorded with a thermocouple and a galvanometer.

* Present address: A.R.C. Institute of Animal Physiology, Babraham, Cambridge.

13 Physiol. 164

190 T. J. BISCOEInterco8tal muscle preparation. Below the level of the eighth rib the diaphragm is inserted

into the costal margin. Therefore, in order to avoid technical complications, only the musclesin the upper seven intercostal spaces were investigated.The thoracic cage was approached through an anterior mid-line incision extending from

the manubrium to below the xiphisternum. Access to the intercostal space selected forstudy was obtained by removing part of the thoracic and abdominal wall below it. A muscleslip 1*0-1.5 cm wide was formed by cutting free a short segment of the rib bounding thespace below. The rib above was transfixed by drills, which were then clamped to the table.The relevant intercostal nerve was crushed centrally and arranged for stimulation, the hemi-thorax being filled with warm liquid paraffin. The muscle was connected to the myographspring by a thin stainless-steel rod and a hook which was engaged through a hole drilled inthe free segment of rib (Fig. 1). The rod passed freely through a hole in the longitudinal axis

Drills M1uscle slip

Rib

Rib.segment.

Fig. 1. Diagram of the experimental arrangement for recordingisometric contractions of intercostal muscle slips.

of a 4 BA screw and was expanded at its upper end. The screw was mounted in the myographspring, and its adjustment allowed the length and tension of the muscle to be set at knownvalues. The muscle temperature was maintained at 37-38° C by drawing the slip into anopen-ended chamber lying between two boxes through which water at 380 C was pumped.The muscle slip was moistened with Krebs's solution. Attempts to maintain the temperatureby inmnersing the muscle in warmed liquid paraffin were unsuccessful. In the few experi-ments in which this was tried the single twitch and tetanus amplitudes diminished by onethird to one half in 15 min.

Soleus and tibialis anterior muscle preparation8. The tibia was fixed at its upper and lowerends with drills, which were clamped to a myograph stand. The motor nerves were exposedfor stimulation near their points of entry into the muscles. For observations on the soleusthe overlying muscles were removed as far as possible. The skin thus formed a sac, whichwas filled with liquid paraffin heated by a coil containing circulating water from the thermo-statically controlled bath. For experiments on tibialis anterior the overlying skin was notincised and the muscle was warmed by a coil wrapped round the limb and also heated bywater pumped from the thermostatic bath. By these means the temperature of the muscleswas maintained at 37-38° C.

Stimulator. The motor nerves were stimulated through bipolar chlorided silver wireelectrodes by square pulses from an Attree (1950) stimulator. Stimuli were supramaximaland of 0 1-0 2 msec duration. For the study of fusion frequencies a range of rates was usedincluding 4, 8, 12, 16, 20, 25, 30, 40, 50, 63, 87 and 124 per second. Tetani of 200 msecduration were delivered in a rapid sequence 5 sec apart to avoid effects arising from post-tetanic potentiation (Buller et at. 1960). When the effects of D-tubocurarine chloride anddecamethonium iodide were being studied, alternate single shocks and tetani were delivered.Two stimulators were arranged so that one delivered pulses at a low rate (28/min), and theother pulses at a high rate (20/sec). The stimulus pattern was obtained by using an EricsonGS 12D selector tube as a scale-of-three counter to operate a relay. The pulses from the low-rate stimulator were used as input signals to the valve. With the discharge at cathode No. Ithe relay was at rest and a single shock from this stimulator also passed across the relay

INTERCOSTAL MUSCLE CONTRACTIONScontacts to the stimulating electrodes. The occurrence of the next pulse transferred theselector tube discharge to cathode No. 2. This manoeuvre led to the activation of the relay,which switched the stimulating electrodes from the low-rate stimulator to the high-ratestimulator. Thus a tetanic stimulus commenced. A subsequent pulse from the low-ratestimulator transferred the discharge to cathode No. 3, the relay returning to the restingstate and the tetanus ceasing.

RESULTS

Change in length of the intercostal muscle had effects similar to thosefound in other muscles by Buller et at. (1960). An increase in length wasparallelled by an increase in initial tension and in the tension developedduring a contraction. With further increase in length the contractiontension eventually reached a maximum; stretching the muscle beyondthis point resulted in a decrease in contraction tension. In contrast tothe results reported by Buller et al. (1960) the contraction height in thepresent experiments seldom returned to its previous level on lowering theinitial tension. The time to the peak of the contraction and the time tohalf relaxation also increased when the length and initial tension wereraised, and in a few experiments one or other of these two parametersreached a peak simultaneously with the twitch height; these measure-ments were therefore always made with the initial muscle tension adjustedfor the maximum contraction height, as was done by Buller et al. (1960).This initial tension was usually about 75 g, but varied considerably.Similar precautions were observed for the comparative measurementsmade on soleus and tibialis anterior.

Single isometric contractionsThese were recorded from intercostal muscle slips at two different

sites: (1) parasternally, from slips of the internal intercostal muscle alone,the transversus thoracis muscle being dissected away from its under side;(2) further laterally, underlying the insertion of serratus anterior andinvolving contractions of both external and internal intercostal fibreswith, sometimes, the transversus thoracis as well.

Records of contractions at these sites in the 4th left intercostal space areshown in Fig. 2A, B where they can be compared with records from soleusand tibialis anterior (Fig. 2D, E). The similarity of the time parametersof the intercostal slip contractions to those of tibialis anterior should benoted. The time parameters of the single contraction of the different inter-costal muscle slips were all very much the same. There were no significantdifferences between right and left sides, between muscles from differentspaces, or between contractions recorded at the two different sites. Themean time to peak contraction was 33 msec (S.D. + 6 msec; n, 264); andthe mean time to half relaxation was 34 msec (S.D. ± 8 msec; n, 264). Theseresults compare with tibialis anterior, where the time to peak was 27 msec

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(S.D. + 2 msec; n, 12) and the time to half relaxation, 27 msec (S.D. + 2 msec;n, 12). Both intercostal muscles and tibialis anterior contrast with thesoleus, where the time to peak was 86 msec (S.D. + 4 msec; n, 23) and thetime to half relaxation, 90 msec (S.D. + 10 msec; n, 23). The times taken bythe intercostal slips to peak contraction and half relaxation are thereforetypical of 'fast' muscle and are comparable to those of tibialis anterior. Norecording was possible from the parasternal internal intercostal musclealone in the first space, because the external intercostal is continuousthroughout this space.

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Fig. 3. Recordings from a series of tetanic contractions at stimulus frequencies(per second) shown above; and on the right graphs of tetanus: twitch ratio againststimulus interval (abscissa in msec) for the complete series. A, soleus; B, 6th rightparasternal internal intercostal; C, tibialis anterior; D, 4thright internal andexternalintercostal in a 4-week-old kitten. The intervals at half maximal tetanus: twitchratio were (msec) A, 105; B, 35; C, 41; D, 57. Calibrations: tension in grams; tetanusduration shown by t = 200 msec.

Tetanic contractionsThe responses of the muscles at various frequencies were compared,

three contractions at each frequency being recorded. The response tosingle shocks was also recorded three times before and after, and once or

twice during, the tetanic series. The average height of these was used tocalculate the tatanus:twitch ratios. Some typical records from the 6thleft internal intercostal, the tibialis anterior and the soleus muscles are

shown in Fig. 3. With soleus, fusion began at less than 8/sec and was

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complete at 30-40/sec, whereas with tibialis anterior and the intercostalslips fusion did not begin until 16-20/sec and was incomplete up to67-80/sec. Also in Fig. 3 are shown graphs of tetanus: twitch ratios plottedagainst stimulus intervals for the whole series of observations with eachmuscle; every point represents the mean of three observations and thepoints lie along S-shaped curves.

8 20 40 124

380C

3320C

30'20C

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t -20 msecFig. 4. Effect of temperature change on the isometric tetanic contraction of theintercostal muscle slip. Stimulation frequency indicated above and temperatureat the side. Tension in grams; t = 20 msec.

Similar graphs were drawn in each experiment and the stimulus intervalat the half-maximal tetanus: twitch ratio was estimated. The mean intervalfor the intercostal muscles was 36 msec (S.D. + 5 msec; n, 24); for thetibialis anterior it was 41 msec (S.D. + 2; n, 3) and for the soleus, 96 msec(S.D. + 6; n, 4). The mean interval for the intercostal muscles is typical of'fast' muscle and comparable to that of the tibialis anterior.

194 T. J. BISCOE

INTERCOSTAL MUSCLE CONTRACTIONS 195

The effects of cooling on the contraction of intercostal muscleOn cooling, the duration of the single isometric contraction increased

and the maximal contraction tension decreased. Also, the frequency ofstimulation to produce tetanic fusion and the maximal tension decreasedas shown in Fig. 4. The tetanus :twitch ratio was plotted against stimulusinterval for the complete series of observations at each temperature.From these graphs the stimulus intervals at half-maximal tetanus twitch

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Fig. 5. Effects of intravenous D-tubocurarine chloride, A, C and decamethoniumiodide B,Donisometriccontractions ofintercostal muscle, soleus andtibialis anterior.Nerve stimulation was by alternate single shocks and tetani, 20/sec. The gaps in thetraces are of 8 min. Tension in g; time marker 30 sec. A: Tubocurarine, 90 fig/kg;3rd right internal intercostal and soleus muscles. B: Decamethonium, 25 pg/kg;4th right internal and external intercostal and soleus muscles. C: Tubocurarine,85 ug/kg; 3rd right internal intercostal and tibialis anterior muscles. D: Deca-methonium, 12.5 ,ug/kg; 4th right internal and external intercostal and tibialisanterior muscles.

ratio were derived, and their values were plotted against temperature.The interval increased on cooling; this is an indication that tetanic fusionoccurred at a lower stimulus frequency as the temperature fell. For thedata of Fig. 4 this change was highly significant, the regression coefficientbeing - 1-4, for which P < 0-01.

Effect of neuromuscular blocking agentsPaton & Zaimis (1951) showed that fast and slow muscles react dif-

ferently to D-tubocurarine and to decamethonium. The soleus is moresensitive to D-tubocurarine and less sensitive to decamethonium whilstthe reverse is true of the tibialis anterior. The same difference was foundin the present investigation. In severity and time course of the paralysis,the intercostal muscles reacted to intravenous D-tubocurarine chloride inthe same way as tibialis anterior, and unlike soleus (Fig. 5A, C). How-ever, in their sensitivity to decamethonium they appeared to occupy anintermediate position, being much more sensitive than the soleus andsomewhat less sensitive than the tibialis anterior (Fig. 5B, D). It wasnoted that the intercostal muscles became paralysed at the same time asthe other two muscles, which was taken as an indication that their bloodsupply was adequate.

The time relations of the intercostal contraction in the kittenBuller et al. (1960) showed that the single isometric contractions was

slower in a kitten than in an adult cat and that tetanic fusion occurred atlower frequencies. This was confirmed by a small series of experiments on4-week-old kittens. Figures 2 and 3 show the comparison between kittenand adult cat responses. In the record of Fig. 2 the time to peak contractionwas 60 msec and the time to half relaxation was 49 msec, as comparedwith adult mean values of 33 and 34 msec respectively. The intervalbetween stimuli for half the maximal tetanus :twitch ratio in the recordof Fig. 3 was 57 msec, compared with a mean of 36 msec for the adult cat.There was considerable variation in the values for the intercostal muscletime parameters between kittens of the same age. The values for the kittenrecorded above were the lowest found in the present work.

DISCUSSION

The contraction time parameters of the soleus muscle and of the tibialisanterior have been recorded by other workers. For soleus at 37-38°CBuller et al. (1960) reported contraction and half relaxation times of about80 msec and a stimulus interval of about 85 msec for half maximal tetanicfusion. Cooper & Eccles (1930) recorded contraction times of 94, 100 and

196 T. J. BISCOE

INTERCOSTAL MUSCLE CONTRACTIONS120 msec for this muscle at 33-37° C and Wills (1942) gave 77-7 msec forthe same parameter but did not record the muscle temperature. In thework reported in this paper the temperature of the muscles was 37-38° C,the time for contraction was 86 msec and the time for half relaxation was90 msec, while the interval for half-maximal tetanic fusion was 96 msec.Gordon & Holbourn (1949) reported the contraction times of the tibialisanterior, superficial and deep fibres as 28-9 msec and 62-6 msec respec-tivelywhen the cat rectal temperature was 36-37° C. In addition, Gordon &Phillips (1949) found that the time to peak contraction was 18-23 msec at amuscle temperature of 380 C and Wills (1942) reported a time to peak of24*3 msec. The tibialis anterior contraction time reported here, 27 msec,is in reasonable agreement with these figures. The differences in the resultsof various authors may be due to the variety of recording techniques usedand also perhaps to the different muscle temperatures. Thus the techniquesused here on soleus and tibialis anterior give results which are in broadagreement with those of other workers, and it is reasonable to concludethat the intercostal results are also valid.Paton & Zaimis (1951) studied the effects of D-tubocurarine chloride

and decamethonium iodide on respiration and on the contractions of thetibialis anterior and soleus muscles. They concluded from these experi-ments and from the similarity of colour, dark red, that the intercostalmuscles should be classified with soleus rather than with tibialis anterior.Thedemonstrationthat the intercostal muscles have the contraction charac-teristics of 'fast' muscles leads to the opposite conclusion, and the greatereffectiveness of D-tubocurarine in the soleus than in the intercostalmuscles is to be expected.Comparison of the intercostal muscles with the diaphragm is not yet

possible. Sabawala & Dillon (1959) noted that intercostal muscle wasmore sensitive to decamethonium than was rabbit lumbrical or guinea-pigdiaphragm. There are, however, no data, all from one species, about therelative effects on neuromuscular transmission of neuromuscular blockingdrugs in the intercostal, diaphragm and other muscles. In addition, morework on the cat diaphragm, which probably has fast and slow parts(Bottazzi, 1915; Briscoe, 1920; Botha, 1957) is required to elucidate theeffects described by Douglas & Matthews (1952) which are quoted above.

It was of interest to note that the intercostal muscles in the kitten werea light pink colour. In the adult cat they were a darker red and occasionallyas dark as soleus (Paton & Zaimis, 1951). The contraction characteristics,i.e. slow in the kitten and fast in the adult cat, were therefore opposite tothose traditionally expected on the basis of muscle colour (Raniver, 1873).The effect of lowering the temperature was to prolong the time charac-

teristics of contraction, as has been shown by other workers (Gordon &

197

1981.J.BSOPhillips, 1949; Maclagen & Zaimis, 1957). These experiments at subnormaltemperatures, and those on the kitten, served as controls which showedthat with this apparatus a slow contraction could be recorded.These results indicate that the intercostal muscles in the upper seven

spaces in the cat are 'fast' muscles.

SUMMARY

1. In the cat the isometric contraction response to indirect stimulationwas recorded from the soleus, tibialis anterior and the muscles in the upperseven intercostal spaces at two sites, parasternally and further laterally.

2. With the single contraction the times to peak were 85-90 msec forsoleus, 25-30 msec for tibialis anterior and 30-35 msec for the intercostalmuscles. The times to half relaxation were 85-95 msec, 23-30 msec and30-35 msec respectively.

3. The interval between stimuli at half the maximal tetanus:twitchratio was 90-105 msec for soleus, 40-43 msec for tibialis anterior andusually 30-40 msec for the intercostal muscles.

4. D-tubocurarine chloride was less effective as a neuro-muscularblocking drug and decamethonium more effective on the intercostal and ontibialis anterior muscles than on soleus.

5. In the intercostal muscles of the 4-week-old kitten and in the cooledadult cat muscle the times to peak and half relaxation and the intervalat half maximal tetanus :twitch ratio were prolonged.

6. It is concluded that the intercostal muscles are 'fast' muscles in thecat.

Mr B. Aldous gave unfailing technical assistance throughout this work. I am grateful toAllen and Hanburys, Ltd., for the gift of the decamethonium iodide.

REFERENCES

ATTREE, V. H. (1950). An electronic stimulator for biological research. J. 8ci. In8trum.27, 43-47.

BIscoE, T. J. (1961). Some properties of intercostal muscle in the cat. J. Phy8iol. 159,31-32P.

BOTHA, G. S. M. (1957). The anatomy of phrenic nerve termination and the motor inner-vation of the diaphragm. Thorax, 12, 50-56.

BoTTAzzi, F. (1915). Nuove ricerche sui muscoli striati e lisci di animali omeotermi. NotaV: le contrazioni del preparato diaframatico provocate da stimoli unici. R.C. Accad.Lincei, 24, 172-183.

BRiscoE, G. (1920). The muscular mechanism of the diaphragm. J. Phy8iol. 54, 46-53.BULLER, A. J., ECCLES, J. C. & ECCLES, R. M. (1960). Differentiation of fast and slow

muscles in the cat hind limb. J. Physiol. 150, 399-416.COOPER, S. & ECCLES, J. C. (1930). The isometric responses of mammalian muscles. J.

Physiol. 69, 377-385.DOUGLAS, W. W. & MATTEEWS, P. B. C. (1952). Acute tetraethylpyrophosphate poisoning

in cats and its modification by atropine or hyoscine. J. Physiol. 116, 202-218.

198 T. J. BISCOE

INTERCOSTAL MUSCLE CONTRACTIONS 199GORDON, G. & HOLBOURN, A. H. S. (1949). The mechanical activity of single motor units in

reflex contractions of skeletal muscle. J. Physiol. 110, 26-35.GORDON, G. & PHILLIPS, C. G. (1949). Slow and rapid components in a flexor muscle. J.

Physiol. 110, 6-7P.MACLAGAN, J. & ZAnas, E. (1957). The effect of muscle temperature on twitch and tetanus

in the cat. J. Physiol. 137, 89-90P.MARTIN, H. N. & HARTWELL, E. M. (1879). On the respiratory function of the internal inter-

costal muscles. J. Phy8ioi. 2, 24-27.PATON, W. D. M. & ZAIMIS, E. J. (1951). The action of D-tubocurarine and of decamethoniumon respiratory and other muscles in the cat. J. Phy8iol. 112, 311-331.

RANVIER, L. (1873). Proprietes et structures diff6rentes des muscles rouges et des musclesblancs, chez les Lapins et chez les Raies. C.R. Acad. Sci., Pari8, 77, 1030-1034.

SABAWALA, P. H. & DILLON, J. B. (1959). Mode of action of depolarizing agents. Actaanaesth. scand. 3, 83-101.

SCHARPEY-SCHAFER, E. & MACDONALD, A. D. (1925). The action of the intercostal muscles.J. Phy8iol. 60, 25-26P.

TALBOT, S. A., LILIENTHAL, J. L. JR., BESER, J. & REYNOLDS, L. W. (1951). A wide rangemechano-electronic transducer for physiological applications. Rev. Wci. Instrum. 22,233-236.

WILLS, J. H. (1942). Speed of response of various muscles of cats. Amer. J. Physiol. 136,623-628.

Note added in proof. Glebovskii (1961) has recorded the contractiontimes of some respiratory muscles in the cat and in the kitten, using anisometric torsion myograph with a mirror on the spring. For the inter-costal muscles in the 6th, 7th and 8th interspaces of the cat he gives theaverage time to peak contraction as 48 9 msec and the average time tohalf relaxation as 51 msec. These times are considerably longer than thoserecorded in the present work. It is not clear from the paper exactly howthe intercostal muscle temperature was maintained at body temperaturenor is it statedhow the muscle temperature was measured. It is possible thata difference in temperature may account for some of the discrepancy be-tween the two sets of results.GLEBOVSKII, V. D. (1961). Contractile properties of respiratory muscles in fully grown andneonate animals. Sechenov Physiol. J. 47, 470-480.