Electrical and Mechanical Properties ofChick Embryo Heart Chambers

Bxj ROBERT J. BOUCEK, M.D., WILLIAM P. MURPHY, JR. , M.D.. AXD

GEORGE H. PAFP, P H . D .

Electrical and mechanical events of the atrium, ventricle and conus of the 72-hour chickembryo heart were recorded by a specially designed instrument. The electrical-mechanicaldelay and the time of muscular contraction differed among the chambers, the atriumbeing the most rapid and the conus, the slowest. Muscular relaxation times of the atriumand ventricle were approximately one-half that of the eonus. Mechanical pei'formance,gaged by the approximations of work and power, was greatest for the ventricle, but-based on the muscle mass (protein), values for the ventricle and conus were similar.Electrical activation of the ventricle followed a consistent pathway, suggesting theexistence of a preferential conduction system prior to the development of the His bundle.

THE chick embryo heart is well suited forthe study of myocardial function and

metabolism. The electrocardiogram of the in-tact chick embryo and of the isolated hearthas characteristics similar to the hearts ofphylogenetically more advanced species.1"5

Embryonic tissue, such as that obtained fromthe 72-hour chick, in addition to its readyavailability has the further advantage thatthere is no coronary circulation, and fluid,electrolytes and metabolites diffuse directlyinto the myocardial fibers and cells.

The small size of this tissue limits the prac-ticability of its use for the direct observationof fine movement details. This report describesan instrument for the simultaneous recordingand analysis of the electrical and mechanicalevents of the chick embryo heart in each ofthe three chambers.

METHODS

The heart was removed from the 72-hour chickembryo and placed in Tyrode solution; the venousinflow and aortic arches attachments were tran-

From the University of Miami School of Medicineand the Howard Hughes Medical Institute, Miami,Fla.

Supported in part by the National Institutes ofHealth grant-in-aid no. H-3565, grant-in-aid of theHeart Association of Greater Miami, and the Develop-mental Fund of the Section of Cardiology', Universityof Miami School of Medicine, Jackson Memorial Hos-pital, Miami, Fla.

^Received for publication May 6, 1959.

seeted to avoid myocardial injury. After inspec-tion to determine age and activity, the heart wasplaced in a drop of a 1:1 mixture of Tyrodesolution and chicken plasma on a glass slide.

The sensing element is a light, dual purposeplatinum lever, one end of which rests on theheart (fig. 1). The second or indifferent proberests in the plasma drop which supports theheart and serves to complete the circuit for theelectrocardiogram.

The original electrocardiographic signal issmall in comparison with the signal observed inman. Therefore, a two-stage triode push-pullamplification is interposed between the pickupand the recorder amplifier. A gain of 400:1 isattained so that with a recorder with a sensitivityof 1 mv./cm. a total sensitivity of 2 ju.v./cm.may be realized. A thermal and electrical shieldaround the pickup and a circuit with an inherentlow noise level permit a stable, interference-freerecording of the chick heart electrocardiogram.

The opposite end of the lever is isolated elec-trically from the probe and its motion serves toindicate the physical activity of the heart (fig. 1).A differential capacitance sensing circuit operatesfrom a pair of fixed capacitor plates excited inequal and opposite phase by a 30-kc. oscilla-tor. The movable plate (the lever) picks up a30-kc. signal of phase and magnitude relativeto its position between these plates. This signalis identified and amplified in proportion to its mag-nitude and phase and is then recorded simul-taneously with the electrocardiogram on dual tracepaper using a direct writing Sanborn recorder.

The heart was always positioned in the plasmaclot in the same manner and the sensing armplaced in the same location on the atrium, ven-tricle or conus. When records of the ventricle

787 Circulation Research, Volume VII, September 1959

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788 BOUCEK, MURPHY, PAPF

CLOSE-UP OF SENSING DEVICES(Front View)

Chick heart,Sensing arm•*— Indiff. electrode

GLASSSLIDE

INDIFFERENT ELECTRODE

ECG

CHICK HEART MOUNT

( Top View)

FIG. 1. Apparatus for sensing electrical and mechanical properties of the chick embryo heart.

were taken, the sensing arm was placed midwayon the ventricle. Records of isolated chamberswere obtained from hearts which were transectedat the atrioventrieular and ventriculoeonal junc-tions after the heart had been fixed in positionby the plasma clot.

To obtain records of hearts with intact circula-tory systems, the embryo was removed from theegg and placed on a glass slide. A small amountof the ectoderm overlying the heart was removedand records were taken for the ventricle andatrium.

Approximate values of work and power forthe chambers were obtained by recording themovement of a weight (1.96 mg. sensing arm)over a distance (height) in a period of time(recox'ding paper). These approximations were notrepresentative of total work or power since dis-placement of the sensing arm was in one directiononly. Furthermore, the work performed againstthe plasma clot was not included in the calculation.

In the analysis of the electrocardiograms, theconduction times were measured from the recordsobtained with fast paper speed (50 mm./sec).The existence of a standard conduction pathwaywas investigated hy partial transection of theatrioventrieular groove in either direction. Ven-tricular conduction was studied by cutting inter-digitating wedges along the ventricular muscu-lature.

Electromechanical phenomena were followed bymeasuring the time between the onset of elec-trical activity and the beginning of contraction(fig. 2). The time required for contraction andrelaxation was measured directly from the records.

Atria, ventricles and coni were obtained bytransection of 24 hearts and protein content ofthe chambers was determined. The tissues of eachchamber were combined and digested by thestandard micro-Kjeldahl method. Xitrogen wasdetermined by the microdiffusion teehnic ofConway and converted to protein by the con-version factor of 6.25.

The ambient temperature was maintained at29 ± 1 C. by a thermostatically controlled heatingunit within the shielding cabinet since tempera-ture control is essential for the development ofcomparable records.

RESULTS

The mechanical properties of! the ventriclesand coni of 27 isolated intact 72-hour chickembryo hearts were analyzed. The only prop-erty of the atrium which could be measuredwas the electromechanical delay because theeffects of the ventricular contraction distortedthe atrial myogram. Since isolation of theatrium by atrioventricular transection causedno obvious change in its activity, the other

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PHYSIOLOGIC PROPEKTIES OF THE CHICK EMBRYO HEART 789

TABLE 1.—Electromechanical Properties of ChickEmbryo Heart

A. DEPOLARIZATION ELECTRO - MECHANICAL DELAY

B. MAXIMUM HEIGHT

C. MUSCULAR CONTRACTION TIME

D. MUSCULAR RELAXATION TIME

Amp .01 (ECG)

Amp. x 20 (Myogrom)

ATRIAL-

VENTRICULAR -^CONDUCTION 1

ELECTRICAL SYSTOLE

.26t.OI

.09±.003

P- WAVE

.02±.0005

INTRAVENTRICULAR

CONDUCTION

.016±.0007

FIG. 2 Top. Electrical and mechanical activity ofthe chick heart ventricle.

FIG. 3 Bottom. Ventricular electrocardiogram from72-hour chick embryo heart. (Time, seconds; rate82.4 ± 2.2 at 29 C.)

mechanical properties of the atrium were de-rived from the study of 18 isolated atrialpreparations (table 1).

Electromechanical delay varied signifi-cantly among the chambers; the atrium con-sistently had the shortest time lag. The delaywas so characteristic for each of the cham-bers that failure to section the ventriculo-conal area properly could be detected by theshortened time interval and the altered myo-gram, which represented a blend of the ven-tricular and conal muscular contraction.Atrial muscular contraction was more rapid

Atrium

Ventricle

Conus

Ele

ctro

mec

hani

cal

dela

y

.029±.002*

.035±.002

.10

±.006

Time in seconds

Con

trac

tion

.12

±.004

.20

±.006

.24

±.006

Rel

axat

ion

.18

±.008

.16

±.005

.34

±.012

Pow

er(m

m.

mgr

.)

.254±.03

.959±.04

.368±.04

Wor

k(m

m.

mg

,)

.029±.003*

.194±.008

.087±.008

* Mean and standard error of the mean.

than that of ventricular contraction, whichwas faster than that of the conus. The rela-tively slow movements of the conus musclewere reflected in the prolonged relaxationtime (.34 sec). Time for muscular relaxa-tion was the same for the atrium and ventricle.

Mechanical performance, gaged by the ap-proximation for work (mm. nig.) and power

,(mm. mg./sec), differed in the three cham-bers. The greatest amount of work and powerwas developed by the ventricle. When thesevalues were expressed on the basis of theamount of protein, the performances of theventricle and conus were similar (table 1).

Contraction of the ventricle resulted insimilar myograms from the mid- and conalareas of the ventricle. The A'entricular myo-gram near the atrium was smaller in size thanthat obtained from the mid- or conal-ventricu-lar regions.

The electrocardiogram of the chick heartwas of a unipolar nature; the indifferent elec-trode was placed at a distance from the heartwhich did not contribute to the tracing. Itwas composed of a distinct P-wave, P-R inter-val, QRS complex and the T-wave (fig. 3).Records from the isolated hearts were similarto those obtained from the hearts in vivo.Electrical and mechanical records from theatrium, ventricle or conus were easily recog-nizable by their distinctive features (fig. 4).

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Amp. .01 ( ECG)

Amp. x2O ( Myogram)FIG. 4. Electrical and mechanical records of tlie heart chambers (72-hour chick embryo).

Paper Speed 5 0 mm/Second

When the sensing electrode was placed mid-way on the ventricle, the initial electricalforce in 27 intact hearts was directed towardthe electrode, inscribing an initial R-wave onthe electrocardiogram; the dominant forcewas in the opposite direction, producing adeep S-wave. The placement of the sensingarm on the ventricle near the atrium recordedprincipally an S-wave. At the outflow end ofthe ventricle, the record was chiefly an R-wave (fig. 5).

Complete transection of the heart at theatrioventricular and ventriculoconal areas re-sulted in idioatrial and idioventricular rhy-thms. Idioconal rhythms were rarely observedand, when present, were slower than the idio-ventricular rates. Atrial contraction appearedto be similar to that of the intact hearts; how-ever, ventricular behaviour was markedly dif-ferent in the two preparations. In the intacthearts, the rate was 82/min., whereas idio-ventricular rates were 22/min. Electro-mechanical delay for the atrium and ventricleof the completely transected hearts was simi-lar to that of the intact preparations.

Isolated conal tissue differed in its rhythmicproperties from the isolated atrium and ven-

tricle. "When idioconal electrical impulses de-veloped, the rate was exceedingly slow andrarely resulted in a mechanical response.When the mechanical response occurred, ithad the same electromechanical delay as theintact conus. Conal electrocardiograms werebizarre, i.e., prolonged conduction times andslow repolarization time.

Partial transection through the atrioven-tricular sulcus at the greater or lesser curva-ture of the heart produced a significant in-crease in atrioventrieular conduction time;mean value for 10 records was 0.12 ± 0.008.Tntraventricular conduction was also signifi-cantly prolonged by the partial atrioventricu-lar transection. The prolonged atrioventricu-lar conduction time was somewhat more prom-inent when the section was made throughthe lesser curvature. The ventricular electro-cardiograms resembled those of the intactheart when the greater curvature portion waspartially transected while a striking differenceoccurred with the • incomplete transectionthrough the lesser curvature (fig. 6).

Partial transection of the ventricle at threedifferent sites caused an increase in the intra-ventricular conduction time (.03 sec), which

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PHYSIOLOGIC PEOPEKTIBS OF THE CHICK EMBRYO HEART 791

was no greater than that observed when thesectioning had been made in the atrioventricu-lar suleus. Multiple incisions in the ventriclecaused a reorienting of the electrical forces(fig- 6).

DISCUSSIOX

The three chambers of the 72-hour chickembryo heart have properties other than thevariations in the intrinsic rhythms observedby previous investigators.0' 7 Variations in theelectromechanical delay, contraction and re-laxation times, and work and power approxi-mations and the sluggishness of the conal areamay be related to differences in the membranepermeability, to the physicochemical union ofthe contractile proteins or to the metabolicprocesses and their biochemical energetics.

Depolarization in the atrium, ventricle andconus was followed by mechanical activity.However, the electromechanical delay differedin the three chambers; the atrial mechanicalresponse occurred less than .03 sec. after de-polarization while the conus reacted like the"slow" muscle fibers which have been de-scribed in the skeletal muscles of the frog.8

Transection of the ventriculoconal junction ormultiple incisions in the ventricle resulted inoccasional electromechanical dissociation inthe conus, i.e., the recording of an electricalevent without a resultant mechanical contrac-tion. Usually, the electrical event which failedto induce a muscular response was weakerthan the effective electrical impulse. It hasbeen suggested that the coupling of electricaland mechanical activities is related to mem-brane depolarization and ion shift.9 If thiswere true, the electromechanical differences inthe three chambers may be the result of varia-tions in membrane potential and ionic permea-bility.

The progressive lag in the electromechanicalresponse and in the duration of contraction ofthe three chambers serves to facilitate thepumping action of the tubular heart (fig. 7).The short brisk atrial activity is completed asthe ventricular contraction commences andthe sluggishness of the conus effects a patentoutflow tract while the ventricle is emptying.The prolonged eonal contraction produces a

Amp 01 (ECO)

VENTRICLE

ATRIUM

Amp .01 (ECG)Amp. x20(Myogrom)

FIG. 5 Top. Ventricular electrocardiogram with thesensing probe positioned at different areas of theventricle.

FIG. 6 Middle. Ventricular conduction followingpartial atrial ventricular and ventricular transection.

FIG. 7 Bottom. Sequence of chamber contraction.

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792 BOUCEK, MURPHY, PAFP

sphincter-like closure of the outflow tract atthe end of systole. Thus, without valves, littleregurgitation of blood occurs.

The electromotive force which activates themechanical properties of the chambers is pre-dictable in its duration. A pacemaker, locatedin the upper portion of the atrium, controlsthe rate and rhythm of the heart. Followingthe passage of the electrical impulse over theatrium to produce the P-wave of the electro-cardiogram, a pause occurs before its entranceinto the ventricle. This delay, the P-R inter-val, has been classically ascribed to the refrac-toriness of the atrioventricular node. How-ever, no recognizable nodal tissue exists in the72-hour chick embryo heart. Perhaps the ven-tricular muscle has a prolonged refractorinesscaused by an unknown mechanism which re-sults in the atrioventricular conduction delay.

The electrical activation of the ventriclemay occur anywhere along the atrioventricu-lar junction. However, in the majority of rec-ords, electrical excitation followed a preferredpathway which was located at the atrioven-tricular area near the lesser curvature of theheart. From this area, the remainder of theventricle is activated (fig. 5). Conal activationthen follows. This repetitive record of elec-trical forces strongly suggests a conductionpathway for the embryonic heart, a pathwaywhich operates prior to the development of thebundle of His. In mammalian hearts, the an-lage of the His bundle is found near the crestof the developing ventricular septum.10 Noventricular septum is present in the 72-hourchick embryo heart. Further evidence for theembryonal conduction pathway was seen inpartial atrioventricular transection. As longas atrioventricular continuity existed in theregion of the lesser curvature, a small R-wavefollowed by a large S-deflectiou occurred, andthis resembled the record of the intact heart.

SUMMARY

An instrument has been developed whichpermits investigation of the electrical andmechanical events of the atrium, ventricle andconus of the 72-hour chick embryo heart. The

electrical properties are as follows: The pace-maker is located high in the atrium. Atrio-ventricular delay occurs in the absence ofnodal tissue. A preferential pathway of ven-tricular conduction antedates the develop-ment of the His bundle.

The mechanical properties are as follows:Electromechanical delay and contraction timeare shortest in the atrium and longest in theconus. Relaxation time is similar in the atriumand ventricle and approximately one-half thatof the conus. Based on muscle mass (protein),the ventricle and conus have comparable workand power approximations.

ACKNOWLEDGMENT

The authors gratefully acknowledge the technicalassistance of Mr. Benjamin Brauzer.

SUMMARIO IN INTERLINGUA

Esseva disveloppate un instrumento quepermitte le investigation del eventos electric emechanic in le atrio, le ventriculo, e le conoin le corde de embryones de gallina de 72horas de etate. Le proprietates electric es lesequentes: Le pacemaker es locate alte in leatrio. II occurre un retardo atrioventricular acausa del absentia de histo nodal. Un via pre-ferential de conduction ventricular precedele disveloppamento del fasce de His.

Le proprietates mechanic es le sequentes:Le retardo electromechanic e le tempore decoutractiones le plus breve in le atrio e le pluslouge in le eono. Le tempore de relaxation essimile in atrio e ventriculo. In le cono, illoes approximativemente duo vices plus longe.Calculate super le base de massa muscular(proteina), le ventriculo e le cono ha approxi-mativemente le mesme carga de labor e lemesine potentia.

REFERENCES1. AVERTHEIM-SALOJIONSOST, J. K. A.: Das

elcktrokardiogramm von huhnerembryonen.Pfluger's Arch. f. d. ges. Physiol. 153: 553,1913.

2. KCLBS, ¥.: Experimenti'Ile untersuchunjjenam huhnerembryo. Cremer's Beitr. z. Phy-siol. 1: 439,1920.

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3. LUEG, W., AND HOFER, K.: Elektrokardio-gramme von embryonalen hiihnerherzen inder gewebeskulturen bei gleichzeitiger kine-matographien des bewegungsablaufs. Deu-tsch. med. Wchnschr. 59: 452, 1933.

4. KATSONUMA, S., AND INADA, G.: tiber elek-trokardiogramme von simultaner herzmus-kelkontraktion in einem gewebekultur me-dium. Nagoya J. M. Sc. 7: 53, 1933.

5. POLLACK, H.: Electrocardiographic studieson chick embryo hearts. I. A technic forrecording electrical changes in isolatedchick embryo hearts. J. Lab. & Clin. Med.16:1194,1931.

6. PATTEN, B. M., AND KRAMER, T. C : Theinitiation of contraction in the embryonic

chick heart. Am. J. Anat. 53: 349, 1933.7. PAPF, G. H.: Conclusive evidence for sino-

atrial dominance in isolated 4S-hour ern-bryonic chick hearts cultivated in vitro.Anat. Ree. 63: 203,1935.

S. KoFFLER, S. W., AND WILLIAMS, E . M. :Properties of the "slow" skeletal musclefibres of the frog. J. Physiol. 121: 31S,1953.

9. BOTTS, J.: The triggering of contraction inskeletal muscle. Lecture and Review SeriesNo. 57-1. Naval Medical Research Institute,National Naval Medical Center, Bethesda,Ma., January 15, 1957.

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ROBERT J. BOUCEK, WILLIAM P. MURPHY, JR. and GEORGE H. PAFFElectrical and Mechanical Properties of Chick Embryo Heart Chambers

Print ISSN: 0009-7330. Online ISSN: 1524-4571 Copyright © 1959 American Heart Association, Inc. All rights reserved.is published by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231Circulation Research

doi: 10.1161/01.RES.7.5.7871959;7:787-793Circ Res. 

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