Download Electrical and Mechanical Properties of Chick Embryo Heart

Electrical and Mechanical Properties of
Chick Embryo Heart Chambers
Bxj ROBERT J. BOUCEK, M.D.,
WILLIAM P. MURPHY, J R . , M.D.. AXD
GEORGE H. PAFP, P H . D .
Electrical and mechanical events of the atrium, ventricle and conus of the 72-hour chick
embryo heart were recorded by a specially designed instrument. The electrical-mechanical
delay and the time of muscular contraction differed among the chambers, the atrium
being the most rapid and the conus, the slowest. Muscular relaxation times of the atrium
and 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, butbased on the muscle mass (protein), values for the ventricle and conus were similar.
Electrical activation of the ventricle followed a consistent pathway, suggesting the
existence of a preferential conduction system prior to the development of the His bundle.
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T
seeted to avoid myocardial injury. After inspection to determine age and activity, the heart was
placed in a drop of a 1:1 mixture of Tyrode
solution and chicken plasma on a glass slide.
The sensing element is a light, dual purpose
platinum lever, one end of which rests on the
heart (fig. 1). The second or indifferent probe
rests in the plasma drop which supports the
heart and serves to complete the circuit for the
electrocardiogram.
The original electrocardiographic signal is
small in comparison with the signal observed in
man. Therefore, a two-stage triode push-pull
amplification is interposed between the pickup
and the recorder amplifier. A gain of 400:1 is
attained so that with a recorder with a sensitivity
of 1 mv./cm. a total sensitivity of 2 ju.v./cm.
may be realized. A thermal and electrical shield
around the pickup and a circuit with an inherent
low noise level permit a stable, interference-free
recording of the chick heart electrocardiogram.
The opposite end of the lever is isolated electrically from the probe and its motion serves to
indicate the physical activity of the heart (fig. 1).
A differential capacitance sensing circuit operates
from a pair of fixed capacitor plates excited in
equal and opposite phase by a 30-kc. oscillator. The movable plate (the lever) picks up a
30-kc. signal of phase and magnitude relative
to its position between these plates. This signal
is identified and amplified in proportion to its magnitude and phase and is then recorded simultaneously with the electrocardiogram on dual trace
paper using a direct writing Sanborn recorder.
The heart was always positioned in the plasma
clot in the same manner and the sensing arm
placed in the same location on the atrium, ventricle or conus. When records of the ventricle
HE chick embryo heart is well suited for
the study of myocardial function and
metabolism. The electrocardiogram of the intact chick embryo and of the isolated heart
has characteristics similar to the hearts of
phylogenetically more advanced species.1"5
Embryonic tissue, such as that obtained from
the 72-hour chick, in addition to its ready
availability has the further advantage that
there is no coronary circulation, and fluid,
electrolytes and metabolites diffuse directly
into the myocardial fibers and cells.
The small size of this tissue limits the practicability of its use for the direct observation
of fine movement details. This report describes
an instrument for the simultaneous recording
and analysis of the electrical and mechanical
events of the chick embryo heart in each of
the three chambers.
METHODS
The heart was removed from the 72-hour chick
embryo and placed in Tyrode solution; the venous
inflow and aortic arches attachments were tranFrom the University of Miami School of Medicine
and the Howard Hughes Medical Institute, Miami,
Fla.
Supported in part by the National Institutes of
Health grant-in-aid no. H-3565, grant-in-aid of the
Heart Association of Greater Miami, and the Developmental Fund of the Section of Cardiology', University
of Miami School of Medicine, Jackson Memorial Hospital, Miami, Fla.
^Received for publication May 6, 1959.
787
Circulation Research, Volume VII, September 1959
BOUCEK, MURPHY, PAPF
788
CLOSE-UP OF SENSING DEVICES
(Front View)
Sensing arm
•*— Indiff. electrode
Chick heart,
GLASS
SLIDE
INDIFFERENT ELECTRODE
ECG
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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 midway
on the ventricle. Records of isolated chambers
were obtained from hearts which were transected
at the atrioventrieular and ventriculoeonal junctions after the heart had been fixed in position
by the plasma clot.
To obtain records of hearts with intact circulatory systems, the embryo was removed from the
egg and placed on a glass slide. A small amount
of the ectoderm overlying the heart was removed
and records were taken for the ventricle and
atrium.
Approximate values of work and power for
the chambers were obtained by recording the
movement of a weight (1.96 mg. sensing arm)
over a distance (height) in a period of time
(recox'ding paper). These approximations were not
representative of total work or power since displacement of the sensing arm was in one direction
only. Furthermore, the work performed against
the plasma clot was not included in the calculation.
In the analysis of the electrocardiograms, the
conduction times were measured from the records
obtained with fast paper speed (50 mm./sec).
The existence of a standard conduction pathway
was investigated hy partial transection of the
atrioventrieular groove in either direction. Ventricular conduction was studied by cutting interdigitating wedges along the ventricular musculature.
Electromechanical phenomena were followed by
measuring the time between the onset of electrical activity and the beginning of contraction
(fig. 2). The time required for contraction and
relaxation was measured directly from the records.
Atria, ventricles and coni were obtained by
transection of 24 hearts and protein content of
the chambers was determined. The tissues of each
chamber were combined and digested by the
standard micro-Kjeldahl method. Xitrogen was
determined by the microdiffusion teehnic of
Conway and converted to protein by the conversion factor of 6.25.
The ambient temperature was maintained at
29 ± 1 C. by a thermostatically controlled heating
unit within the shielding cabinet since temperature control is essential for the development of
comparable records.
RESULTS
The mechanical properties of! the ventricles
and coni of 27 isolated intact 72-hour chick
embryo hearts were analyzed. The only property of the atrium which could be measured
was the electromechanical delay because the
effects of the ventricular contraction distorted
the atrial myogram. Since isolation of the
atrium by atrioventricular transection caused
no obvious change in its activity, the other
PHYSIOLOGIC PROPEKTIES OF THE CHICK EMBRYO HEART
789
TABLE 1.—Electromechanical Properties of Chick
Embryo Heart
DEPOLARIZATION
ELECTRO - MECHANICAL
DELAY
Conus
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B.
MAXIMUM
C.
MUSCULAR
HEIGHT
CONTRACTION
D.
MUSCULAR
RELAXATION
TIME
.254
±.03
.029
±.003*
.959
±.04
.194
±.008
.368
±.04
.087
±.008
.12
±.004
±.008
.035
±.002
.20
.16
±.006
±.005
.10
Work
(mm. mg,)
.18
.029
±.002*
±.006
TIME
Power
(mm. mgr.)
Ventricle
A.
Relaxation
Atrium
Contraction
Electro
mechanical
delay
Time in seconds
.24
.34
±.006
±.012
* Mean and standard error of the mean.
Amp .01 (ECG)
Amp. x 20 (Myogrom)
ELECTRICAL SYSTOLE
ATRIALVENTRICULAR - ^
CONDUCTION
1
.26
t.OI
.09
±.003
P- WAVE
.02
±.0005
INTRAVENTRICULAR
CONDUCTION
.016
±.0007
FIG. 2 Top. Electrical and mechanical activity of
the chick heart ventricle.
FIG. 3 Bottom. Ventricular electrocardiogram from
72-hour chick embryo heart. (Time, seconds; rate
82.4 ± 2.2 at 29 C.)
mechanical properties of the atrium were derived from the study of 18 isolated atrial
preparations (table 1).
Electromechanical delay varied significantly among the chambers; the atrium consistently had the shortest time lag. The delay
was so characteristic for each of the chambers that failure to section the ventriculoconal area properly could be detected by the
shortened time interval and the altered myogram, which represented a blend of the ventricular and conal muscular contraction.
Atrial muscular contraction was more rapid
than that of ventricular contraction, which
was faster than that of the conus. The relatively slow movements of the conus muscle
were reflected in the prolonged relaxation
time (.34 sec). Time for muscular relaxation was the same for the atrium and ventricle.
Mechanical performance, gaged by the approximation for work (mm. nig.) and power
,(mm. mg./sec), differed in the three chambers. The greatest amount of work and power
was developed by the ventricle. When these
values were expressed on the basis of the
amount of protein, the performances of the
ventricle and conus were similar (table 1).
Contraction of the ventricle resulted in
similar myograms from the mid- and conal
areas of the ventricle. The A'entricular myogram near the atrium was smaller in size than
that obtained from the mid- or conal-ventricular regions.
The electrocardiogram of the chick heart
was of a unipolar nature; the indifferent electrode was placed at a distance from the heart
which did not contribute to the tracing. It
was composed of a distinct P-wave, P-R interval, QRS complex and the T-wave (fig. 3).
Records from the isolated hearts were similar
to those obtained from the hearts in vivo.
Electrical and mechanical records from the
atrium, ventricle or conus were easily recognizable by their distinctive features (fig. 4).
BOUCEK, MURPHY, PAFP
790
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FIG. 4. Electrical and mechanical records of tlie heart chambers (72-hour chick embryo).
When the sensing electrode was placed midway on the ventricle, the initial electrical
force in 27 intact hearts was directed toward
the electrode, inscribing an initial R-wave on
the electrocardiogram; the dominant force
was in the opposite direction, producing a
deep S-wave. The placement of the sensing
arm on the ventricle near the atrium recorded
principally an S-wave. At the outflow end of
the ventricle, the record was chiefly an Rwave (fig. 5).
Complete transection of the heart at the
atrioventricular and ventriculoconal areas resulted in idioatrial and idioventricular rhythms. Idioconal rhythms were rarely observed
and, when present, were slower than the idioventricular rates. Atrial contraction appeared
to be similar to that of the intact hearts; however, ventricular behaviour was markedly different in the two preparations. In the intact
hearts, the rate was 82/min., whereas idioventricular rates were 22/min. Electromechanical delay for the atrium and ventricle
of the completely transected hearts was similar to that of the intact preparations.
Isolated conal tissue differed in its rhythmic
properties from the isolated atrium and ven-
tricle. "When idioconal electrical impulses developed, the rate was exceedingly slow and
rarely resulted in a mechanical response.
When the mechanical response occurred, it
had the same electromechanical delay as the
intact conus. Conal electrocardiograms were
bizarre, i.e., prolonged conduction times and
slow repolarization time.
Partial transection through the atrioventricular sulcus at the greater or lesser curvature of the heart produced a significant increase in atrioventrieular conduction time;
mean value for 10 records was 0.12 ± 0.008.
Tntraventricular conduction was also significantly prolonged by the partial atrioventricular transection. The prolonged atrioventricular conduction time was somewhat more prominent when the section was made through
the lesser curvature. The ventricular electrocardiograms resembled those of the intact
heart when the greater curvature portion was
partially transected while a striking difference
occurred with the • incomplete transection
through the lesser curvature (fig. 6).
Partial transection of the ventricle at three
different sites caused an increase in the intraventricular conduction time (.03 sec), which
PHYSIOLOGIC PEOPEKTIBS OF THE CHICK EMBRYO HEART
791
was no greater than that observed when the
sectioning had been made in the atrioventricular suleus. Multiple incisions in the ventricle
caused a reorienting of the electrical forces
(fig- 6).
DISCUSSIOX
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The three chambers of the 72-hour chick
embryo heart have properties other than the
variations in the intrinsic rhythms observed
by previous investigators.0' 7 Variations in the
electromechanical delay, contraction and relaxation times, and work and power approximations and the sluggishness of the conal area
may be related to differences in the membrane
permeability, to the physicochemical union of
the contractile proteins or to the metabolic
processes and their biochemical energetics.
Depolarization in the atrium, ventricle and
conus was followed by mechanical activity.
However, the electromechanical delay differed
in the three chambers; the atrial mechanical
response occurred less than .03 sec. after depolarization while the conus reacted like the
"slow" muscle fibers which have been described in the skeletal muscles of the frog.8
Transection of the ventriculoconal junction or
multiple incisions in the ventricle resulted in
occasional electromechanical dissociation in
the conus, i.e., the recording of an electrical
event without a resultant mechanical contraction. Usually, the electrical event which failed
to induce a muscular response was weaker
than the effective electrical impulse. It has
been suggested that the coupling of electrical
and mechanical activities is related to membrane depolarization and ion shift.9 If this
were true, the electromechanical differences in
the three chambers may be the result of variations in membrane potential and ionic permeability.
The progressive lag in the electromechanical
response and in the duration of contraction of
the three chambers serves to facilitate the
pumping action of the tubular heart (fig. 7).
The short brisk atrial activity is completed as
the ventricular contraction commences and
the sluggishness of the conus effects a patent
outflow 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 the
sensing probe positioned at different areas of the
ventricle.
FIG. 6 Middle. Ventricular conduction following
partial atrial ventricular and ventricular transection.
FIG. 7 Bottom. Sequence of chamber contraction.
792
BOUCEK, MURPHY, PAFP
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sphincter-like closure of the outflow tract at
the end of systole. Thus, without valves, little
regurgitation of blood occurs.
The electromotive force which activates the
mechanical properties of the chambers is predictable in its duration. A pacemaker, located
in the upper portion of the atrium, controls
the rate and rhythm of the heart. Following
the passage of the electrical impulse over the
atrium to produce the P-wave of the electrocardiogram, a pause occurs before its entrance
into the ventricle. This delay, the P-R interval, has been classically ascribed to the refractoriness of the atrioventricular node. However, no recognizable nodal tissue exists in the
72-hour chick embryo heart. Perhaps the ventricular muscle has a prolonged refractoriness
caused by an unknown mechanism which results in the atrioventricular conduction delay.
The electrical activation of the ventricle
may occur anywhere along the atrioventricular junction. However, in the majority of records, electrical excitation followed a preferred
pathway which was located at the atrioventricular area near the lesser curvature of the
heart. From this area, the remainder of the
ventricle is activated (fig. 5). Conal activation
then follows. This repetitive record of electrical forces strongly suggests a conduction
pathway for the embryonic heart, a pathway
which operates prior to the development of the
bundle of His. In mammalian hearts, the anlage of the His bundle is found near the crest
of the developing ventricular septum.10 No
ventricular septum is present in the 72-hour
chick embryo heart. Further evidence for the
embryonal conduction pathway was seen in
partial atrioventricular transection. As long
as atrioventricular continuity existed in the
region of the lesser curvature, a small R-wave
followed by a large S-deflectiou occurred, and
this resembled the record of the intact heart.
electrical properties are as follows: The pacemaker is located high in the atrium. Atrioventricular delay occurs in the absence of
nodal tissue. A preferential pathway of ventricular conduction antedates the development of the His bundle.
The mechanical properties are as follows:
Electromechanical delay and contraction time
are shortest in the atrium and longest in the
conus. Relaxation time is similar in the atrium
and ventricle and approximately one-half that
of the conus. Based on muscle mass (protein),
the ventricle and conus have comparable work
and power approximations.
ACKNOWLEDGMENT
The authors gratefully acknowledge the technical
assistance of Mr. Benjamin Brauzer.
SUMMARIO IN INTERLINGUA
Esseva disveloppate un instrumento que
permitte le investigation del eventos electric e
mechanic in le atrio, le ventriculo, e le cono
in le corde de embryones de gallina de 72
horas de etate. Le proprietates electric es le
sequentes: Le pacemaker es locate alte in le
atrio. II occurre un retardo atrioventricular a
causa del absentia de histo nodal. Un via preferential de conduction ventricular precede
le disveloppamento del fasce de His.
Le proprietates mechanic es le sequentes:
Le retardo electromechanic e le tempore de
coutractiones le plus breve in le atrio e le plus
louge in le eono. Le tempore de relaxation es
simile in atrio e ventriculo. In le cono, illo
es approximativemente duo vices plus longe.
Calculate super le base de massa muscular
(proteina), le ventriculo e le cono ha approximativemente le mesme carga de labor e le
mesine potentia.
REFERENCES
1. AVERTHEIM-SALOJIONSOST,
SUMMARY
An instrument has been developed which
permits investigation of the electrical and
mechanical events of the atrium, ventricle and
conus of the 72-hour chick embryo heart. The
J.
K.
A.:
Das
elcktrokardiogramm von huhnerembryonen.
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1913.
2. KCLBS, ¥.:
Experimenti'Ile untersuchunjjen
am huhnerembryo. Cremer's Beitr. z. Physiol. 1: 439,1920.
PHYSIOLOGIC PROPERTIES OF THE CHICK EMBRYO HEART
3. LUEG, W., AND HOFER, K.:
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gramme von embryonalen hiihnerherzen in
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4. KATSONUMA, S., AND INADA, G.: tiber elektrokardiogramme von simultaner herzmuskelkontraktion in einem gewebekultur medium. Nagoya J. M. Sc. 7: 53, 1933.
5. POLLACK, H.: Electrocardiographic studies
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recording electrical changes in isolated
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16:1194,1931.
6. PATTEN, B. M., AND KRAMER, T. C :
793
chick heart. Am. J. Anat. 53: 349, 1933.
7. PAPF, G. H.: Conclusive evidence for sinoatrial dominance in isolated 4S-hour ernbryonic chick hearts cultivated in vitro.
Anat. Ree. 63: 203,1935.
S. KoFFLER,
S. W., AND WILLIAMS, E .
M. :
Properties of the "slow" skeletal muscle
fibres of the frog. J. Physiol. 121: 31S,
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9. BOTTS, J.: The triggering of contraction in
skeletal muscle. Lecture and Review Series
No. 57-1. Naval Medical Research Institute,
National Naval Medical Center, Bethesda,
Ma., January 15, 1957.
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The
10. MALL, F. P.: On the development of the
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Electrical and Mechanical Properties of Chick Embryo Heart Chambers
ROBERT J. BOUCEK, WILLIAM P. MURPHY, JR. and GEORGE H. PAFF
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Circ Res. 1959;7:787-793
doi: 10.1161/01.RES.7.5.787
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Copyright © 1959 American Heart Association, Inc. All rights reserved.
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