Download Parasympathetic control of the heart. I. An interventriculo

J Appl Physiol 96: 2265–2272, 2004.
First published February 20, 2004; 10.1152/japplphysiol.00620.2003.
Parasympathetic control of the heart. I. An interventriculo-septal ganglion is
the major source of the vagal intracardiac innervation of the ventricles
Tannis A. Johnson,1 Alrich L. Gray,1 Jean-Marie Lauenstein,1 Stephen S. Newton,1 and V. John Massari1,2
1
Department of Pharmacology and 2Specialized Neuroscience Research Program,
Howard University College of Medicine, Washington, District of Columbia 20059
Submitted 16 June 2003; accepted in final form 25 September 2003
cardiovascular research, we have reexamined the distribution
and counted the number of neurons contained within intracardiac ganglia throughout the cat heart.
Activation of the vagus nerve can cause substantial effects
on ventricular functions. Thus, for example, increased vagal
tone can 1) decrease ventricular contractility (24), 2) prolong
ventricular refractoriness (38), and 3) terminate some types of
ventricular arrhythmias in humans (15). The origins of the
vagal postganglionic neurons that innervate the ventricles,
however, are poorly understood. Previous physiological data
from this laboratory have suggested that a left ventricular
cranioventricular (CV) ganglion selectively mediated negative
inotropic effects of vagal stimulation on the left ventricle
without influencing negative chronotropic or negative dromotropic actions (17, 20). These data imply that the axons of the
CV ganglion should selectively project to the walls of the left
ventricle. Furthermore, other ventricular ganglia should exist
that innervate the walls of the right ventricle. We have tested
these hypotheses by using a fluorescent retrograde tracing
method to determine the source(s) of the vagal innervation of
the right and left ventricular walls.
MATERIALS AND METHODS
substantial interest in the neuroanatomic innervation of the heart for a considerable period of time
(25), the locations, projections, and functions of the intracardiac ganglia are incompletely understood. The number of
ganglia that have been characterized in the heart varies across
species and ranges from as low as 4 in the rat (9, 27) to as many
as 10 in humans (5, 32). Although many intracardiac ganglia
are found in the atria, ganglia can also be found on the
ventricular surface of the heart in diverse species, including
humans (5, 28); dogs (4); cows, sheep, deer, and porpoises
(14); rabbits and monkeys (25); and pigs (6). In the cat, it was
suggested that intracardiac ganglia are found exclusively
within the atria (10); however, in a recent study (20), it has
been shown that at least one ventricular ganglion is found in
the cat heart. Because the cat is an important animal model in
All experiments were reviewed and approved by the Institutional
Animal Care and Use Committee of Howard University in accordance
with guidelines established by the National Institutes of Health for the
humane treatment of animals.
Four adult cats (2–4 kg) were used to determine the topography of
intracardiac ganglia. The animals were deeply anesthetized with
pentobarbital sodium (55 mg/kg iv) and perfused sequentially via a
cannula inserted retrogradely in the abdominal aorta with 1 liter of 0.1
M PBS (pH 7.4) containing heparin sulfate (2,500 U) and 3 liters of
4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.4). There were
no readily apparent differences in the gross anatomy of the cat hearts
that were examined. The hearts were removed, postfixed in the same
fixative for 1 h, and then transferred to 30% sucrose in phosphate
buffer for 48 h. The hearts were subsequently frozen by immersion in
isopentane chilled to a slurry over liquid nitrogen, embedded in
optimum-cutting temperature (OCT) compound (Pella, Redding, CA),
and cut in a cryostat into 50-␮m sections. One series of transverse
sections was mounted onto gelatinized slides and stained with hematoxylin and eosin. A second series of sections from each heart was
processed for the immunocytochemical visualization of the general
neuronal marker protein gene product 9.5 (PGP 9.5) using a Vectastain Elite ABC kit and rabbit anti-PGP 9.5 antibody (Biogenesis)
diluted 1:2,000. A third series was unstained. Tissues were dehydrated
with alcohols, cleared in xylene, coverslipped with Permount, and
examined in a Nikon FXA photomicroscope equipped with bright-
Address for reprint requests and other correspondence: V. J. Massari, Dept.
of Pharmacology, Howard Univ. College of Medicine, 520 W St., N.W.,
Washington, DC 20059 (E-mail: [email protected]).
The costs of publication of this article were defrayed in part by the payment
of page charges. The article must therefore be hereby marked “advertisement”
in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
autonomic control; cardioinhibitory; intracardiac ganglia; parasympathetic nervous system; vagus nerve
ALTHOUGH THERE HAS BEEN
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Johnson, Tannis A., Alrich L. Gray, Jean-Marie Lauenstein,
Stephen S. Newton, and V. John Massari. Parasympathetic control
of the heart. I. An interventriculo-septal ganglion is the major source
of the vagal intracardiac innervation of the ventricles. J Appl Physiol
96: 2265–2272, 2004. First published February 20, 2004; 10.1152/
japplphysiol.00620.2003.—The locations, projections, and functions
of the intracardiac ganglia are incompletely understood. Immunocytochemical labeling with the general neuronal marker protein gene
product 9.5 (PGP 9.5) was used to determine the distribution of
intracardiac neurons throughout the cat atria and ventricles. Fluorescence microscopy was used to determine the number of neurons
within these ganglia. There are eight regions of the cat heart that
contain intracardiac ganglia. The numbers of neurons found within
these intracardiac ganglia vary dramatically. The total number of
neurons found in the heart (6,274 ⫾ 1,061) is almost evenly divided
between the atria and the ventricles. The largest ganglion is found in
the interventricular septum (IVS). Retrogradely labeled fluorescent
tracer studies indicated that the vagal intracardiac innervation of the
anterior surface of the right ventricle originates predominantly in the
IVS ganglion. A cranioventricular (CV) ganglion was retrogradely
labeled from the anterior surface of the left ventricle but not from the
anterior surface of the right ventricle. These new neuroanatomic data
support the prior physiological hypothesis that the CV ganglion in the
cat exerts a negative inotropic effect on the left ventricle. A total of
three separate intracardiac ganglia innervate the left ventricle, i.e., the
CV, IVS, and a second left ventricular (LV2) ganglion. However, the
IVS ganglion provides the major source of innervation to both the left
and right ventricles. This dual innervation pattern may help to coordinate or segregate vagal effects on left and right ventricular performance.
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PARASYMPATHETIC CONTROL OF THE LEFT AND RIGHT VENTRICLES
otic procaine penicillin G (30,000 U/kg sc) daily. Animals were killed
under deep barbiturate anesthesia (pentobarbital sodium, 55 mg/kg) by
intravascular perfusion with fixatives after a 3-day survival time as
described above. Every third 50-␮m transverse frozen section of the heart
taken from the aortic arch to within 3 mm of the apex of the ventricle was
mounted on gelatinized slides, coverslipped with 50% aqueous glycerol,
and examined using FITC fluorescence optics. Only neurons with an
identifiable nucleus (⫻200 magnification) were counted. Cell counts
were multiplied by three to compensate for examination of every third
section. Retrogradely labeled cells were identified by the presence of a
yellow nucleus and a clear cytoplasm. For each ganglion, we determined
the total number of neurons and the number of retrogradely labeled
neurons. Statistical analyses of the data were performed using a one-way
ANOVA. Homogeneity of variance was evaluated using the Levene test
(23), and multiple comparisons were evaluated using Tukey’s multiplerange test when group variances were equal or Dunnett’s T3 post hoc test
when group variances were unequal (22, 33).
RESULTS
Microscopic analysis of serial sections through the heart
immunocytochemically stained with an antibody to the general
Fig. 1. Schematic illustrations of the locations (A–H) of intracardiac ganglia in the cat. L1–L8, 8 representative levels of the heart. A,
posterior atrial ganglion; B, sinoatrial ganglion; C, atrioventricular ganglion; D, cranioventricular ganglion; E, right ventricular ganglion;
F, interatrio-septal ganglion; G, interventriculo-septal ganglion; H, left ventricular ganglion 2; I, sporadic neurons were sometimes
observed here; aa, aortic arch; gcv, great coronary vein; ivc, inferior vena cava; la, left atrium; lau, left auricle; lca, left anterior descending
coronary artery; lv, left ventricle; paa, pulmonary arteries; ra, right atrium; rau, right auricle; rv, right ventricle; svc, superior vena cava.
*Locations of neurons within the intracardiac ganglia.
J Appl Physiol • VOL
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field, dark-field, and Nomarski differential interference contrast optics. The locations of intracardiac ganglia were plotted onto drawings
of transverse sections taken through eight representative levels of the
heart (Fig. 1).
Experiments to determine the origin(s) of ganglia innervating the
ventricles were performed on nine cats of either sex. Cats were sedated
with a mixture of ketamine (22 mg/kg im) and acepromazine (0.22 mg/kg
im) and given atropine (0.05 mg/kg im) to reduce respiratory secretions.
Anesthesia was induced and maintained with isoflurane by inhalation.
Subsequently, the animal was intubated with a cuffed endotracheal tube,
and a thoracotomy was performed. Positive end-expiratory pressure
artificial respiration was maintained thereafter. Body temperature was
maintained at 37°C with warming pads. The pericardium was incised and
carefully retracted. In each of eight animals, a total of 10 ␮l of the
retrograde tracer diamidino yellow (1.0% in ethylene glycol containing
2% DMSO) was injected into four sites on the anterior surface of either
the left or right ventricle using a Hamilton syringe (Fig. 2). In a control
animal, 10 ␮l of diamidino yellow was injected into the pericardial space
but not into the myocardium. The pericardium was then closed, and the
muscles and skin were sutured in layers. Before the end of surgery, all
animals received torbugesic (0.2 mg/kg im) for pain. Postoperatively, all
cats received the analgesic drug ketoprofen (1 mg/kg sc) and the antibi-
PARASYMPATHETIC CONTROL OF THE LEFT AND RIGHT VENTRICLES
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neuronal marker PGP 9.5 (36) revealed eight separate regions
that consistently contained multiple closely associated clusters
of neurons that we have referred to as ganglia (Figs. 1 and 3).
Two ganglia were found on the right atrium [the posterior atrial
(PA) and sinoatrial (SA) ganglia], one ganglion was found on
the left atrium [the atrioventricular (AV) ganglion], one ganglion was found within the interatrial septum [the interatrioseptal (IAS) ganglion], one ganglion was found on the right
ventricle [the right ventricular (RV) ganglion], two ganglia
were found on the left ventricle (the CV ganglion and the LV2
ganglion), and one ganglion was found within the interventricular septum [the interventriculo-septal (IVS) ganglion]. Most
of the ganglia were found within fatty and connective tissues
between the surface of the heart and cardiac muscle, or surrounding the great vessels, and were in close proximity to
nerve bundles. In some cases, however, particularly within the
IVS ganglion, substantial numbers of neurons were found
embedded in areas of myocardium that were completely surrounded by cardiac muscle cells and had little or no surrounding fatty matrix.
J Appl Physiol • VOL
The PA ganglion is found in a fat pad located between the
superior vena cava and the aorta on the posterior medial aspect
of the right atrium. It is one of the largest ganglia on the heart.
The PA ganglion is also the most rostrally located intrinsic
cardiac ganglion. It begins just superior to the common pulmonary artery and ends at the superior aspect of the left atrium.
In cross section, the PA ganglion is found ventral to the
pulmonary artery and dorsal to the aorta and right atrium. The
PA ganglion is comprised of large teardrop-shaped or small
linear clusters of neurons widely separated and completely
surrounded by fatty connective tissue. Clusters may be composed of as many as 60 neurons.
The SA and AV ganglia are located within fat pads on the
surfaces of the atria. The SA ganglion is found on the right
atrium located at the junction of the right pulmonary veins and
the superior vena cava. The cells in this ganglion lie adjacent
to the atrial myocardium and are oriented in a columnar or
teardrop-shape fashion along the external border of the atrial
myocardium. Occasionally, small ganglia can be found within
the atrial myocardium. The outer border of the ganglion is
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Fig. 2. Schematic illustrations of the locations (*) of retrograde
tracer injections into the right ventricle (A) or left ventricle (B)
of the cat heart. Abbreviations as in legend to Fig. 1.
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comprised of fatty tissue (21). The AV ganglion is located at
the junction of the inferior vena cava and the inferior left
atrium. The cells in this ganglion are found in several oval
clusters dispersed in connective and fatty tissue (21).
The CV ganglion is found in a fat pad located at the
cranioventricular margin of the left ventricle. The rostral aspect
of the ganglion originates at the ventral left lateral aspect of the
conus trunci. Caudally, this ganglion begins to migrate dorsally
toward the dorsal left lateral aspect of the right ventricle. The
CV ganglion is found in a large mass of fatty tissue containing
scattered clusters of ganglia comprised of a few cells (18, 20).
The RV ganglion is a relatively small ventricular ganglion
located on the rostral right lateral aspect of the conus trunci and
is found in small groups of four to five cell clusters embedded
in fatty connective tissue. The IVS ganglion is the largest
ventricular ganglion. It is found deeply embedded in the
ventricular myocardium at the posterior aspect of the interventricular septum. This ganglion is composed of large round
clusters of cells surrounded by fatty connective tissue. Individual clusters typically contain 40–60 cells. This ganglion begins
at the rostral aspect of the ventricular myocardium and ends
just below the aortic outlet. The LV2 ganglion is a smaller
ventricular ganglion found on the epicardial surface of the left
J Appl Physiol • VOL
ventricle in small elongated and somewhat scattered clusters
embedded in fat tissue and often in close association with
epicardial blood vessels. Although PGP 9.5 immunoreactivity
permitted the initial mapping of the locations of intracardiac
ganglia, actual counts of both labeled and unlabeled neurons
within these ganglia (Fig. 4) were made at ⫻200 magnification
using fluorescence microscopy with a FITC excitation and
barrier filter set (Nikon) because this optical system permitted
the ready differentiation of neuronal cytoplasm, nuclei, and
axons from other tissue elements within the heart. Nuclear
labeling of retrogradely labeled neurons with diamidino yellow
was also readily observed with this optical system but was
confirmed by using a UV optical filter that visualizes diamidino
yellow (Nikon). By contrast, it was often difficult to estimate
cell counts with PGP 9.5 because the nuclei of PGP 9.5immunoreactive neurons as well as the cytoplasm are both
strongly labeled. Therefore, the presence or absence of nuclear
boundaries within a cell could not be used to help differentiate
neurons from fragments. Other cardiac landmarks were also
poorly distinguished in these tissues as well. While examining
adjacent control sections, however, we observed that by using
FITC optics we could much more clearly distinguish and count
neurons within intracardiac ganglia on the basis of their mor-
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Fig. 3. Protein gene product 9.5 (PGP 9.5) immunocytochemical staining of intracardiac ganglia. A–H are photomicrographs that
correspond, respectively, to the schematic drawing of ganglia labeled A through H in Fig. 1. Arrows point to examples of ganglion
cells. f, Fat cells; g, ganglion; lam, left atrial myocardium; n, intracardiac nerve; ram, right atrial myocardium. Nomarski optics.
Final magnification: ⫻220 in A–F; ⫻110 in G and H.
PARASYMPATHETIC CONTROL OF THE LEFT AND RIGHT VENTRICLES
Fig. 4. Nos. of neurons (means ⫾ SE) found within individual intracardiac
ganglia of the cat. PA, posterior atrial ganglion; SA, sinoatrial ganglion; AV,
atrioventricular ganglion; IAS, interatrio-septal ganglion; RV, right ventricular
ganglion; CV, cranioventricular ganglion; IVS, interventriculo-septal ganglion; LV2, left ventricular ganglion 2.
J Appl Physiol • VOL
Percent labeling in all other ganglia was not significantly
different from one another (P ⬎ 0.16) and averaged 6.51 ⫾
2.54%. Note particularly that very few retrogradely labeled
neurons were found in the CV ganglion (Figs. 5 and 6).
Subsequent to multiple microinjections of diamidino yellow
into the anterior apical surface of the left ventricle (Fig. 2),
there was a statistically significant difference in the percentage
of the total number of neurons that were retrogradely labeled
within individual intracardiac ganglia [F(7,22) ⫽ 16.76, P ⬍
0.001; Fig. 7]. In this case, more than three-fourths of the
retrogradely labeled neurons found in the heart were located in
three ventricular ganglia, i.e., the CV, IVS, and LV2 ganglia.
Percent labeling in the IVS ganglion was significantly larger
than that observed in any other intracardiac ganglion (mean ⫽
42.8 ⫾ 7.6%; P ⬍ 0.05). There was no significant difference in
retrograde labeling observed between the CV and LV2 ganglia
(mean ⫽ 18.4 ⫾ 3.2%; P ⬎ 0.05); however, labeling in the CV
ganglion was significantly greater than that observed in the SA,
AV, IAS, or RV ganglia (P ⬍ 0.05). Percent labeling in all
ganglia other than the CV, IVS, and LV2 ganglia averaged
4.8 ⫾ 0.93%. When diamidino yellow was injected into the
pericardium in the control animal, 49 ⫾ 0.54% of all neurons
within each intracardiac ganglion were labeled.
DISCUSSION
Considerable uncertainty exists concerning the anatomic
location and precise physiological functions of the intracardiac
ganglia of multiple species, including the cat. Because the
sympathetic input to the heart largely arises from neurons
located in the intrathoracic ganglia (3), intracardiac ganglia are
believed to be predominantly parasympathetic, although a few
neurons containing catecholamines have been found on atrial
ganglia (19). Some evidence also indicates that intracardiac
ganglia may contain sensory neurons and local circuit neurons
(2). Although it has been long known that vagal efferents to the
heart richly innervated parasympathetic ganglia located on the
atria, it was generally believed that vagal parasympathetic
control either did not extend to the ventricles or had minimal
physiological relevance (29). In the cat, Calaresu and St. Louis
(10) reported that intracardiac ganglia were found surrounding
Fig. 5. Percentage of total number of labeled neurons in individual intracardiac ganglia subsequent to injections of a retrograde tracer into the right
ventricle. Abbreviations as in legend to Fig. 4.
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phology and autofluorescence rather than their content of PGP
9.5. Autofluorescence was not a problem because it was readily
distinguished from diamidino yellow by rapidly rotating different filter sets into the optical path. Furthermore, when on the
rare occasion in these relatively thick sections that the same
neuron could tentatively be identified in two adjacent sections,
neurons that were identified morphologically with FITC optics
were PGP 9.5 immunoreactive. The FITC optical system also
permitted the easy visualization and identification of fat cells,
epithelial cells, cardiac muscle, blood vessels, valves, and other
cardiac landmarks that further simplified the not inconsequential problem of relating the microscopic anatomy to the gross
anatomy of the heart. Finally, the regional pattern of distribution of intracardiac ganglia that was observed using PGP 9.5
immunoreactivity was identical to that seen using FITC optics
or in tissues stained with hematoxylin and eosin.
In 3 of a total of 14 animals, a few sporadic clusters of
intracardiac ganglia were seen in epicardial fat within another
intraventricular site (Fig. 1, I). Because these neurons were so
infrequently noted, however, they are not discussed further,
although they may represent some standard variation in the
neuroanatomy of the cat heart. The cat heart was observed to
contain a mean of 6,274 ⫾ 1,061 neurons. From this total, 51%
(3,210 ⫾ 587) were found within the ventricles. The number of
neurons observed within individual intracardiac ganglia varied
significantly [F(7,39) ⫽ 6.50, P ⬍ 0.001]. The two largest
ganglia in the heart were the IVS and PA ganglia (Fig. 4).
Injections of diamidino yellow into the myocardium resulted
in elongated but erratically shaped areas of intensely fluorescent labeling ⬃3–5 mm in length, which traversed the area
from the epicardial surface to well within the cardiac muscles
but did not extend into the ventricular lumens. Subsequent to
multiple microinjections of diamidino yellow into the anterior
apical surface of the right ventricle (Fig. 2), there was a
statistically significant difference in the percentage of the total
number of neurons that were retrogradely labeled within individual intracardiac ganglia [F(6,23) ⫽ 15.77, P ⬍ 0.001; Fig.
5]. In fact, more than two-thirds of the labeled neurons in the
heart were found in the IVS ganglion (Fig. 5). Percent labeling
in the IVS ganglion was statistically larger than that of any
other ganglion in the heart (mean ⫽ 66.5% ⫾ 6.7; P ⬍ 0.05).
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Fig. 6. Injections of a retrograde tracer into
the right ventricle do not cause labeling in
the CV ganglion (A) but result in substantial
labeling in the IVS ganglion (B). Arrows in
B indicate examples of ganglion cells with
labeled nuclei. Note the absence of labeled
nuclei within neurons in the CV ganglion in
A. Fluorescence microscopy. Original magnification, ⫻220.
Fig. 7. Percentage of total number of labeled neurons in individual intracardiac ganglia subsequent to injections of a retrograde tracer into the left
ventricle. Abbreviations as in legend to Fig. 4.
J Appl Physiol • VOL
glia, i.e., the SA and AV ganglia (21). We have now determined the distribution of two additional atrial ganglia, i.e., the
PA ganglion and the IAS ganglion. Furthermore, we have
detected a total of four intraventricular ganglia, one on the right
ventricle, two on the left ventricle, and one within the interventricular septum. Although there is some homology between
species in the distribution of intracardiac ganglia, there are also
substantial interspecies differences in the number of atrial or
ventricular ganglia, their distribution, and in estimates of the
total number of neurons found in the heart (5, 6, 10, 27).
The distribution of parasympathetic postganglionic nerves to
many regions of the heart, including both ventricles, has
largely been inferred from physiological studies (1, 8, 12, 29),
not neuroanatomic studies. We have previously determined the
intracardiac location and physiological effects of the CV ganglion on cardiac functions. It was reported that microinjections
of a ganglionic blocking drug into the CV ganglion attenuated
the negative inotropic effects of vagal stimulation on the left
ventricle, without influencing either vagally mediated negative
chronotropic or negative dromotropic effects (18, 20). These
physiological data predict that, at a minimum, neurons in the
CV ganglion should project to the left ventricle and not to other
regions of the heart. In the present study, a retrograde tracer
was placed into the anterior apical surface of the left ventricle
because some data indicate that the autonomic innervation of
the ventricles is not homogeneous. Thus, for example, the
sympathetic postganglionic innervation of the anterior apical
surface of the left ventricle arises from the left stellate ganglion, while the corresponding innervation of the posterobasal
wall arises from the right stellate ganglion (30). We observed
that the CV ganglion was robustly labeled subsequent to
microinjections of tracer into the left ventricle but was not
significantly labeled subsequent to tracer injections into the
right ventricle. These neuroanatomic data are therefore consistent with our previous physiological hypothesis that the CV
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the great vessels and on the atria, or within the interatrial
septum. However, “ganglion cells were never seen in the kitten
ventricles” (Ref. 10; p. 61). This conclusion was based, however, on an examination of the ventricles in only one kitten. In
analogy with the data in the cat, early data suggested that the
rat and dog ventricles also contained few or no intracardiac
ganglia (8, 11). Recent evidence, however, indicates that at
least one intraventricular ganglion is present in both the cat
(20) and the rat heart (34, 35), and multiple intraventricular
ganglia are present in the dog heart (39). In the present report,
therefore, we have examined sections throughout the atria and
ventricles of the cat heart for the presence of intracardiac
ganglia. Our data indicate that there are eight regions of the cat
heart that consistently contain ganglia. Within the atria, we
have previously defined the location of two intracardiac gan-
PARASYMPATHETIC CONTROL OF THE LEFT AND RIGHT VENTRICLES
J Appl Physiol • VOL
the left ventricle. It is interesting to speculate that this dualinnervation pattern may help to coordinate or segregate vagal
effects on ventricular performance. It would be interesting
indeed to inject two separate retrograde tracers into the left and
right ventricles to see whether the same neurons in the IVS
ganglion project to both targets or whether separate populations of neurons within this ganglion project to individual
ventricles.
Greater understanding of the pathways and mechanisms
whereby the vagus nerve exerts its effects on the ventricles
may be helpful in understanding autonomic responses during
acute myocardial ischemia. Autonomic responses during acute
myocardial ischemia play a major role in predicting the probability of a fatal outcome. If activation of the sympathetic
nervous system occurs, malignant ventricular fibrillation and
sudden death are likely (31). If, on the other hand, acute
myocardial ischemia is followed by activation of cardiac vagal
efferents, a protective antifibrillatory effect is seen and the
probability of survival is enhanced (16, 37). These results may
be due to the fact that enhanced vagal tone increases ventricular fibrillation threshold and reduces the arrhythmogenic effects of sympathetic activation (37). Human studies indicate
that vagal activity is reduced for several months after myocardial infarction, whereas sympathetic tone is enhanced (32, 34,
48). It is believed that this altered autonomic balance may in
part mediate the increased incidence of sudden cardiac death
after myocardial infarction (26). The autonomic nervous system also has a profound effect on the pathogenesis of numerous
arrhythmias, including supraventricular tachycardias, atrioventricular block, and ventricular arrhythmias associated with
myocardial ischemia and sudden death (13, 26). These data
emphasize the clinical importance of understanding how the
parasympathetic nervous system regulates ventricular functions. This knowledge is essential to the future rational design
of experimental interventions in heart disease.
GRANTS
This research was supported in part by grants from the National Heart,
Lung, and Blood Institute (RO1-HL-51917) and American Heart Association
to V. J. Massari. Additional funding was provided by the Gustavus and Louise
Pfeiffer Research Foundation to A. L. Gray and the Specialized Neuroscience
Research Program (1U54-NS-39407, M. A. Haxhiu, Principal Investigator).
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ganglion selectively mediates a negative inotropic effect on the
left ventricle because the CV ganglion does not effect chronotropic or dromotropic cardiac functions (18, 20) and does not
project to the right ventricle to potentially influence the physiological function of this chamber. The present data further
indicate, however, that although the physiological effects mediated by the CV ganglion are selective, it is not the exclusive
source of vagal efferents to the left ventricle. Rather, two other
intraventricular ganglia also significantly mediate vagal control
of left ventricular functions. These include the IVS and LV2
ganglia. Percent labeling in the IVS ganglion was significantly
larger than that found in any other intracardiac ganglion, and
more than three-fourths of the retrogradely labeled neurons
found in the heart were located in these three ventricular
ganglia. By contrast, the percent labeling in each of the other
ganglia averaged ⬍4%. Further studies will be required to
determine the precise physiological roles of the IVS and LV2
ganglia in the control of left ventricular functions and the
potential extent of their influence on left ventricular contractility.
To control for potential labeling due to leakage of tracer
from the injection sites, tracer was injected into the pericardial
space. This resulted in a uniform 49 ⫾ 0.54% degree of
neuronal labeling within each intracardiac ganglion. By contrast, injections of tracers into either the left or right ventricular
wall resulted in a highly statistically significant difference (P ⬍
0.001) in percent labeling across the different intracardiac
ganglia. We conclude from these data that leakage of the tracer
from its injection sites was minimal. We have also considered
the possibility that the degree of retrograde ganglionic labeling
seen with diamidino yellow might potentially be proportional
to the distance between labeled ganglia and the sites of injection of this tracer. In this way, relatively close ganglia would be
labeled, whereas distant ganglia would not. Several lines of
evidence indicate that failure to find retrogradely labeled neurons within various intracardiac ganglia subsequent to injections of the tracer into either ventricle was not due to relative
distance from the injection site or inadequate survival time.
Thus, for example, the present data indicate that injections of
tracer into the right ventricle failed to label neurons in a closely
adjacent ganglion actually contained within the right ventricle
(the RV ganglion) while significantly labeling neurons approximately equidistant from the injection sites in the interventricular septum (the IVS ganglion). Furthermore, the 4-day survival time that was used in the present report has also been
demonstrated to provide ample staining of cardioinhibitory
vagal preganglionic neurons in the nucleus ambiguus of the
brain stem of the cat after microinjections into the SA or AV
ganglia (7), a distance that is substantially greater than that
separating all intracardiac ganglia from injection sites in either
the left or right ventricle.
The present data indicate that the vagal postganglionic
control of right ventricular function is mediated primarily from
the IVS ganglion, which contained more than two-thirds of the
retrogradely labeled neurons found in the heart. This was
statistically significantly greater than that of any other intracardiac ganglion. By comparison, average percent labeling in
all other ganglia was not significantly different from one
another and averaged ⬃6% of the total labeling in the heart.
The IVS ganglion, therefore, is the largest source of neurons
innervating the anterior surfaces of both the right ventricle and
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