Download Ventricular Anatomy for the Electrophysiologist (Part II)

SPECIAL REVIEW
Ventricular Anatomy for the
Electrophysiologist (Part II)
서울대학교 의과대학 병리학교실 서 정 욱
이화여자대학교 의학전문대학원 김 문 영
ABSTRACT
The conduction fibers and Purkinje network of the ventricular myocardium have their peculiar location and
immuno-histochemical characteristics. The bundle of His is located at the inferior border of the membranous
septum, where the single trunk ramifies into the left and right bundle branches. The left bundle branches are
clearly visible at the surface. The right bundles are hidden in the septal myocardium and it is not easy to
recognize them. The cellular characters of the conduction bundles are modified myocardial cells with less
cytoplasmic filaments. Myoglobin is expressed at the contractile part, whereas CD56 is expressed at the
intercalated disc. A fine meshwork of synaptophysin positive processes is noted particularly at the nodal
tissue. C-kit positive cells are scattered, but their role is not well understood. Purkinje cells are a peripheral
continuation of bundles seen at the immediate subendocardium of the left ventricle.
Key words: ■ conduction system ■ Purkinje network
Introduction
The functional assessment of abnormal cardiac
rhythm and a targeted treatment based on
electrophysiologic studies are successful advances in
cardiology.1 Morphological assessment or confirmation
of hearts with such abnormalities is rare, not only due
to the limited availability of human hearts but also
inherent technological limitations of existing
technology.2 Classical morphological approaches and
immuno-histochemical or molecular biologic
assessment on a heart transplant recipient will be a
unique chance to study the conduction system of the
Correspondence: Jeong-Wook Seo, Department of Pathology, Seoul
National University College of Medicine and Hospital, 103 Daehangno,
Jongro-gu, Seoul 110-799, Korea
Tel: 82-2-2072-2550, Fax: 82-2-743-5530
E-mail: [email protected]
* Kim MY is a senior student from Ewha Woman’s University School of Medicine.
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Journal of Cardiac Arrhythmia
■
pathology
■
arrhythmia
■
electrophysiology
human heart. In this brief review, the histological
characteristics of conduction cells, stained by
conventional and immuno-histochemical staining, are
demonstrated in the second part of the review.3
The characteristic location of the ventricular
conduction system
The atrioventricular node is situated in its
subendocardial location at the triangle of Koch. A
cluster of short spindle cells are arranged compactly
around the AV nodal artery. The long axis of the AV
node is approximately 0.7 cm. Fatty tissue and some
nerve fibers are seen around the node. The bundle of
His is 0.5~0.7 cm long and 0.1 cm in diameter (Figure
1, 2). The bundle has a short traversing segment to the
anterior superior direction and then ramifies into the
SPECIAL REVIEW
Histological and immuno-histochemical
characteristics of the conduction system
right and left bundles (Figure 2). The left bundle and
its anterior and posterior fascicles are recognized
histologically at the subendocardium of the left
ventricular surface of the ventricular septum. The right
bundle is located between the thick bundles of the
right ventricular myocardium and the true septal
myocardium. The histological verification of the right
bundle is not easier than the left side.
Masson trichrome staining is one of the convenient
methods to differentiate the conduction tissue from
collagenous stroma and pathologic fibrous changes of
the myocardium are demonstrable as well through the
trichrome staining (Figure 3). It is understood that
bundles and fascicles are covered with fibrous sheath
so that rapid conduction from the myocardium is
insured, although electrical conduction between cells
needs a special gab junction protein or intercalated
disc as revealed by electron microscopy. Side to side
contact by a parallel alignment of muscle bundles is
not conductive. It is therefore important to have an
axial alignment to propagate contractile stimulation.
Cells of the conduction system are mainly
myocardial cells. The cytoplasm contains myofibrils
and they do not have cytoplasmic processes as seen in
neural cells. The conduction cells have less myofibrils
than myocardial cells and intimate contact with nerve
endings are noted. Conduction cells at the microscopic
level are spindle cells with smaller cytoplasm. The
immuno-histochemical reaction against myoglobin is
positive (Figure 3). The intensity of staining by
immuno-histochemistry is often influenced by fixation
and other factors so that visual intensity of staining
may not be an indicator of the amount of protein.
Neural cell adhesion molecule (NCAM, CD56) is a
member of the immunoglobulin super family,
mediating intercellular adhesion in the nervous system
and skeletal muscle and CD56 was up-regulated at the
ischemic myocardium in human and animal models.4
There was a strong positive reaction for CD56 at the
MV
MV
NC
RC
9
8
7
6
5
1
16
10
17
11
18
12
19
13
20
14
21
15
2
3
4
Figure 1.Left ventricular surface of the ventricular septum. The initial cut was made at the commissure of right and noncoronary cusps (RC, NC) of the aortic valve. After that, 4 cm long tissue blocks, 2 cm above and below the aortic valve,
were designed to make 9 pieces of vertical sections, each with 4 mm thickness (#1-9). Block #9 is toward the anterior
wall of the left ventricle, while block #4 toward the posterior wall. The rest of the septal surface was sectioned horizontal
to make blocks (#10-21).
MV; mitral valve, NC; non-coronary cusp, RC; right coronary cusp
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B
A
TT
MV
HB
AV
TV
TV
D
C
AO
INF
HB
RC
TV
Figure 2. A. histological section at piece #3 of Fig 1 showing the atrioventricular node (AV) over the fibrous annulus at
the atrial surface. B. A section at piece #2 of Fig 1 showing the non-branching His bundle (HB) at the inferior border of
the membranous septum, where the posterior fascicle (long open arrows) ramifies. C. A section at piece #1 of Fig 1
showing a non-branching bundle (HB), the anterior fascicle of the left bundle (long arrows) at the surface and the
intramyocardial right bundle (short arrows). D. A section at piece #7 of Fig 1 showing the right part of the ventricular
septum extending to the infundibulum (INF) of the right ventricle. Short arrows indicate the plane between myocardial
layers where the right bundle is located (Masson’s trichrome stain).
AO; aorta, MV; mitral valve, RC; right coronary cusp of aortic valve, TT; tendon of Todaro, TV; septal leaflet of tricuspid valve
intercalated disc of the ventricular myocardium
(Figure 4). The atrioventricular node and bundles did
not show such strong reaction. It is also noted that the
length of myocytes represented as the length between
two intercalated discs was longer at the
subendocardial left bundles. Synaptophysin, another
marker for neural cells is negative at the myocardial
cells, but a fine meshwork of synaptophysin positive
fibers is seen around the individual conduction cells. The
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Journal of Cardiac Arrhythmia
staining pattern of synaptophysin at the atrial
myocardium was a slender network between the
myocytes (Figure 5). This pattern was the most prominent
at the atrial myocardium and AV node. His bundle is
faintly stained and ventricular myocardium is the
weakest. C-Kit is known to be one of the markers of
stem cells5 and interstitial Cajal-like cells related with
atrial arrhythmia.6,7 The C-Kit positive cells were rare
isolated cells at His bundle (Figure 5). Atrioventricular
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B
A
cs
PF
PF
MC
MC
LV Septum
D
C
PF
PF
MC
MC
Figure 3. A. Histological section at piece #16 of Fig 1 showing a cross section of the distal part of the subendocardial
left bundle forming Purkinje fibers (PF) (arrow) separated from the underlying myocardium (MC). B. A section at piece
#16 of Fig 1 showing subendocardial smooth muscle nodules overlying left bundles (open arrow & dot line). C. A section
at piece #1 of Fig 1 showing thick and continuous subendocardial smooth muscle fibers (open arrow & dot line) stained
violet with Masson trichrome but appear negative for myoglobin as shown in D. (A~C; Masson’s trichrome stain, D;
Myoglobin)
node has some, but rest of area show few positive
cells.
Previous studies on the conduction system of the
heart showed subsarcolemmal SR (sarcoplasmic
reticulum), but no T-tubule or corbular SR. Connexin
45 and neurofilament M (NF-M) were found, but
connexin 43 and atrial natriuretic peptide were
lacking.1,8
Myocytes of the sinoatrial and atrioventricular (AV)
nodes characteristically have small, dispersed gap
junctions predominantly composed of Cx45. The His
bundle co-expresses Cx45 with Cx43, while its
downstream branches prominently express Cx40.
Cx43 becomes abundant in the more distal portions of
the system, while Cx45 is expressed continuously
from the AV node to the ends of the Purkinje fibers.9
Purkinje network
The Purkinje fibers are part of the ventricular
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A
B
Figure 4. A. An intense positive reaction for CD56 at intercalated discs (arrows) of ventricular septal myocardial cells.
B. A less intense positive reaction for CD56 at bundle branches (double head arrow) and long interval intercalated discs.
A
B
Figure 5. A. The staining pattern of synaptophysin at atrial myocardium was a slender network between the myocytes.
B. The staining pattern of C-Kit positive cells at His bundle. (x400) (A; Synaptophysin, B; C-kit)
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Journal of Cardiac Arrhythmia
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conduction system and were originally discovered by
Tawara.10,11 These fibers conduct excitation (electrical
activation) rapidly from the bundle of His to the
ventricular myocardial tissue. The Purkinje fibers are
myocardial cells with vacuolated cytoplasm located in
the ventricular walls of the heart, just beneath the
endocardium. 12 Purkinje fibers, being modified
myocardial cells, contain some contractile protein, but
the amount is small and glycogen and other organelles
occupy the cytoplasm. It has been argued that
vacuolated Purkinje cells are only seen in the ungulate
heart, but the vacuolated character is noted in human
heart too. There are, however, muscular cells, smooth
muscle cells, other than Purkinje fibers found in the
subendocardium (Figure 3C), particularly in hearts
with failure.
From the point of view of ultrastructural composition,
the cells of different parts of the cardiac conduction
system are partly similar. In contrast to the heart
contractile cardiomyocytes, the cells of the cardiac
conduction system including Purkinje fibers have a
small amount of myofibrils, small mitochondria,
lighter cytoplasm and higher glycogen content, but no
T-tubular system.
The electrophysiologic demonstration of the
Purkinje network showed a propagation of excitation
starting from the apex, continuing rapidly towards the
base of the ventricle resulting in a contractile motion
from the apex to the base.12 Another study using
electromechanical wave imaging visually confirmed
that artificially created sinus rhythm and right-atrial
pacing, consisted of a contraction wave that started at
the apex right at the beginning of the QRS complex,
propagated along the septum and then the leftventricular posterior wall and finally to the base.13
Ventricular pacing, on the other hand, revealed a
disharmonized contraction of the ventricle, confirming
that Purkinje fibers have a crucial role in the synchronization
of ventricular contractions.13 A richly trabeculated
endocardium, in which the endocardial invaginations
carry the Purkinje tissue partway into the left
ventricular endocardium, further increases the speed
and area of early ventricular activation. This is a factor
in ventricular synchronous activation together with the
distribution of the conduction branches and the rapid
velocity of the conduction system.16
Rapid ventricular arrhythmias in post-myocardial
infarction in the canine heart arise from ectopic foci
(triggered) within the Purkinje fiber network located
in the subendocardium of the infarct zone in the left
ventricle. These spontaneously occurring arrhythmias
predictably occur between 24 to 48 hours after
occlusion. In Purkinje cells dispersed from the
subendocardium of the infarct zone (Purkinje cells
from a 48-hour infarcted heart), the density and
function of several sarcolemmal ion channels are
altered compared to normal Purkinje cells. Little work
has been done on remodeled Purkinje cells,
particularly Purkinje cells involved in the initiation
and perpetuation of cardiac arrhythmias in diseased
hearts.17
The Purkinje fibers are distinguished from heart
muscle cells by a distinct pattern of gene expression.
They up-regulate Cx42, a conduction cell-specific
gap-junction protein, unique ion channels, and genes
typically expressed in neuronal cells. In addition,
conduction cells induce a unique set of myofibrillar
protein genes, such as slow-twitch skeletal muscle
myosin heavy chain (sMyHC), atrial myosin heavy
chain (aMyHC), and skeletal muscle-type myosin
binding protein-H (MyBP-H). Purkinje fibers also
down-regulate heart muscle-specific myofibrillar
proteins, such as troponin and myosin binding proteinC (cMyBP-C), which are essential for normal heart
muscle contractility. 14 Studies on experimental
animals with ischemic heart disease demonstrate
alteration in the distribution of gap junction
immunolabel to the sides of the myocyte, called
‘lateralization’. Gap junction mediated passage of
ionic/molecular signals appears responsible for the
spread of the ischaemia-reperfusion injury from
myocyte to myocyte that leads to rigour contracture
and cell death.
Gap junction channels are almost absent in the sinus
node, there are few of them in the AV node, but there
are a lot of them in fast conducting muscle cells such
as His and Purkinje fibers. This gives rise to the
anisotropic propagation of depolarization along the
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cardiac muscle fiber orientation.15
Heterogeneity of gap junction distribution
combined with reduced Cx43 levels appears to act cooperatively to create an arrhythmogenic substrate at
less severe levels of overall gap junction reduction
than predicted in theoretical models. The findings in
mouse models lend considerable support to the view
that the nature and extent of the Cx43 reduction in the
failing human ventricle is, in practice, of sufficient
magnitude to increase susceptibility to arrhythmia.9
The Cx43 gap junction arrangement is particularly
disordered in hypertrophic cardiomyopathy, the most
common cause of sudden cardiac death due to
arrhythmia in young adults. In cardiac-restricted Cx43
knockout mice selected for longer term survival,
reduction of coupling resulting from declining
ventricular Cx43 leads to propagation of the impulse
across numerous Purkinje/ working ventricular
myocyte junctions that normally remain dormant,
thereby creating abnormal activation patterns and
wave-front collisions in the ventricular myocardium.
Conclusion
It is important that the His bundle and the left
bundle branches form a continuum with the distal part
of the ventricular conduction system, named the
Purkinje network. This will allow myocardial
contraction to begin from the apex after the activation
of the Purkinje network. The key morphological
feature of Purkinje fibers in ungulate hearts is
vacuolated cytoplasm, which is a manifestation of
small numbers of contractile elements and a large
amount of glycogen. Molecular characterization of the
activity of connexin 43 is one of the important factors
involved in arrhythmias in heart failure, myocardial
infarcts and hypertrophic cardiomyopathy.
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Journal of Cardiac Arrhythmia
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