Mireille Masson-Pévet

Pacemaker cell types in the rabbit sinus node: A correlative ultrastructural and electrophysiological study

In isolated preparations of the rabbit sinus node, we have investigated the fine structure of the tissue at many sites which had been electro-physiologically identified by means of microelectrode recordings. For each of these sites we quantified the myofilament density of the cells, since this appeared to be a useful parameter for characterizing cell types in the sinus node region; and because myofilaments were present both in isolated form and organized in myofibrils, the degree of organization was also measured in a semi-quantitative fashion. The electrical activity of cells at a given site was characterized by the activation moment relative to the cardiac cycle and furthermore by the rate of diastolic depolarization and the maximum rate of rise of the action potential. From the centre of the node toward the periphery a very gradual increase in myofilament density was observed in all directions. It was found that the rate of diastolic depolarization, which feature is generally accepted as being basic to the automaticity of the sinus node, was inversely related to the volume percentage of myofilaments. This means that a relation exists between the pacemaker action and the cell type. The anatomically less developed cells, i.e. the cells with the lowest density of organelles, which are located in the central portion of the node, are the most specialized pacemakers. No clear relation was found between the myofilament density and the rate of rise of the action potential. In the direction of the crista terminalis we observed an increase in the rate of rise and an increase in conduction velocity concomitant with the increase in myofilament density. Toward the interatrial septum, however, the increase in myofilament density was not accompanied by an increase in rate of rise; in this direction the impulse conduction was blocked. A correlation between cell type and impulse conduction could thus not be established.

Sinus node and atrium cells from the rabbit heart: A quantitative electron microscopic description after electrophysiological localization

Abstract Electrophysiologically identified cell groups in the sinus node from the rabbit have been compared with atrial fibers with the electron microscope. The point counting method has been used to estimate the volume density of the following structures: nucleus, mitochondria, myofilaments, sarcoplasmic reticulum tubules and subsarcolemmal vesicles. These data were collected in leading pacemaker cells, latent pacemaker cells and atrium cells from the crista terminalis. It has been found that organized structures in leading pacemaker cells occupy about 50% of the cell volume, as compared with over 90% in atrial fibers. Leading pacemaker cells consequently appear extremely “empty”. It has also been found that the group of cells which show the characteristic features of leading pacemaker cells at the ultrastructural level as observed in the correlated experiments is larger than the leading center found in electrophysiology and thus it seems impossible with the actual observation methods to delineate the leading pacemaker center using only cytological criteria.

Asymmetry of the sino-atrial conduction in the rabbit heart

Abstract In isolated right atria of the rabbit heart the impulse conduction from the sinus node to the interatrial septum was studied, using the microelectrode technique. After the electrophysiological experiments several preparations were subjected to a correlative morphological investigation, using light or electron microscopy. By making incisions in the preparation we could demonstrate that the pacemaker region was bordered at the medial (septal) side by a zone in which conduction was blocked completely; the entering wavefronts died out gradually in both antegrade and retrograde directions. The double component action potentials that can be recorded in this region in the spontaneously beating intact preparation, appeared to be an almost passive summation of the wavefronts that entered this zone from the pacemaker side (first component) and from the septal side (second component). The tissue at the septal side was activated by a front that encircled the conduction block. Microscopically the area of the block was not uniform: it showed transition in cell type and tissue architecture, as was observed in a previous study also at the lateral (cristal) border of the node, where the pacemaker impulse is conducted with increasing speed toward the atrium. Neither did we find a difference in spread of electrotonus between the medial and lateral border of the sinus node. The block is therefore thought to be caused by the electrophysiological properties of the cells, most likely their low excitability.

Functional and morphological organization of the cat sinoatrial node

The feline sinoatrial node has a unifocal impulse generation as previously described for rodents. Its main component is collagen. The primary pacemaker consists of at most 2000 cells, but appears to function normally with less than 500 cells. Primary pacemaker cells are found in the area where empty cells are predominant. A negative correlation between myofilament density and diastolic depolarization rate, known to exist in the rabbit and guinea-pig, is absent in the cat. Gap junctions are seen in the center and in the periphery of the nodal region, but they are extremely rare. The electrophysiological characteristics of the primary pacemaker of the cat are quite similar to those of the rabbit, although the nodal morphology is very different. Abrupt transitions from one cell type into another are observed in the feline sinoatrial node. From this morphological point of view the feline sinoatrial node resembles the canine and human sinoatrial nodes more than the lapine sinoatrial node.

The development of beat-rate synchronization of rat myocyte pairs in cell culture

SummaryWhen two spontaneously beating neonatal rat heart cells in tissue culture were allowed to grow together they synchronized their originally independent beats to a common rhythm, as measured with an opto-electronic technique. Both single isolated cells and cell pairs exhibited a highly irregular beating pattern. Beating irregularity was strongly and positively correlated with mean interbeat interval. Synchronization of beating occurred in 50% of the pairs studied within one beating interval. In the remaining cell pairs, the first synchronized beat was followed by a 4–65 s period of partial synchronization. The time difference between contraction moments of two cells in a pair respective to each other (latency) changed upon synchronization from a random value to a fixed value. In a few cases the latency decreased during 20 to 30 s after the first synchronized beat before a steady-state value was reached. The mean interbeat interval (IBI) of the synchronized cell pairs was governed by the mean IBI of the originally faster beating cel. In 83% of the cases the mean IBI of the cell pairs was between that of the originally isolated beating cells. We conclude from the experiments described that physical coupling (i.e. gap junction formation) is virtually complete before beating synchronization occurs.

The caveolae in rabbit sinus node and atrium

SummaryCaveolae or membrane vesicles are commonly observed in smooth and skeletal muscle as well as in working heart muscle. Using sections of fixed tissue and replicas of freeze-cleaved material, we show in this study that caveolae are also very numerous in sinus node cells of the rabbit, and to a lesser degree, in the atrial cells.Caveolae increase the plasma membrane surface area by 115% in the leading sinus node, and by 56% in the atrial cells. In these two cell types, the membrane of the caveolae contains four times fewer intramembranous particles than the rest of the plasma membrane, and this difference applies to both PF and EF faces. The role of the caveolae is still unclear, but it does not seem that they have a pinocytotic function.

Little-excitable transitional cells in the rabbit sinoatrial node: a statistical, morphological and electrophysiological study

It is demonstrated that systolic and diastolic depolarization rate are correlated with the percentage of myofilaments in the cells of the rabbit sinoatrial node. It appears that, in the rabbit sinoatrial node, little-excitable transitional cells exist in the zone of propagation at the septal side of the typical nodal cells.

On the Ultrastructural Identification of Pacemaker Cell Types within the Sinus Node

The heart beat originates within a small group of cells in the sinus node. In the rabbit this site of earliest discharge or leading pacemaker center, is located in the intercaval region, 0.5 – 2 mm away from the medial border of the crista terminalis. It contains typical nodal cells, and is part of a much larger group of cells that all show diastolic depolarization. This latter group of cells, which under standard conditions (see Bleeker et al., 1980) follow the discharge of the leading pacemaker cells and which never reach threshold spontaneously are called transitional cells. From its site of origin the excitation wave propagates through the sinus node preferentially in an oblique cranial direction toward the crista terminalis. In all other directions, the spread of excitation is considerably slower; in the direction of the interatrial septum conduction fails (Fig. 1).

Intact isolated sinus node cells from the adult rabbit heart

Abstract Till now, the membrane currents underlying spontaneous activity in the sinus node have been studied with the voltage clamp method, on pieces of tissue which were derived mainly from the area of transitional cells in the sinus node [6]. In order to obtain a preparation to study the pacemaker currents in leading pacemaker cells, we have developed a method which enables us to get viable, intact, single sinus node cells. Electrophysiologically indentified leading and latent pacemaker centers from the rabbit sinus node were dissociated in a medium containing collagenase and DNA-ase. A reproducible yield of 1 × 10 5 ultrastructurally intact isolated cells has been obtained from the leading center of the node. These cells can be kept for at least 2 days after the isolation in salt solution containing 2.2 m m calcium chloride, without any loss of integrity. The dissociation procedure described is suitable for typical nodal cells and to a lesser degree for transitional cells derived from the latent pacemaker areas of the sinus node. Because working heart cells are destroyed by this method, almost all intact myocardial cells in the suspension are of nodal origin.