Development of myocardial fiber organization in the rat heart.

Confocal laser-scanning microscopy of phalloidine-stained actin fibers is a relatively new tool for studying the development of myocardial fiber organization. It seems to show orientation of myocytes in rather early embryonic stages. To further evaluate the differentiation of the myocardium, this technique was compared with transmission electron microscopy in rat embryos aged between 11 and 18 days. Although the confocal images of actin filament patterns pointed to early myocyte orientation, the electron micrographs revealed that even at 17 days the ventricular myocardium was far from mature. Myofibrils never completely filled the myocytes, and lack of organization was the rule rather than the exception. The organized structure as revealed by confocal microscopy was based on cell-to-cell continuity, whereas electron microscopy showed crossing and disarray within individual myocytes. Exceptions were in the ventricular trabeculations, which showed precocious myofiber differentiation. The trabeculations probably support ventricular systole in those stages in which the free walls do not yet provide efficient contractions. The other exception was the wall of the outflow tract, which showed well-oriented myofibrils from early stages onwards. Apparently, the outflow tract has a different function in these stages. The differences found between confocal microscopy and electron microscopy suggest that some caution is indicated in the interpretation of fluorescent images of relatively low magnification.

Development of the atrioventricular valve tension apparatus in the human heart.

Using various microscopical techniques we studied the development of the atrioventricular valves in human hearts between 5 and 19 weeks of development. Within the atrioventricular cushions two different layers could be recognized that remained present in all ages studied. The atrial layer, being present at the side of the atrioventricular orifice, was positive for laminin while the ventricular layer, that was connected to the myocardium, was positive for fibronectin and collagen III. Fate-mapping of these two layers, morphometrics, and scanning electron microscopy, supplemented with in vivo labeling of cushion tissue in chicken hearts have lead to new insights in the process of valve development. The cushions became freely movable prevalvular leaflets by delamination of ventricular myocardium underneath the cushion tissue. This myocardium gradually retracted towards annulus and papillary muscles and finally disappeared, resulting in fibrous, non-myocardial valves. The atrial layer of the cushions remained present as a jelly-like surface on the valve leaflets while the ventricular layer of the cushions became the compact fibrous tissue of the leaflets and the chords. Chordal development was first visible at 10 weeks of development when gaps were formed in the ventricular layer of the cushions on top of the papillary muscles. These gaps enlarged into the interchordal spaces while the cushion tissue in between the gaps lengthened to form the chords. We conclude that the leaflets as well as the chords of the atrioventricular valves are derived from atrioventricular cushion tissue. Myocardium is only important for loosening of the leaflets while keeping connection with the developing papillary muscles. Errors in delamination or retraction of myocardium or remodeling of cushion tissue into chords form the basis for various congenital valve anomalies.

Ultrastructural changes of the myocardium in the embryonic rat heart.

BACKGROUND: Ultrastructural changes of the embryonic heart have been described, and quantitative studies have reported the changes of cellular organelles in late fetal and postnatal development. However, no specific data are available on the quantitative morphology of the individual segments and intersegmental junctions of the early embryonic heart, although these components must have different functions. METHODS: We measured the absolute volumes of glycogen, Golgi complex, myofibrils, mitochondria, and the surface areas of the rough endoplasmic reticulum and mitochondrial cristae in the different regions of the embryonic rat heart by using stereological tools. RESULTS: During embryonic development, the cardiac segments and intersegmental junctions increase their glycogen volume. The sinoatrial junction and primary fold show a more rapid increase than all the other cardiac regions, whereas the atrioventricular canal shows a high level of glycogen content throughout the period studied. The Golgi complex and rough endoplasmic reticulum show a conspicuous decrease from day 15 onward. The cellular content of myofibrils and mitochondria and the surface area of the mitochondrial cristae show a gradual increase from day 11 to day 17 of development, but full maturation apparently takes place in late fetal and early postnatal stages. At day 15 of development, the cellular volumes of myofibrils and mitochondria show a temporary decrease. CONCLUSIONS: The glycogen content cannot be explained on the basis of metabolism alone. The storage of glycogen is hypothesized to serve mechanical cell stability and may also be related to a target mechanism for ingrowing nerves. Myofibrillar and mitochondrial contents of the myocytes indicate a relatively late differentiation of the venous pole of the heart. Uninterrupted maturation is only started at the time of septation.

Nuclear and cellular size of myocytes in different segments of the developing rat heart.

BACKGROUND: In the embryonic heart, the individual cardiac segments show different growth rates. For the analysis of changing form in relation with changing function, data on number and shape of cardiomyocytes are necessary. Such data will give insight into the process of hypertrophy and/or hyperplasia as they may take place in the myocardium in the embryonic period. METHODS: We have measured the volumes of the nuclei and myocytes as well as the surface areas of the nuclear envelope and cellular membrane using stereological tools in rat embryos from 11 days postcoitum to 17 days postcoitum. From the data of the cellular volume of the myocytes and the myocardial volume of the individual segments, we have calculated the total number of myocytes during the developmental period. RESULTS: It is shown that the sinus venosus, sinu-atrial junction, and atrium increase their cellular volume during development, whereas the other cardiac segments show no difference in cellular volume. Similarly, the surface area of the cell membrane of the sinus venosus and sinu-atrial junction had increased during development. The nuclear volume and the surface area of the nuclear envelope did not differ during the period studied. The total number of myocytes showed a conspicuously smaller increase in the atrio-ventricular canal and distal outlet segment than in the other segments. CONCLUSIONS: The increase of the cellular volume in the segments sinus venosus and sinu-atrial junction seems to be due to a late differentiation process. In general, however, the increase of the myocardial volume in the individual cardiac segments is caused by hyperplasia of the cardiomyocytes in these segments and not by hypertrophy. The surface area of cells has a fixed relationship with cell volume, indicating that no important changes take place in the developmental period studied.

Growth of the myocardial volumes of the individual cardiac segments in the rat embryo.

BACKGROUND: Although the growth of the developing heart in relation to an increase of ventricular systolic pressure and the growth of the entire embryo during development has been described, no data are available on the growth of the individual segments and intersegmental junctions. Because these different portions are known to function differently, the need for data on their individual development is obvious. METHODS: We have measured the volumes of these different compartments by Cavalieri's point counting method in rat embryos from 11 to 17 days. RESULTS: It is shown that sinus venosus and sinu-atrial junction as well as the main compartments atrium, inlet, and proximal outlet segment grow roughly proportional to the total myocardial volume. Atrio-ventricular canal and distal outlet segment show a restricted growth and their proportional volumes decrease in time. The inlet segment is the most important part of the ventricular mass at 11 days of gestation, when it is still larger than the proximal outlet segment and, thus, takes the greater part in systolic action of the ventricular mass. The growth of the primary fold increases from day 13 onwards and can be considered as part of the wall of the inlet segment which gives rise to the main part of the ventricular septum. CONCLUSIONS: The timing of the septal volume increase fits with qualitative descriptions of ventricular septation. The atrio-ventricular canal and distal outlet segment have an important constrictive function in early stages, when valves are not yet present. Slow conduction and contraction patterns have been reported to be a characteristic feature of these portions of the embryonic heart. With development of valves these segments are loosing their mechanical function and, thus, their proportional volume declines.