Growth of the myocardial volumes of the individual cardiac segments in the rat embryo.код для вставкиСкачать
THE ANATOMICAL RECORD 243:93-100 (1995) Growth of the Myocardial Volumes of the Individual Cardiac Segments in the Rat Embryo M.W.M. KNAAPEN, B.C.M. VROLIJK, AND A.C.G. WENINK Department of Anatomy and Embryology, University of Leiden, The Netherlands ABSTRACT 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-atrialjunction 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 septa1 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. 0 1995 Wiley-Liss, Inc. Key words: Heart, Development, Myocardium, Stereology During development the embryonic blood pressure and blood volume increase linearly with body weight and so does the myocardial volume (Van Mierop and Bertuch, 1967; Clark et al., 1986).However, it has been reported that the growth rate of the entire embryo shows sudden changes in certain stages, which could be due to changes in cell activity and cell differentiation of several organs and/or tissues (Goedbloed, 1972). More specifically, cardiac growth rate changes have been described in relation to the embryonic crown-rump length (Mandarim-de-Lacerda, 1991). These might reflect certain changes in cardiac differentiation. Indeed, some cardiac components have specific functions during embryonic life, such as the sphincter-like function of the junctions which contain endocardia1 cushion tissue (Goerttler, 1955; Jaffee, 1965). Moreover, different action potential and contraction patterns have been related to different expression patterns of myosin isoforms in the various segments of the developing heart (de Jong et al., 1987; Evans et al., 1988; Agata et al., 1993). Thus, myocardial growth seems to be governed 0 1995 WILEY-LISS, INC. by the expression of several genes which regulate typical cell activities and cell differentiations of different segments in the developing heart. Yet, the individual growth of these segments and intersegmental junctions has never been described. We have approached this subject by measuring the myocardial volume of the entire heart as well as the myocardial volumes of the individual segments in rat embryos ranging from 11to 17 days of development. MATERIAL AND METHODS We used 35 Wistar rat embryos ranging from day 11 to day 17 post coitum. After ether anaesthesia and cervical dislocation of the pregnant females, the embryos were excised from the uterus. Then, the embryos were Received August 18,1994;accepted February 15,1995. Address reprint requests to Dr. A.C.G. Wenink, Department of Anatomy, PO Box 9602,2300RC Leiden, the Netherlands. 94 M.W.M. KNAAPEN ET AL. fixed by perfusion in the right atrium with half strength Karnovsky buffered in Cacodylate, pH 7.2. After perfusion the embryos were fixed by immersion in the same fixative for 24 hr. Complete embryos of 11 days were washed in Cacodylate buffer and embedded in Epon 812. From older embryos, only the hearts were embedded. After the embedding procedure, they were serially sectioned at 2 Fm and stained with toluidineblue. Morphologic Determination of Cardiac Segments The boundaries of the individual segments and intersegmental junctions of the heart were morphologically defined in the sectioned embryos. In Figures 1 and 2, light microscopic photographs of hearts from 11 days and 14 days of development are shown. A reconstruction of a 12 day old embryonic heart shows the successive arrangement of the individual segments and intersegmental junctions (Fig. 3). At day 11 post coitum, the sinus venosus is that part of the heart which receives all the veins of the embryo. The sinu-atrial junction is the myocardium that lies between the sinus venosus and atrium. In the young embryos, where the sinus venosus has two sinus horns, the sinu-atrial junction is a constricted area, a t the site where the lumina of the sinus horns discharge into the lumen of the atrium. When the sinus venosus incorporates into the atrium, the sinu-atrial junction becomes a double layer of cells and looks like two valves between sinus venosus and atrium. The atrium is the segment which receives the blood from the sinus venosus in the younger embryos. In the older embryos, where the atrium is septated, the blood from the venae cavae enters that part of the right atrium which is derived from the incorporated sinus venosus. The right atrium is connected with the left atrium, receiving the lung veins, by the oval foramen. The atrio-ventricular junction is the canal lying between atrium and ventricular inlet segment. In the younger embryos, 11and 12 days post coitum, the atrio-ventricular canal is a tubular structure containing cushion tissue. In the older embryos, the myocardium of the atrio-ventricular canal is still recognized by the location of these cushions, although the canal is relatively shorter. Furthermore, in these stages of development the cells of the atrio-ventricular canal show several cytoplasmic extensions, whereas the cells of the atrium and inlet segment are more rounded. The ventricular part of the heart consists of two separate segments. The inlet segment is the compartment which receives the blood from the atrium via the atrio-ventricular canal in the very young embryos. In older embryos only the blood of the left atrium is received by the inlet segment via the left part of the atrio-ventricular junction. The inlet segment has apical trabeculations. The primary fold is the junction lying between the inlet segment and outlet segment. It lies in a more or less sagittal plane and it delimits the primary foramen, i.e., the communication between the ventricular inlet and outlet segments. In the inner curvature of the heart tube, the primary fold is continuous with the right side of the atrio-ventricular wall. In the outer curvature of the heart tube, the primary fold corresponds with a groove between the two ventricular segments, when seen from the outside of the heart. The primary fold develops into the larger part of the ven- Fig. 1. In the embryonic heart of a 11-day rat embryo, the atrium (Atr) is connected to the inlet segment (Inl) by the atrio-ventricular canal (AV). The primary fold (PF) forms the junction between inlet segment and proximal outlet segment (PO). The distal outlet segment (DO) contains endocardial cushion tisuue. x 125. tricular septum in the cardiac apex. The outlet segment can be divided in two parts, the proximal outlet segment, which is trabeculated as the immediately neighbouring inlet segment, and the distal outlet segment which is smooth and contains the endocardial outlet ridges. This segment is connected with the mesenchymal arterial trunk, which gives rise to the great arteries. Stereology The total myocardial volumes of the hearts were estimated by the Cavalieri method (Gundersen and Jensen, 1987). We counted the number of points on a grid hitting the myocardium in ten sections, taken systematically at equal distances. The magnification was x 40 and the point area of the point grid used was 225 mm'. The myocardial volumes of the individual segments were also estimated using this method. For the large segments such as atrium, ventricular inlet, and outlet segment, we used a point area of 225 mm2 and a magnification of x 40.For the smaller segments, sinus venosus, sinu-atrial junction, atrio-ventricular canal, and distal outlet segment the magnification was x 100 and the point area was 25 mm'. In the smallest embryos, only this latter grid and magnification were used. Calculation of the total myocardial volume and the myocardial volumes of the individual segments was done by Cavalieri's formula: Volume = M - 2x a x d x ZP Here M is the magnification, a is the point area in mm', d is the distance between the counted sections in MYOCARDIAL GROWTH 95 Fig. 2. In the embryonic heart of a 14-day rat embryo, the atrium (Atr) is connected to both inlet segment (Inl) and proximal outlet segment (PO). The sinus venosus (SV) incorporates into the right atrium where the sinu-atrial (SA) junction functions as the venous valves. The distal outlet segment (DO) contains still endocardia1cushion tissue. ~ 4 0 . mm, and X P is the total of counted points on the ten sections. The absolute volumes of the myocardium of the entire heart and of the individual cardiac segments were examined by the statistical package SPSS (version 5.0).In the statistical analysis, the developmental stages were fixed in days of gestation according to Berkson (Snedecor and Cochran, 1980). Multivariate analysis of variance with a significance level of 0.05 was used to compare the growth curves of the individual segments. RESULTS The Total Myocardial Growth During the developmental stages from 11 days to 17 days post coitum, the total myocardial volume showed an exponential growth curve (Fig. 4). The myocardial volume increased from 0.0416 mm3 a t day 11 to 2.7785 mm3 a t day 17. In Figure 5 the total myocardial volume for each developmental day studied is given on a logarithmic scale. It seems to show a change of slope a t day 14. This suggests that there are two different growth rates. The growth rate from day 11 to day 14 differs with slight significance (Manova, P = 0.034) from the growth rate between 14 and 17 days of development. The Myocardial Growth of the Individual Segments The growth curves of the myocardial volumes of the individual segments are shown on a linear and logarithmic scale respectively (Figs. 6, 7). In Table 1, the mean myocardial volumes of the individual segments Fig. 3. 3-D Graphic reconstructions (ventral view) to show the sequential arrangement of the cardiac segments and intersegmental junctions of a 12-day rat embryo in relation to each other. The compartments are shown from the venous to the arterial pole, and in each figure the more downstream region has been added. a: sinus venosus (SV). b sinus venosus and sinu-atrial junction (SA). c: the same with addition of the atrium (Atr). d the atrio-ventricular canal (AV) has been added. e: the inlet segment (Inl) has been connected with the atrio-ventricular canal. f: the same with addition of the primary fold (PF).Note that, in the inner curvature, the primary fold is continuous with the atrio-ventricular canal (see e). g: the same with addition of the proximal outlet segment (PO). h the complete heart after addition of the distal outlet segment (DO). for each developmental day studied are given. The increase in myocardial volume of the sinu-atrial junction did not differ significantly from that of the sinus veno- 96 M.W.M. KNAAPEN ET AL. Volume (mm3) Volume (mm3) 10 r ll .i...i 0.5 0 10 0.1 11 12 13 14 Age in days 15 1 I 16 17 0.01 10 11 12 4 13 14 15 16 17 5 Age in days Volume (mm3) Volume (mm3) 1 0.1 0.01 0.4 0.001 0.2 0 1 - 4.0001 10 11 12 13 ’ 14 1 15 16 Age in days ‘SV Fig. 4.The growth of the total myocardial volume and its standard deviation of the embryonic rat heart in mm3 from day 11 to day 17 post coitum. Fig. 5. As Figure 4, with the volumes given on a logarithmic scale. *SA OATR B A V *INL +PF 17 7 *PO +DO tersegmental junctions in mm3 from 11 day to 17 day embryonic rathearts. SV, sinus venosus; SA, sinu-atrial junction; Atr, atrium; AV, atrio-ventricular canal; Inl, inlet segment; PF, primary fold; PO, proximal outlet segment; DO, distal outlet segment. Fig. 7. As Figure 6, with the volumes given on a logarithmic scale. Fig. 6. The myocardial volumes of the individual segments and in- sus and neither from the atrium although the inter- when the myocardial volume of the primary fold was cepts were of course different. The growth curves of the added to the proximal outlet segment the growth three main compartments; atrium, inlet segment, and curves of the inlet segment and the enlarged proximal proximal outlet segment are not identical. The curves outlet segment differed significantly also (P = 0.005). of the proximal outlet segment and the inlet segment The growth curves of atrio-ventricular and distal outlet are significantly different (P = 0.022). The primary segment were remarkable (Fig. 7). They differed with fold, including the ventricular septum, differs signifi- small significance (P = 0.042) from each other, but cantly from the atrium (P < 0.001), inlet segment (P = they differed with high significance (P < 0.001) from 0.002), and proximal outlet segment (P = 0.015) (Fig. the other segments and intersegmental junctions. The relative growth of the individual segments is 8). However, there are reasons to consider the primary fold, giving rise to the ventricular septum, as part of evaluated in Figure 10, where each individual volume the inlet segment. Thus, when the myocardial volume is expressed as the percentage of the total myocardial of the primary fold was added to the myocardial volume volume. The relative myocardial volumes of sinus of the inlet segment, the resulting curve did not differ venosus and sinu-atrial junction remained the same significantly from the growth curve of the atrium and during the period studied. The residual values of the that of the proximal outlet segment (Fig. 9). However, atrium to the total myocardial volume did not differ 97 MYOCARDIAL GROWTH TABLE 1. The myocardial volumes of the individual segments and intersegmental junctions in mm3 at each day of gestation in the period studied' Age 11 12 13 14 15 16 17 S.V. S.A. Atr. A.V. 0.0005 0.0015 0.0036 0.0073 0.0162 0.0173 0.0317 0.0004 0.0015 0.0025 0.0080 0.0141 0.0195 0.0317 0.0077 0.0195 0.0585 0.1525 0.2774 0.4949 0.6104 0.0059 0.0106 0.0119 0.0241 0.0308 0.0373 0.0436 Inl. 0.0120 0.0246 0.0504 0.1483 0.3012 0.4835 0.7095 P.F. 0.0014 0.0039 0.0127 0.0518 0.1055 0.1881 0.4122 P.O. 0.0091 0.0157 0.0522 0.1319 0.3002 0.5095 0.8601 D.O. 0.0047 0.0101 0.0229 0.0444 0.0681 0.0882 0.0793 'SV, Sinus Venosus; SA, Sinu-Atrial junction; Atr, Atrium; AV, Atrio-Ventricular canal; Inl, Inlet segment; PF, Primary Fold; PO, Proximal Outlet segment; DO, Distal Outlet segment. significantly indicating that the relative growth curve of the atrium shows the same evolution as the total myocardial volume although there was a small increase of the relative atrial volume between day 11and day 14, and a small decrease from day 16 to day 17. The inlet segment showed an equal contribution to the myocardial volume of nearly 27% in all the developmental stages. However, it is remarkable that a t day 11,the inlet segment showed the highest proportion of the total myocardial volume, which is 0.01196 mm3 in absolute myocardial volume. The proximal outlet segment increased slightly from day 11to day 17 of development, and its growth curve differed not significantly from that of the total myocardial volume. An interesting phenomenon is shown by the junctions atrio-ventricular canal, primary fold and distal outlet segment. The atrio-ventricular canal showed a rapid decrease from 14% at day 12 to 4% a t day 14, whereafter the relative growth of the atrio-ventricular canal declined slowly. Between day 12 and 14 the relative myocardial volume of the primary fold increased from 4% at day 12 to 10% a t day 14. The distal outlet segment showed a gradual decline during the developmental stages studied. DISCUSSION Our data show a steady increase of both the total myocardial volume and the myocardial volumes of the individual segments and junctions. The total myocardial volume increases from 0.0416 mm3 at day 11 to 2.7785 mm3 at day 17. The growth rate of the total myocardial volume is higher between day 11 and day 14 than from day 14 to day 17. This supports the data of Goedbloed (1972) who found that in the developing rat embryo there are changes in growth rates. He suggested that these changes correlate with the changes in cell activity and cell differentiation. Around day 14, Goedbloed (1972)found a sudden change in growth rate of the volume of the entire rat embryo. Mandarim-deLacerda (1991) found also a change in growth rate, in human embryos, when the volumes of the hearts were plotted against the crown-rump length. He suggested that this phenomenon could be due to a more intensive ventricular growth to supply the entire embryo with blood. The systolic pressure shows a linear relationship to the embryonic weight (Van Mierop and Bertuch, 1967) and the volume of the entire embryo correlates linearly with the total myocardial volume of the developing heart (Clark et al., 1986). Goedbloed (1972) re- ported a growth change of the entire embryo at day 14 post coitum. If this change in growth rate is correlating to cardiovascular changes which reflect increasing blood volume and blood pressure, then typical differentiation processes in specific areas of the developing heart have to be hypothesized. It is remarkable that around day 14 the septation has been reported to have proceeded to such an extent that a separate right blood stream into the outlet is evolving (Wenink et al., 1994). Thus, systolic pressure increases a t that time, especially in the ventricles (Van Mierop and Bertuch, 1967). Besides, the amount of myofibrils increases and they become more orientated, whereas the volume of the cytoplasm relatively decreases in the myocytes of the ventricular wall (Manasek, 1970). Therefore, the developmental regulation of myosin heavy chain isoforms could change the contractile characteristics of the developing atria and ventricles in response to changes in blood volume and blood pressure during development, as supposed by Evans (1988). Sinus Venosus and Sinu-Atrial Junction The myocardial volumes of the sinus venosus and sinu-atrial junction as percentages of the total myocardial volume remain the same, whereas the absolute myocardial volumes of these segments show a slight increase during the period studied. Also the sinu-atrial junction shows the same growth curve as the atrium, indicating that this junction is partly sinus venosus and partly atrium. In the young embryos, the sinuatrial junction does not seem to function to avoid backward flow from the atrium to the sinus venosus. In the older embryos the sinu-atrial junction is partly transformed to the venous valves due to incorporation of the sinus venosus into the right atrium (Odgers, 1935; Wenink, 1987; Steding, 1990). In this stage of development, the venous valves might have acquired the function of avoiding backward flow, because the blood pressure in the atrium increases, although it is much lower than in the ventricular part (Faber, 1968). The right venous valve gives rise to the Eustachian and Thebesian valves, whereas the left venous valve fuses partly with the atrial septum. However, these cardiac components do not seem to be of great functional importance and morphological variations of the valvular apparatus in the right atrium such as the absence of the Thebesian valve are frequent (Steding, 1990; Felle and Bannigan, 1994). Our own measurements are of course inadequate to reflect such variations. 98 M.W.M. KNAAPEN ET AL. Atrium, Inlet, and Proximal Outlet Segment Volume (mm3) 'r I 1 0.01 O.lI i I 0.001 L 10 11 12 13 14 15 Age in days O- ATR * INL + PF * P O Volume (mm3) lor I 0.001 10 1 11 I 13 12 14 15 Age in days 0 ATR f PO + IP Proportional volume (%) 35 r 15t Undoubtedly, these segments must show functional changes during development. In the straight heart tube stage, when atrium, inlet, and proximal outlet segment are sequentially arranged to each other (Wenink, 19871, the myocardium of the heart tube shows peristaltoid contractions (Goss, 1938). By day 11 of development, that is the youngest stage studied presently, the heart has already looped but it maintains the sequential arrangement of those segments. As we have shown, the inlet segment has then the highest proportion of the total myocardial volume. In this stage the proximal outlet segment hardly collects any blood from the atrium via the atrio-ventricular canal and the inlet segment directs all blood to the distal outlet segment via the primary foramen. Therefore, in this stage of development the inlet segment is probably the main contraction compartment to supply the embryo with blood. From 13 days onwards the cushions of the atrio16 17 ventricular canal direct also a part of the bloodflow into the proximal outlet segment through a developing groove from the atrio-ventricular canal into the proxi8 mal outlet segment (Wenink et al., 1994). From that time the proximal outlet segment, too, starts to function like a contraction compartment which directs the bloodflow to the distal outlet segment. This might explain our observation (Fig. 10) that the proximal outlet segment is steadily increasing its proportional volume. In later stages of development the embryo gradually requires a greater blood volume to supply all the tissues of the embryo as well as an increase in blood pressure (Van Mierop and Bertuch, 1967). The suggestion that the atrium functions as the collecting compartment whereas inlet and proximal outlet segment act as the contraction compartments is supported by the fact that inlet and proximal outlet segment show higher and longer action potentials than the atrium does (Paff and Boucek, 1962; Agata et al., 1993). The rate of the action potentials increases gradually as more myocarI dium becomes contractile (GOSS,1938). 16 17 Apparently, the functional changes of the heart tube are as follows. The straight heart tube shows peristal9 toid contractions. The venous part has then the greatest contractility (Hall, 1954). After looping, a more synchronous contraction pattern develops which is reflected by changes in myosin heavy chain expression patterns (de Jong et al., 1987). As we have shown, the ventricular inlet segment has the greatest relative myocardial volume in these stages. Gradually, the Fig. 8. The myocardial volumes of the atrium and both ventricles, (i.e., inlet and proximal outlet) and the primary fold on a logarithmic scale in relation to the developmental stages from 11-day to 17-day rat embryos. EL- 10 5 0 10 11 12 13 14 15 16 Age in days -SV *SA O A T R B A V *INL Figs. 8-10. +PF 17 10 *PO *DO Fig. 9. The myocardial volumes of atrium, inlet segment including the myocardial volume of the primary fold (IP), and proximal outlet segment during the period studied on a logarithmic scale in relation to the developmental stages from 11-day to 17-day rat embryos. Fig. 10. The ratio of the individual cardiac segments and intersegmental junctions to the total myocardial volume in the period from 11-day to 17-day post coitum. SV, sinus venosus; SA, sinu-atrialjunctions; Atr, atrium; AV, atrio-ventricular canal; Inl, inlet segment; PF, primary fold; PO, proximal outlet segment; DO, distal outlet segment. MYOCARDIAL GROWTH 99 which squeeze the endocardial cushions together, so as to avoid backward flow (Goerttler, 1955; Jaffee, 1965). Indeed, the myocardial walls of the atrio-ventricular canal and distal outlet segment have been reported to show a slow conduction velocity and slow contraction Primary Fold (de Jong et al., 1992). The primary fold is a special junction because it gives By day 13, septation has proceeded to create two rise to the main part of the ventricular septum. There- blood streams (Rogers and Morse, 1986; Wenink, 1987). fore, in relevant stages, we have included the ventric- From that time onwards the formation of valves at the ular septum in our counts of the primary fold. The atrio-ventricular and ventriculo-arterial junctions bebasal portion of the fold, in the inner curvature of the comes more important in avoiding backward flow of heart, is not supposed to grow significantly, but our the blood when the pressures in both ventricles start to data fail to show this because all of the fold was increase (Van Mierop and Bertuch, 1967; Clark et al., counted as one single structure. The growth curve of 1986).In the human embryo, the atrioventricular cushthe primary fold shows a remarkable increase of the ion tissue has been reported to grow only for a limited volume from day 11to day 17. period (Mandarim-de-Lacerda, 1991; Wenink, 1992), Before day 14 of development the volume shows the whereas the distal outlet segment may become thicker, greatest increase. This period corresponds with the but without changing its length (Thurkow and time when the atrio-ventricular canal starts to direct Wenink, 1993). All these data point a t a diminishing the blood into both the inlet and proximal outlet seg- role of the atrio-ventricular canal and the distal outlet ments (Lamers et al., 1992; Wenink et al., 1994). As is segment as temporary sphincters. Our present findshown in Figures 8 and 10, the primary fold differs in ings, in particular the steep decrease of the relative growth rate from the growth rate of the inlet segment myocardial volumes between day 12 and day 14 (Fig. 6) as well as the proximal outlet segment and the growth support the notion of a changing function in this develrate of the total myocardial volume. It is important to opmental period. note that the growth rates of inlet segment and proxiCONCLUSIONS mal outlet segment are also different from the total myocardial volume and from each other. We have deSeveral important conclusions can be extracted from liberately considered inlet segment, primary fold and the growth curves. 1)The sinus venosus and sinu-atrial proximal outlet segment as three separate entities. junction grow proportional to the total myocardial volThus, the volume increase of the primary fold at 14 ume. They remain relatively small components, but days is supposed to reflect ventricular septation. Goor there is no change in proportion during development. (1970) has suggested that ventricular septation takes 2) A similar exponential growth is seen in atrium, inlet place by expansion of the two neighbouring ventricular segment, and proximal outlet segment. These are the cavities. This would mean an equal contribution from components that do most of the work of the heart. In both the inlet segment and the proximal outlet seg- the youngest stages, the inlet segment is the most imment. However, the different growth curves of the en- portant segment to propel the blood into the distal outtities allow still another hypothesis. If the volumes of let segment. 3) The atrio-ventricular canal and distal primary fold and inlet segment are added and consid- outlet segment show conspicuously less growth than all ered as one entity, then this new entity no longer has a other components. However, their percentage of the todifferent growth curve from either that of the proximal tal volume is relatively high at day l l , indicating their outlet segment or that of the total myocardial volume importance in squeezing the endocardial cushions to(Fig. 9). This suggestion that the ventricular septum gether to avoid backward flow in a heart yet without would belong more to the left ventricle than to the valves. 4) The primary fold, although being a juncright is also supported by the myofiber architecture in tional structure, does not have the same constricting the mature heart (Anderson and Becker, 1980). function as the atrio-ventricular canal and distal outlet segment a t day 11. It can be considered as part of the Atrio- Ventricular Canal and Distal Outlet Segment wall of the inlet segment and gives rise to the main The slow growth of the atrio-ventricular canal and part of the ventricular septum. distal outlet segment, i.e., the decrease of a high relaACKNOWLEDGMENTS tive volume at day 11to a lower proportion at day 17, indicates that their function may be important only This study was supported by grant 90288 from the during a limited period of cardiac development. These Netherlands Heart Foundation. The authors gratefully are the regions in the developing heart which contain acknowledge the help of Dr E.A. van der Velde, Deendocardial cushion tissue. Before valve formation partment of Medical Statistics, who reviewed the stathey must have a particular contraction pattern. The tistical part of this study. ventricular segments contract faster than the atrium does, and in addition the repolarization phenomenon of LITERATURE CITED the atrium occurs simultaneously with depolarization Agata, N., H. Tanaka, and K. Shigenobu 1993 Developmental of the ventricle (Paff and Boucek, 1962). Also the changes in action potential properties of the guinea-pig myocardium. Acta Physiol. Scand., 149:331-337. atrium has a lower blood pressure than the ventricular part (Faber, 1968). Thus, in a heart which does not yet Anderson, R.H., and A.E. Becker 1980 Cardiac Anatomy. Churchill Livingstone, Edinburgh London New York, 5.14-5.26. possess atrio-ventricular valves and ventriculo-arterial Clark, E.B., N. Hu, J.L. Dummett, G.K. Vandekieft, C. Olson, and R. valves the myocardial walls of atrio-ventricular canal Tomanek 1986 Ventricular function and morphology in chick embryo from stages 18 to 29. Am. J. Physiol., 250:H407-H413. and distal outlet segment function like sphincters atrial and ventricular segments develop their own specific myosin isoform expression pattern (de Jong et al., 1987), and we believe that this functional evolution is also illustrated by the present data. 100 M.W.M. KNAAPEN ET AL. Evans, D., J.B. Miller, and F.E. Stockdale 1988 Developmental patterns of expression and coexpression of myosin heavy chains in atria and ventricles of the avian heart. Dev. Biol., 127:376-383. Faber, J.J. 1968 Mechanical function of the septating embryonic heart. Am. J . Physiol., 3:475-481. Felle, P., and J.G. Bannigan 1994 Anatomy of the valve of the coronary sinus (Thebesian valve). Clin. Anat., 7:lO-12. Goedbloed, J.F. 1972 Embryonic and postnatal growth of rat and mouse. Acta Anat., 82:305-336. Goerttler, K. 1955 Uber Blutstromwirkung als Gestaltungsfaktor fur die Entwicklung des Herzens. Beitrage Path. Anat., 115:33-56. Goor, D.A., J.E. Edwards, and C.W. Lillehei 1970 The development of the interventricular septum of the human heart: Correlative morphogenetic study. Chest, 58:453-467. Goss, C.M. 1938 The first contractions of the heart in rat embryos. Anat. Rec., 70505-524. Gundersen, H.J.G., and E.B. Jensen 1987 The efficiency of systematic sampling in stereology and its prediction. J . Microsc., 147:229263. Hall, E.K. 1954 Further experiments on the intrinsic contractility of the embryonic rat heart. Anat. Rec., 118:175-184. Jaffee, O.C. 1965 Hemodynamic factors in the development of the chicken embryo heart. Anat. Rec., 151:69-76. de Jong, F., W.J.C. Geerts, W.H. Lamers, J.A. Los, and A.F.M. Moorman 1987 Isomyosin expression patterns in tubular stages of chicken heart development: A 3-D immunohistochemical analysis. Anat. Embryol., 177231-90. de Jong, F., T. Opthof, A.A.M. Wilde, M.J. Janse, R. Charles, W.H. Lamers, and A.F.M. Moorman 1992 Persisting zones of slow impulse conduction in developing chicken hearts. Circ. Res., 71: 240-250. Lamers, W.H., A. Wessels, F.J. Verbeek, A.F.M. Moorman, S. Viragh, A.C.G. Wenink, A.C. Gittenberger-de Groot, and R.H. Anderson 1992 New findings concerning ventricular septation in the human heart. Circulation, 86:1194-1205. Manasek, F.J. 1970 Histogenesis of the embryonic myocardium. Am. J. Cardiol., 25:149-168. Mandarim-de-Lacerda, C.A. 1991 Growth allometry of the myocardium in human embryos (from stages 15 to 23). Acta Anat., 141: 251-256. Odgers, P.N.B. 1935 The formation of the venous valves, the foramen secundum and the septum secundum in the human heart. J. Anat., 69:412-422. Paff, G.H., and R.J. Boucek 1962 Simultaneous electrocardiograms and myograms of the isolated atrium, ventricle and conus of the embryonic chick heart. Anat. Rec., 142:73-79. Rogers, C.S., and D.E. Morse 1986 Atrial septation in the rat: A light microscopic and histochemical study. J . Submicrosc. Cytol., 18: 313-324. Snedecor, G.W., and W.G. Cochran 1980 Statistical Methods, 17th edition. The Iowa State University Press. Steding, G., X. Jinwen, W. Seidl, J. Manner, and H. Xia 1990 Developmental aspects of the sinus valves and the sinus venoms septum of the right atrium in human embryos. Anat. Embryol., 181: 469-475. Thurkow, E.W., and A.C.G. Wenink 1993 Development of the ventriculoarterial segment of the human embryonic heart: A Morphometrics Study. Anat. Rec., 236:664-670. Van Mierop, L.H.S., and C.J. Bertuch 1967 Development of arterial blood pressure in the chick embryo. Am. J. Physiol., 212:43-48. Wenink, A.C.G. 1987 Embryology of the heart. In: Paediatric Cardiology. R.H. Anderson, E.A. Shinebourne, F.J. Macartney, and M. Tynan, eds. Churchill Livingstone, Edinburgh. Wenink, A.C.G. 1992 Quantitative morphology of the embryonic heart: An approach to development of the atrioventricular valves. Anat. Rec., 234:129-135. Wenink, A.C.G., L.J. Wisse, and P.M. Groenendijk 1994 Development of the inlet portion of the right ventricle in the embryonic rat heart: The basis for tricuspid valve development. Anat. Rec., 239: 216-223.