Nguemo Filomain, Fleischmann Bernd K, Schunkert Heribert, Hescheler Juergen, Reppel Michael
Institute of Neurophysiology, University of Cologne, Cologne, Germany.
Cell Physiol Biochem. 2007;20(6):809-24. doi: 10.1159/000110441.
BACKGROUND/AIMS: The expression and regulation of Ca2+ signaling in embryonic cardiomyocytes has been shown to be different from those in adult heart cells, particularly the L-type Ca2+ channel current (I(CaL)) increases during development. However, little is known about the underlying reasons for this increase of I(CaL ) density and developmental changes in the process of I(CaL ) inactivation, a critical regulator of intracellular Ca2+ homeostasis. In the present work, we therefore studied functional differences of I(CaL) between embryonic and fetal cardiomyocytes and its interaction with intracellular Ca2+ homeostasis and Ca2+-induced Ca2+ release (CICR). Moreover, we examined the process of voltage- (VDI) and Ca2+-dependent inactivation (CDI) of I(CaL) during murine embryonic heart development.
The electrophysiological characteristics of I(CaL ) inactivation were analyzed in embryonic ventricular cardiomyocytes of early (E9.5-11.5) and late developmental, fetal (LDS, E16.5-18.5) stages and of adult mice using the whole-cell patch-clamp technique.
Fast, Ca2+-dependent inactivation kinetics (tau(f)) were significantly accelerated in LDS-derived cardiomyocytes (2.53 +/- 1.43 ms, n=9) as compared to EDS (5.09 +/- 2.19 ms, n=8, p>=0.009), whereas slow, voltage-dependent inactivation time constants (tau (s)) were unchanged. In cardiomyocytes derived from LDS we observed an increase in the maximal gating charge (Q(max)), suggesting an increase in the number of L-type Ca2+ channels at the sarcolemma, whereas the channel open probability (P(o)) was unchanged. Accordingly a significantly higher I(CaL ) density was found in LDS (-14 +/- 2.26 pA/pF, n=14) versus> EDS-derived cardiomyocytes (-10.03 +/- 1.43 pA/pF, n=13, p)( 0.05). Since ryanodine (10 microM) failed to alter tau(f) at both developmental stages, a major contribution of CICR and bulk Ca2+ to the acceleration of the fast inactivation kinetics during heart development seems to be unlikely.
Our data suggest that the increase of local subsarcolemmal Ca2+, evoked by the higher expression of L-type Ca2+ channels at the sarcolemma, rather than bulk Ca2+ accelerates I(CaL ) inactivation during embryonic heart development.
背景/目的：胚胎心肌细胞中Ca2+信号的表达和调节已被证明与成年心脏细胞不同，尤其是L型Ca2+通道电流（I(CaL)）在发育过程中增加。然而，关于I(CaL)密度增加的潜在原因以及I(CaL)失活过程中的发育变化（细胞内Ca2+稳态的关键调节因子）知之甚少。因此，在本研究中，我们研究了胚胎和胎儿心肌细胞之间I(CaL)的功能差异及其与细胞内Ca2+稳态和Ca2+诱导的Ca2+释放（CICR）的相互作用。此外，我们研究了小鼠胚胎心脏发育过程中I(CaL)的电压依赖性失活（VDI）和Ca2+依赖性失活（CDI）过程。
使用全细胞膜片钳技术分析早期（E9.5 - 11.5）和晚期发育、胎儿期（LDS，E16.5 - 18.5）以及成年小鼠胚胎心室心肌细胞中I(CaL)失活的电生理特性。
与早期发育阶段（EDS，5.09 ± 2.19 ms，n = 8）相比，LDS来源的心肌细胞（2.53 ± 1.43 ms，n = 9）中快速、Ca2+依赖性失活动力学（tau(f)）显著加快（p >= 0.009），而缓慢的电压依赖性失活时间常数（tau(s)）不变。在LDS来源的心肌细胞中我们观察到最大门控电荷（Q(max)）增加，表明肌膜上L型Ca2+通道数量增加，而通道开放概率（P(o)）不变。因此，LDS来源的心肌细胞（-14 ± 2.26 pA/pF，n = 14）中的I(CaL)密度显著高于EDS来源的心肌细胞（-10.03 ± 1.43 pA/pF，n = 13，p < 0.05）。由于在两个发育阶段，10 μM的ryanodine均未能改变tau(f)，因此CICR和大量Ca2+对心脏发育过程中快速失活动力学加速的主要贡献似乎不太可能。
我们的数据表明，肌膜上L型Ca2+通道较高表达引起的局部肌膜下Ca2+增加，而非大量Ca2+，加速了胚胎心脏发育过程中的I(CaL)失活。
