Joint Study on Evolution of Skeletal Muscle E-C Coupling

高等脊椎动物骨骼肌兴奋-收缩偶联(E-C偶联)的一个显著特征是，收缩不需要钙内流。这是因为膜去极化直接触发了肌浆网(SR)中钙的释放。负责这一信号转导的分子是一种二氢吡啶受体，它被认为负责电压传感器(潜在的E-C偶联)和L类型的钙通道的双重功能。在无脊椎动物肌肉纤维的E-C偶联中，外源钙离子内流是收缩所必需的，并且存在DHP受体，但只负责L型钙离子通道功能。因此，高等脊椎动物的E-C耦合机制与无脊椎动物的E-C耦合机制有质的区别。必须有一个进化的步骤，以获得新的去极化诱导的钙释放机制。主对象…本研究更多的是考察代表动物进化不同阶段的物种，并比较E-C耦合的生物物理和形态特征。我们报告说，进化步骤可能发生在文昌鱼(Branchiostoma Lanceolatum)和七鳃鳗(Lampetra Plan Eri)之间的低等脊索动物。在单个快肌纤维上的全细胞电压钳实验表明，这种进化变化伴随着膜内电荷运动成分的出现，该成分被DHP的衍生物硝苯地平所阻断，并显著减慢了钙通道的激活动力学。在七鳃鳗、小鼠和角鱼的纤维中，细胞内钙的释放(使用敏感的荧光染料FUF3测量)与膜去极化有关，对外部施加的钴耐受，并被硝苯地平抑制。而文昌鱼纤维中的Ca~(2+)释放则被Co~(2+)抑制。这些结果有力地支持了这一假说，即硝苯地平敏感的膜内电荷运动是脊椎动物骨骼肌E-C偶联的信号转导机制。海洋无脊椎动物和低等脊椎动物代表着这种不同物质的独特‘基因库’，应该为我们提供关于DHP-受体分子进化的年代学证据。较少

A distinctive feature of the excitation-contraction coupling (E-C coupling) of skeletal muscle in higher vertebrates in that Ca^<2+> -influx is not necessary for contraction. This is because membrane depolarisation directly triggers Ca^<2+> release from the sarcoplasmic reticulum (SR). The molecule responsible for this signal transduction in the transverse tubular (T-tubular) membrane is a dihydropyridine (DHP) receptor, which is thought to be responsible for the dual function, of voltage sensor (underlying E-C coupling) and the L-type Ca^<2+> channel. In E-C coupling in invertebrate muscle fibers, influx of external Ca^<2+> is necessary for contraction, and a DHP -receptor is present, but responsible for only the L-type Ca^<2+> channel function. Therefore, the E-C coupling mechanism in higher vertebrates is qualitativele different from that in invertebrates. There must have been an evolutionary step to acquire the novel depolarisation-induced Ca^<2+> release mechanism. The main object … More ive of the present research project is to examine species representing different stages in animal evolution, and compare the biophysical and morphological properties of E-C coupling.We reported that the evolutionry step might have occurred in lower chordates between amphioxus (Branchiostoma lanceolatum) and lamprey (Lampetra planeri). Whole-cell voltage clamp experiments on single fast muscle fibers suggest that this evolutionary change is accompanied by the appearance of a component of intramembrane charge movement, which is blocked by nifedipine, a DHP derivative, and by a remarkable slowing-down of the activation kinetics of the Ca^<2+> channel. In the fibers of lamprey, mouse, and dogfish, intracellular Ca^<2+>-release (measured using the Ca^<2+> sensitive flurescence dye fluo3) is associated with membrane depolarisation, is resistant to externally applied Co^<2+>, and is suppressed by nifedipine. In contrast, in the amphioxus fiber, Ca^<2+>-release is suppressed by Co^<2+>. The results strongly support the hypothesis that nifedipine-sensitive intramembrane charge movement is the signal tansduction mechanism of vertebrate skeletal muscle E-C coupling.Marine invertebrates and lower vertebrates represent a unique 'gene-bank' of such diverse material and should provide us with chronological evidence about the molecular evolution of the DHP-receptor. Less