Calcium Signaling, Metabolism, and EC Coupling in Heart

项目总结/摘要 申请人的长期目标是继续研究钙信号，兴奋收缩（EC） 偶联，以及这些过程对心肌能量代谢的依赖性。具体目标 研究兔和小鼠心室细胞（包括心脏特异性钠钙交换蛋白 基因敲除小鼠）以：1）研究来自梗塞周围的重塑细胞中的钙信号传导和EC偶联 兔子区这将包括评估L型钙通道功能丧失是否会 解释EC偶联的失败以及横小管、ryanodine和 二氢吡啶受体参与这些细胞的衰竭; 2）研究代谢的影响， 抑制兔心室肌细胞偶联体的功能和结构。这将包括 测量偶联体中L-型钙通道的最小数量以及代谢途径 抑制会影响它们的功能。特别是，钙火花和尖峰形成和细胞微结构的改变 横小管系统作为代谢过程中耦合子功能丧失的原因 3）研究钠-钙交换蛋白基因敲除小鼠对代谢抑制的抗性， 应力这将包括对代谢抑制阻止激活的假设的研究。 在野生型小鼠中逆转钠-钙交换，导致钙诱导的钙离子通道被破坏。 EC偶联的释放机制。相反，假设钠-钙交换体敲除 小鼠不需要钠-钙交换来进行EC偶联，因此对 代谢抑制抑制钠钙交换激活对钙峰的影响 将检查延迟。这些实验的目的之一是解释 触发过程中的二进制裂隙钙。方法包括测量钙峰的概率和它们的 在用代谢抑制剂处理之前和之后的兔和小鼠的潜伏期分布。此外 方法包括最近开发的用于重建血管的三维结构的程序。 横小管系统和兰尼碱和二氢吡啶受体的三维分布， 梗塞周围细胞和用代谢抑制剂处理的细胞。

Project Summary/Abstract The applicant¿s long-term aims are to continue studies on calcium signaling, excitation-contraction (EC) coupling, and the dependence of these processes on energy metabolism in cardiac muscle. The specific aims are to study ventricular cells from rabbits and mice (including cardiac-specific sodium-calcium exchanger knock-out mice) to: 1) investigate calcium signaling and EC coupling in remodeled cells from the peri-infarct zone in rabbits. This will include an assessment of whether loss of L-type calcium channel function can account for failure of EC coupling and whether significant alterations in transverse-tubules, ryanodine and dihydropyridine receptors are involved in the failure of these cells; 2) investigate the effect of metabolic inhibition on the function and structure of couplons in rabbit ventricular myocytes. This will include a measurement of the minimum number of L-type calcium channels in a couplon and the way that metabolic inhibition affects their function. In particular, alterations in calcium spark and spike formation and cellular microarchitecture of the transverse-tubule system as a cause of the functional loss of couplons during metabolic inhibition will be considered; 3) study the resistance of sodium-calcium exchanger knock-out mice to metabolic stress. This will include an investigation of the hypothesis that metabolic inhibition prevents activation of reverse sodium-calcium exchange in wild-type mice, resulting in disruption of the calcium-induced calcium release mechanism of EC coupling. In contrast, it is hypothesized that sodium-calcium exchanger knock-out mice do not require sodium-calcium exchange for EC coupling and are therefore resistant to the effects of metabolic inhibition. The consequences of inhibiting sodium-calcium exchange activation on calcium spike latency will be examined. These experiments are, among other things, designed to explain the importance of diadic cleft calcium in the trigger process. Methods include measuring calcium spike probabilities and their latency distributions in rabbits and mice before and after treatment with metabolic inhibitors. In addition the methods include recently developed procedures for reconstructing the 3-dimensional architecture of the transverse¿tubule system and the 3-dimensional distribution of ryanodine and dihydropyridine receptors in peri-infarct cells and cells treated with metabolic inhibitors.