Reorganization of calcium signaling in heart failure

描述（由申请人提供）：本研究将主要在斯洛伐克共和国进行，作为NIH资助#HL074045的扩展。本研究的总体目标是阐明心力衰竭时哺乳动物心肌细胞二分体中钙信号机制的变化。这些结构含有L型钙通道（DHPR）簇，与钙离子功能性偶联的兰尼碱受体钙释放通道簇相反。兴奋-收缩偶联过程中钙信号的改变有助于衰竭心脏的性能下降。将确定在DHPR激活钙释放过程中钙信号步骤的变化及其与二联体的超微结构组织和兴奋-收缩偶联单元中通道的拓扑结构的关系。为此，将对全细胞钙电流、全细胞钙瞬变和二进钙释放事件（钙峰）的动力学进行测量。这些测量将补充的体积和表面密度的管状系统和池和纵向肌浆网的形态学分析，和共定位之间的钙通道和钙释放通道，使用电子显微镜和共聚焦显微镜。钙电流和钙释放的动力学的定量分析将被用于估计对兴奋-收缩偶联的分级和效率的控制至关重要的钙信号传导的参数。计算机模拟实验将用于确定可能的机制，并设计具有高预测能力的实验。了解心脏电-心偶联的异常在心力衰竭中是重要的，因为它可能成为治疗干预的战略位点。此外，定义通道簇的空间组织和它们的钙信号之间的关系对于理解诸如突触传递、神经内分泌分泌和血管张力调节的过程具有更广泛的意义。

DESCRIPTION (provided by applicant): This research will be done primarily in Slovak Republic as an extension of NIH grant #HL074045. The overall goal of this proposal is to elucidate the changes of calcium signaling mechanisms in dyads of mammalian cardiac muscle cells during heart failure. These structures contain clusters of L-type calcium channels (DHPRs) in opposition to clusters of ryanodine receptor calcium release channels functionally coupled by calcium ions. Alterations in calcium signaling during excitation-contraction coupling contribute to the decreased performance of failing hearts. The changes in calcium signaling steps during activation of calcium release by DHPRs and their relationship to the ultrastructural organization of dyads and the topology of channels in the excitation-contraction coupling units will be determined. To this end, measurements of the kinetics of whole-cell calcium currents, whole-cell calcium transients, and dyadic calcium release events (calcium spikes) will be carried out. These measurements will be supplemented by morphological analysis of the volume and surface densities of the tubular system and of cisternal and longitudinal sarcoplasmic reticulum, and of colocalization between calcium channels and calcium release channels, using electron microscopy and confocal microscopy. Quantitative analysis of the kinetics of calcium current and calcium release in parallel will be used to estimate parameters of calcium signaling critical for control of gradation and efficiency of excitation-contraction coupling. Computer simulation of experiments will be used to ascertain possible mechanisms and to design experiments with high predictive power. Understanding the abnormalities of cardiac E-C coupling in heart failure is important since it might become a strategic site for therapeutic intervention. Furthermore, defining the relationships between the spatial organization of channel clusters and their calcium signaling has a broader significance for understanding processes such as synaptic transmission, neuroendocrine secretion and regulation of vascular tone.