Investigate the effect of mitochondrial energy state on Ca2+ sparks and handling

心脏病是美国人的最大杀手。越来越多的证据表明， 钙离子调节异常与心脏病的发生发展密切相关。因此 了解病理条件下细胞内钙信号的调控是非常重要的。 钙诱导钙释放的局部调控依赖于L类钙离子的空间组织 通道和Ryanodine受体(RyR)在二分体。类似地，线粒体对钙的摄取 由于它们非常接近钙离子释放部位，这是刺激氧化所需的过程 工作变化过程中的磷酸化。然而，线粒体对CICR的反馈还不太清楚。 由于线粒体是活性氧物种(ROS)的主要来源，它们可能会潜在地影响 胞质氧化还原状态，进而改变RyR开放概率。在这项拟议的研究中，一种双光子激光器 显微镜系统将被用来直接检查能量状态如何动态发生剧烈变化 在不同实验条件下对钙离子火花特性的影响。细胞内钙离子(或ROS)，A^m， 和NADH将同时记录在分离的豚鼠心肌细胞上，并离线分析 已成像。线粒体之间的时空耦合！去极化和钙离子火花将 用定量的方法进行分析。此外，还建立了线粒体和钙离子的计算模型 将开发释放单元来定量研究线粒体能量学之间的相互作用 和局部钙离子的处理。最后，建立了心肌细胞与底物结合的完整模型 新陈代谢、细胞电生理学、pH调节和E-C偶联将被用来研究 能量产生、离子通道、钙处理和pH变化的潜在机制，以及 在缺血-再灌流期间心脏收缩功能的降低。通过将 实验和计算结果，这些研究将允许一个完整的理解起源 缺血后损伤和心力衰竭的发展，并显著刺激新心脏的发展 疾病疗法。

Heart disease is the single largest killer of the American. A growing body of evidence has shown that there is a close relationship between Ca2+ handling abnormalities and development of heart disease. Therefore it is fundamentally important to understand the regulation of Ca2+ signaling under pathological conditions. Local control of Ca2+-induced Ca2+ release (CICR) depends on the spatial organization of L-type Ca2+ channels and ryanodine receptors (RyR) in the dyad. Analogously, Ca2+ uptake by mitochondria is facilitated by their close proximity to the Ca2+ release sites, a process required for stimulating oxidative phosphorylation during changes in work. Mitochondrial feedback on CICR, however, is less well understood. Since mitochondria are a primary source of reactive oxygen species (ROS), they could potentially influence the cytosolic redox state, in turn altering RyR open probability. In this proposed study, a two photon laser microscope system will be used to directly examine how acute changes in energy state dynamically influence Ca2+ spark properties under various experimental conditions. Cytosolic Ca2+ (or ROS), A^m, and NADH will be recorded simultaneously in isolated guinea pig cardiomyocytes and analyzed offline using imaged. The spatiotemporal coupling between mitochondria! depolarization and Ca2+ sparks will be analyzed using a quantitative approach. Furthermore, a computational model of mitochondria and Ca2+ release unit will be developed to quantitatively investigate the interaction between mitochondrial energetics and local Ca2+ handling. Finally, an integrated model of the cardiomyocyte incorporating substrate metabolism, cellular electrophysiology, pH regulation and E-C coupling will be developed to investigate the mechanisms underlying alterations in energy production, ion channels, Ca2+ handling and pH, as well as the resulting reduction of cardiac contractile function during ischemia-reperfusion. By combining the experimental and computational results, these studies will allow for a complete understanding the origin of post-ischemic injury and development of heart failure, and significantly spur the development of novel heart disease therapies.