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

心脏病是美国人最大的单一杀手。越来越多的证据表明， Ca 2+处理异常与心脏病的发生密切相关。因此 理解病理条件下Ca 2+信号传导的调节是非常重要的。 Ca 2+诱导的Ca 2+释放（CICR）的局部控制依赖于L型Ca 2+的空间组织 通道和ryanodine受体（RyR）。类似地，线粒体对Ca 2+的吸收是 由于它们靠近Ca 2+释放位点，这是一个刺激氧化反应所需的过程， 在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（ROS）的主要来源，因此它们可能会影响细胞内活性氧的释放。 胞质氧化还原状态，反过来改变RyR开放概率。在这项研究中，双光子激光器 一个显微镜系统将被用来直接检查如何在能源状态的急剧变化动态 在不同的实验条件下影响Ca ~（2+）火花特性。胞质Ca 2+（或ROS），A μ m， 和NADH将在分离的豚鼠心肌细胞中同时记录，并使用 成像。线粒体之间的时空耦合！去极化和Ca 2+火花将是 用定量方法分析。此外，线粒体和Ca 2+的计算模型 释放单元将被开发用于定量研究线粒体能量学之间的相互作用 和局部Ca 2+处理。最后，一个整合模型的心肌细胞纳入基板 代谢，细胞电生理，pH调节和E-C耦合将被开发来研究 能量产生、离子通道、Ca 2+处理和pH值改变的潜在机制，以及 在缺血-再灌注期间导致心脏收缩功能的降低。通过组合 实验和计算结果，这些研究将允许一个完整的理解的起源， 缺血后损伤和心力衰竭的发展，并显着刺激新心脏的发展， 疾病治疗。

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.