Subcellular organization and Ca+2 signaling in heart failure

描述(由申请人提供)： 钙信号转导功能障碍在许多情况下发生在心肌中，并与 心脏疾病多种多样。这个项目试图定量地表征钙离子 发生在心肌细胞中的信号缺陷，并探讨其原因 功能障碍。将对单个心室肌细胞进行共聚焦钙离子成像 测量高时间和空间分辨率的钙火花和细胞内的钙信号。 细胞内钙离子通过笼状钙离子的光释放而迅速变化，而膜 用全细胞膜片钳方法控制电压。计划中的实验将 揭示钙电流、钙火花和钙离子其他成分之间的联系 信号。 初步实验提出了一个中心假说： 兴奋-收缩偶联可能是钙离子信号缺陷的基础 观察到的。这种重塑可能涉及细胞骨架蛋白的重组，改变 L型钙通道蛋白的定位和/或兰尼定受体钙释放 横管的通道或重新组织。为了验证这一假设，转基因 将使用小鼠、大鼠疾病模型和新的细胞培养方法来检测 钙信号缺陷的分子原因。将开展具体试验，以 描述细胞骨架元素在钙信号缺陷中的作用。 因此，计划中的工作应该会拓宽我们对心脏和心脏中钙信号转导的理解 阐明它是如何在疾病中变得功能失调的。此外，这项工作应该提供 深入了解心脏细胞的亚细胞结构，并阐明 横管、细胞骨架结构、电压门控和配基门控通道。

DESCRIPTION (provided by applicant): Ca2+ signaling dysfunction occurs in heart muscle under many conditions and is associated with diverse cardiac pathologies. This project seeks to characterize quantitatively Ca2+ signaling defects that occur in cardiac ventricular myocytes and to investigate the causes of the dysfunction. Confocal Ca2+ imaging of single ventricular myocytes will be carried out to measure Ca2+ sparks and the cellular Ca2+ signal at high temporal and spatial resolution. Intracellular Ca2+ will be rapidly changed by photorelease of caged Ca2+ while membrane potential is controlled with a whole cell patch clamp method. The planned experiments will reveal the links between the Ca2+ current, the Ca2+ sparks and other components of the Ca2+ signal. Preliminary experiments suggest a central hypothesis: a disruption of the machinery of excitation-contraction coupling may underlie the Ca2+ signaling defects that have been observed. Such remodeling may involve reorganization of the cytoskeletal proteins, altered positioning of L-type Ca2+ channel proteins and/or the ryanodine receptor Ca2+ release channels or re-organization of the transverse-tubules. To investigate the hypothesis, transgenic mice, rat models of disease and new cell culture methods will be used to examine the molecular causes of the Ca2+ signaling defects. Specific experiments will be carried out to characterize the role of cytoskeletal elements in the Ca2+ signaling defects. The planned work should therefore broaden our understanding of Ca2+ signaling in heart and clarify how it may become dysfunctional in disease. Additionally, the work should provide insight into the subcellular organization of the heart cell and illuminate the links between transverse tubules, cytoskeletal structures, and voltage- and ligand-gated channels.