Ryanodine Receptor Channels in Heart Failure

描述(由申请人提供)：细胞内钙的异常处理已被确定为心力衰竭(HF)的主要原因，心力衰竭是美国的主要死亡原因。尽管进行了大量的努力，但钙信号与HF相关的确切机制仍然知之甚少。本研究的总体目标是明确肌浆网(SR)钙释放通道/Ryanodine受体(RyR)在心衰中的作用。在心肌中，RyRs是通过钙诱导的钙释放(CICR)机制被胞内钙激活的。我们实验室最近获得的数据表明，除了胞质内的钙外，RyRs还受管腔部位SR内的钙的控制。腔内钙传感器的存在调节SR钙的释放，对于我们理解正常和病变心肌的细胞内钙信号具有深远的意义。例如，通过将RyR通道的功能活动与SR的负载状态联系起来，管腔钙传感器在RyR改变时稳定钙释放。因此，任何影响RyR特性的因素都可以通过SR钙负荷的变化以及伴随而来的RyR通道活动的腔内钙依赖变化来补偿。我们的初步数据表明，这种反馈机制可能在心力衰竭时受到损害，可能导致SR钙释放功能的慢性抑制。在目前的提案中，我们将解决以下问题：1)RyRs的固有门控特性(即受胞浆和腔内钙的调节)在HF中是否发生了变化？2)这些变化的确切分子机制是什么？3)RyRs的变化是如何导致心衰患者钙循环异常的？为此，将利用重建系统中的单通道方法以及分离的心室肌细胞和整个心脏跳动中的荧光钙成像技术，使用几种成熟的心力衰竭动物模型，在多个水平上确定RyR特性和心力衰竭之间的关系。本研究的结果将加深我们对心衰时钙处理改变的机制和作用的理解。

DESCRIPTION (provided by applicant): Abnormal handling of intracellular Ca has been identified as a major cause of heart failure (HF), a leading cause of death in the US. Despite intense effort, the precise mechanisms underlying the HF-related alterations of Ca signaling remain poorly understood. The overall goal of this proposal is to define the role of alterations of the sarcoplasmic reticulum (SR) Ca release channel/ryanodine receptor (RyR) in HF. In cardiac muscle, RyRs are activated by cytosolic Ca via the mechanism of Ca-induced Ca release (CICR). Data obtained recently in our laboratory shows that in addition to cytosolic Ca, RyRs are also controlled by Ca inside the SR at luminal sites. The presence of a luminal Ca sensor regulating release of SR Ca has profound implications for our understanding of intracellular Ca signaling in normal and diseased myocardium. For example, by linking the functional activity of RyR channels to the loading state of SR, the luminal Ca sensor stabilizes Ca release in the face of alterations of RyRs. Consequently, any factors affecting the properties of RyRs are compensated by changes in SR Ca load and concomitant luminal Ca-dependent changes in RyR channel activity. Our preliminary data suggest that this feedback mechanism might be compromised in HF, potentially leading to chronic depression of SR Ca release function. In the present proposal we will address the following questions: 1) Are the intrinsic gating properties of RyRs (i.e. regulation by cytosolic and luminal Ca) altered in HF? 2) What are the precise molecular mechanisms of these alterations? And 3) How do the proposed changes in RyRs contribute to abnormal Ca cycling in HF? To this end, single channel approaches in a reconstitution system and fluorescent calcium imaging techniques in isolated ventricular myocytes and whole beating hearts will be utilized to determine relationships between RyR properties and HF at multiple levels, using several well-established animal models of HF. The results of this study will improve our understanding of the mechanisms and role of altered Ca handling in HF.