Mechanisms of Long-term Cardiac Ion Channel Regulation

心脏离子通道的长期调节机制

• 批准号: 6900271
• 负责人: Jonathan Satin
• 金额: $ 36.83万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-15 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6900271
• 关键词: G protein; atrial fibrillation; biological signal transduction; calcium channel; calcium ion; cardiac myocytes; cell membrane; confocal scanning microscopy; electrophysiology; heart failure; heart function; laboratory mouse; laboratory rat; polymerase chain reaction; protein binding; protein kinase A; protein protein interaction; recombinant proteins; sarcolemma; second messengers; telemetry; voltage gated channel; western blottings

DESCRIPTION (provided by applicant): Heart failure (HF) and atrial fibrillation (AF) represent two of the most prevalent heart diseases in the U.S. In moderate to severe HF and in AF numerous phenotypic changes occur including a reduction of voltage-gated calcium channel current density. Changes in Ca channel expression may have far-reaching effects on heart function due to their central role in excitation contraction coupling. Also, Ca channels have been implicated in excitation-transcription coupling. Thus alterations of Ca channel expression in the heart can also impact on disease progression. The principal voltage-gated Ca channel in the heart (CaV1.2) is extensively studied in the realm of its biophysical characteristics and second messenger modulation. However, very sparse information is available regarding longer term regulation of Ca channel expression. In this proposal we introduce a novel mechanistic hypothesis governing the functional expression of cardiac Ca channels in the sarcolemma of cardiac myocytes. In our preliminary data we show evidence for a ras-related monomeric G-protein interaction with the CaV1.2 accessory subunit CaVb2. We propose to test the global hypothesis that G-protein interaction with CaVb subunits reduces Ca channel current density by competing for CaV1.2 binding. Moreover, our preliminary data shows strong evidence that this is a dynamic process that is modulated by chronic PKA activity. There are four specific aims that guide our experimental design: Aim 1 will characterize the time course and sub-cellular localization of nascent G-protein and CaVb complexes. Aim 2 will characterize the intracellular second messenger modulation of G-protein-CaVb regulation of Ca channel current expression. Aim 3 will characterize the modulation of Ca channel current and G-protein modulation in native cells. Aim 4 will characterize the function of these G-proteins in heart function. These aims encompass approaches ranging from investigations of a purely molecular nature (protein-protein interactions), to in vitro model systems (heterologous expression of recombinant proteins), to native cell systems (primary isolates and cultures of heart cells), to the impact of a single class of G-proteins on tissue level heart function. This proposal may identify a novel class of therapeutic targets for alleviation of heart dysfunction in HF and AF.

描述（由申请人提供）：心力衰竭 (HF) 和心房颤动 (AF) 是美国最常见的两种心脏病。中度至重度 HF 和 AF 会发生多种表型变化，包括电压门控钙通道减少电流密度。 Ca 通道表达的变化可能对心脏功能产生深远的影响，因为它们在兴奋收缩耦合中发挥着核心作用。此外，Ca 通道与兴奋转录耦合有关。因此，心脏中 Ca 通道表达的改变也会影响疾病进展。心脏中主要的电压门控 Ca 通道 (CaV1.2) 在其生物物理特性和第二信使调制领域得到了广泛研究。然而，关于 Ca 通道表达的长期调节的信息非常稀疏。在本提案中，我们引入了一种新的机制假说，该假说控制心肌细胞肌膜中心脏 Ca 通道的功能表达。在我们的初步数据中，我们显示了 ras 相关单体 G 蛋白与 CaV1.2 辅助亚基 CaVb2 相互作用的证据。我们建议测试以下总体假设：G 蛋白与 CaVb 亚基的相互作用通过竞争 CaV1.2 结合来降低 Ca 通道电流密度。此外，我们的初步数据显示强有力的证据表明这是一个受慢性 PKA 活动调节的动态过程。有四个具体目标指导我们的实验设计：目标 1 将表征新生 G 蛋白和 CaVb 复合物的时间过程和亚细胞定位。目标 2 将表征细胞内第二信使 G 蛋白-CaVb 对 Ca 通道电流表达的调节。目标 3 将表征天然细胞中 Ca 通道电流的调节和 G 蛋白的调节。目标 4 将描述这些 G 蛋白在心脏功能中的功能。这些目标涵盖的方法范围从纯分子性质（蛋白质-蛋白质相互作用）的研究，到体外模型系统（重组蛋白质的异源表达），到天然细胞系统（心脏细胞的原代分离物和培养物），再到一类 G 蛋白对组织水平心脏功能的影响。该提议可能会确定一类新的治疗靶点，用于缓解 HF 和 AF 的心功能障碍。