IP3R-Dependent Signaling in Excitation-Contraction Coupling during Heart Failure

心力衰竭期间兴奋收缩耦合中的 IP3R 依赖性信号传导

• 批准号: 7331064
• 负责人: Timothy Lee Domeier
• 金额: $ 2.02万
• 依托单位: LOYOLA UNIVERSITY CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7331064
• 关键词: Action Potentials; Affinity; Agonist; Arrhythmia; Binding; Buffers; Calcium; Calcium Channel; Calcium Oscillations; Calcium Signaling; Cardiac; Cardiovascular Diseases; Confocal Microscopy; Coupling; Dyes; Endothelin-1; Event; Fluo 4; Fluorescence Microscopy; Frequencies; Functional disorder; Goals; Heart; Heart Atrium; Heart Diseases; Heart failure; Individual; Investigation; Ion Channel; Knowledge; Lasers; Localized; Measures; Mediating; Membrane Potentials; Microscopy; Modeling; Monitor; Morbidity - disease rate; Muscle Cells; Oryctolagus cuniculus; Patch-Clamp Techniques; Resolution; Role; Ryanodine Receptor Calcium Release Channel; Sarcoplasmic Reticulum; Scanning; Signal Transduction; Sudden Death; Testing; Variant; Ventricular; direct application; indo 1; inositol-1,4,5-triphosphate receptor; insight; mortality; novel; receptor expression; voltage; voltage clamp

DESCRIPTION (provided by applicant): Cardiac Excitation-contraction coupling (ECC) is mediated by calcium-induced calcium release, where calcium influx through voltage gated calcium channels opens ryanodine receptors and triggers massive release of calcium from intracellular stores. Alterations in the cellular events of ECC predispose the heart to arrhythmia. IP3 receptor (IP3R) activity exerts positive inotropic and arrhythmogenic effects on ECC in atrial myocytes. During heart failure, the expression of IP3R is increased; whether this results in changes in ECC or arrhythmogenesis remains unclear. The goal of this proposal is to investigate the role of IP3R-dependent signaling on ECC (Specific Aim 1) and arrhythmogenesis (Specific Aim 2) during heart failure. Studies will utilize ventricular myocytes isolated from normal rabbits as well as from the well-characterized rabbit heart failure model. IP3R-dependent signaling will be induced using application of the I PS-liberating agonist Endothelin-1 or via direct application of IP3. Inhibition of IP3R-dependent signaling will be achieved using both pharmacological inhibition and the expression of an IP3 affinity trap which binds and buffers intracellular IP3. Fluorescence microscopy with calcium sensitive dyes will monitor intracellular calcium (epifluorescence microscopy, indo-1) and subcellular calcium release events (laser scanning confocal fluorescence microscopy, fluo-4). Further, patch clamp techniques will be used to record membrane potential and ion channel currents. Specific Aim 1 investigates the hypothesis that IP3R-dependent signaling exerts positive inotropic effects on ECC in ventricular myocytes, particularly during heart failure. In this Aim the effects of IP3R-dependent signaling on basal intracellular calcium, action potential-induced calcium transients, and elementary calcium release events (calcium sparks and puffs) will be examined. Specific Aim 2 tests the hypothesis that IP3R-dependent signaling contributes to arrhythmogenesis in heart failure ventricular myocytes. Here, the effects of IP3R-dependent signaling on the frequency of arrhythmogenic calcium signals (spontaneous calcium release, calcium waves, and calcium alternans) and changes in membrane potential (Early and Delayed Afterdepolarizations, spontaneous action potentials) will be investigated. Arrhythmia represents the major cause of sudden death during heart failure. It is expected that results from this proposal will give needed insight into the mechanisms of arrhythmia during heart failure, which may aid treatment of individuals with cardiac disease.

描述（由申请人提供）：心脏兴奋-收缩耦合（ECC）是由钙诱导的钙释放介导的，其中钙通过电压门控钙通道流入打开兰尼碱受体并触发细胞内储存的钙的大量释放。 ECC 细胞事件的改变使心脏容易出现心律失常。 IP3 受体 (IP3R) 活性对心房肌细胞 ECC 产生正性肌力和致心律失常作用。心力衰竭时，IP3R的表达增加；这是否会导致 ECC 或心律失常发生的变化仍不清楚。该提案的目标是研究心力衰竭期间 IP3R 依赖性信号传导对 ECC（具体目标 1）和心律失常发生（具体目标 2）的作用。研究将利用从正常兔子以及充分表征的兔子心力衰竭模型中分离出的心室肌细胞。 IP3R 依赖性信号传导将通过应用 I PS 释放激动剂 Endothelin-1 或直接应用 IP3 来诱导。 IP3R依赖性信号传导的抑制将通过药理学抑制和结合并缓冲细胞内IP3的IP3亲和陷阱的表达来实现。使用钙敏感染料的荧光显微镜将监测细胞内钙（落射荧光显微镜，indo-1）和亚细胞钙释放事件（激光扫描共焦荧光显微镜，fluo-4）。此外，膜片钳技术将用于记录膜电位和离子通道电流。具体目标 1 研究了以下假设：IP3R 依赖性信号传导对心室肌细胞中的 ECC 产生正性肌力作用，特别是在心力衰竭期间。在此目标中，将检查 IP3R 依赖性信号传导对基础细胞内钙、动作电位诱导的钙瞬变和基本钙释放事件（钙火花和钙喷发）的影响。具体目标 2 检验了 IP3R 依赖性信号传导导致心力衰竭心室肌细胞心律失常发生的假设。在此，将研究 IP3R 依赖性信号传导对致心律失常钙信号（自发钙释放、钙波和钙交替）频率和膜电位变化（早期和延迟后除极、自发动作电位）的影响。心律失常是心力衰竭期间猝死的主要原因。预计该提案的结果将为心力衰竭期间心律失常的机制提供必要的见解，这可能有助于心脏病患者的治疗。