Coupled Gating of L-type Calcium Channels in Heart

心脏 L 型钙通道的耦合门控

基本信息

批准号：
8645692
负责人：
Luis F Santana
金额：
$ 38.22万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-04-20 至 2017-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8645692
关键词：
1,2-diacylglycerol A kinase anchoring protein Action Potentials Address Adrenergic Agents Amino Acid Substitution Arrhythmia Autistic Disorder Calcineurin Calmodulin Cells Cellular Membrane Coupled Coupling Cyclic AMP Cyclic AMP-Dependent Protein Kinases Data Development Diglycerides Disease Electrophysiology (science)Event Frequencies Funding Gene Expression Genetic Transcription Goals Health Heart Heart failure Human Image Individual Knock-in Mouse Knockout Mice L-Type Calcium Channels Label Lead Light Location Long QT Syndrome Membrane Methods Modality Modeling Modification Molecular Biology Mus Muscle Cells Optics Phosphorylation Site Phosphotransferases Physiological Play Population Probability Public Health Research Resolution Role Sarcolemma Scaffolding Protein Shapes Signal Transduction Surface System Techniques Testing Timothy syndrome Total Internal Reflection Fluorescent Transgenic Mice Transgenic Organisms Translating Ventricular Wild Type Mouse Work adrenergic calcineurin phosphatase design heart function innovation interest mutant novel optogenetics patch clamp red fluorescent protein research study response transcription factor transcription factor NF-AT c3 treatment strategy

项目摘要

DESCRIPTION (provided by applicant): The work proposed in this application will test the hypothesis that L-type Cav1.2 channel activity varies along the sarcolemma of ventricular myocytes due to signaling micro-domains created by a subpopulation of these channels interacting with the scaffolding protein AKAP150. Preliminary data suggest the novel concept that AKAP150-associated Cav1.2 channels play a critical role in shaping action potential waveform, arrhythmogenesis, excitation-contraction (EC) coupling, and excitation-transcription (ET) coupling in ventricular myocytes. A key discovery is that small clusters of AKAP150-associated Cav1.2 channels are capable of undergoing coordinated openings and closings ("coupled gating"). The frequency of coupled gating events increases in cells expressing a mutant Cav1.2 channel that causes arrhythmias and autism in humans with long QT syndrome 8 (LQT8). The project has three specific aims designed to investigate the physiological and pathophysiological implications of these findings. Specific aim 1 is to test the hypothesis that AKAP150 is required for coupled gating of wild type (WT) and Cav1.2-LQT8 channels. Specific aim 2 is to test the hypothesis that coupled gating of WT and Cav1.2-LQT8 channels modulates EC coupling in ventricular myocytes. Finally, Specific aim 3 is to test the hypothesis that loss of AKAP150 protects against arrhythmias in WT and LQT8 mice. The methods that will be used to achieve these aims include patch-clamp electrophysiology, optical clamping, light- and chemically-induced dynamic targeting of kinases to cellular membranes, light-induced activation of adrenergic signaling (i.e., optogenetics), confocal, and TIRF microscopy. Experiments will involve a transgenic mouse specially created for this project and that expresses fluorescently labeled Cav1.2-LQT8 channels in ventricular myocytes as well as other transgenic, knock in, and knock out mice created by collaborators. This work will generate fundamental information on the mechanisms by which AKAP150 and Cav1.2 channels control of excitability, gene expression, and EC coupling in the heart under physiological and pathological conditions.

描述（由申请人提供）：本申请中提出的工作将检验以下假设：由于信号传导微型域，由于这些通道的子群与这些通道与支架蛋白AKAP150相互作用而产生的这些通道产生的信号微型域，l型CAV1.2通道活性沿着心室心肌细胞的肌瘤变化。初步数据表明，与AKAP150相关的CAV1.2通道在塑造动作电势波形，心律失常发生，激发 - 收缩（EC）耦合以及兴奋 - 转录（ET）耦合在心室心肌细胞中起着关键作用。一个关键的发现是，与AKAP150相关的CAV1.2通道的小簇能够进行协调的开口和关闭（“耦合门控”）。在表达突变体CAV1.2通道的细胞中，耦合门控事件的频率会增加，该通道在患有长QT综合征8（LQT8）的人类中引起心律不齐和自闭症。该项目具有三个特定的目的，旨在研究这些发现的生理和病理生理意义。具体目的1是测试野生型（WT）和CAV1.2-LQT8通道所必需的AKAP150的假设。具体目的2是测试以下假设：WT和CAV1.2-LQT8通道的耦合耦合可以调节心室心肌细胞中的EC耦合。最后，具体目的3是测试AKAP150损失可预防WT和LQT8小鼠中心律失常的假设。将用于实现这些目标的方法包括斑板夹电生理学，光学夹紧，光和化学诱导的激酶对细胞膜的动态靶向，光诱导的肾上腺素信号传导（即光遗传学）的光激活，concocalics，Cumcocal和Tirf Microscopy。实验将涉及专门为该项目创建的转基因小鼠，并表达荧光标记为CAV1.2-LQT8通道中的心室肌细胞以及其他转基因，并敲击并敲除了由协作者创建的小鼠。这项工作将生成有关AKAP150和CAV1.2通道控制兴奋性，基因表达和EC在生理和病理条件下心脏中对兴奋性，基因表达和EC偶联的机制的基本信息。