权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Molecular mechanisms of regulation of L-type Ca channels

L型Ca通道调节的分子机制

基本信息

批准号：
17590189
负责人：
KAMEYAMA Masaki
金额：
$ 2.18万
依托单位：
Kagoshima University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17590189/
关键词：
calcium channels calmodulin calmodulin kinase II patch clamp cardiac myocytes GST融合蛋白

项目摘要

Activity of the cardiac L-type Ca^<2+> channel (Ca_v1.2) is modulated by dual feedback mechanisms, i.e., Ca^<2+>-dependent facilitation (CDF) and Ca^<2+>-dependent inactivation (CDI). Although calmodulin (CaM) has been suggested to mediate both CDF and CDI, Ca^<2+>/CaM-dependent protein kinase II (CaMKII) is also suggested to play a primary role in CDF. In this study, we investigated the roles and relations of Ca^<2+>, CaM and CaMKII in the basal activity, CDF and CDI using the patch-clamp method in guinea-pig ventricular myocytes. In the cell-attached mode, inhibitors of CaM significantly reduced both CDF and CDI, whereas those of CaMKII only modulated the time courses of CDF and CDI. In the inside-out mode, CaM (0.1-3μM) + ATP (2.4-3 mM)(at [Ca^<2+>]_i <10 nM) produced dose-dependent channel activity with a bell-shaped relationship between [CaM] and channel activity and with a maximum activation of 200-300% of control. Increasing of [Ca^<2+>]_i (~500 nM) shifted the [CaM]-channel activity curve toward left (i.e., to lower [CaM]). Thus, at a fixed concentration of CaM (e.g. 0.5 μM), Ca^<2+> showed biphasic effects: facilitation at [Ca^<2+>]_i <500 nM, and inactivation at [Ca^<2+>]_i >500 nM. This Ca^<2+>-dependent effect of CaM was considered to represent CDF and CDI. Based on these data, a simple model for Ca^<2+>-and CaM-dependent modulation of the Ca_v1.2 channel has been proposed, in which the channel had two CaM-binding sites, one for the basal activity and CDF and the other for CDI. In conclusion, it is suggested that CaM plays key roles in maintaining the basal activity and in producing CDF and CDI of the channel, and that CaMKII and Ca^<2+> modulate the interaction between the channel and CaM.

心脏L型Ca^2+通道（Ca_v1.2）的活性受双重反馈机制调节，即，Ca^2+依赖性易化（CDF）和Ca^2+依赖性失活（CDI）。虽然钙调素（CaM）被认为介导CDF和CDI，但Ca 2 +/CaM依赖性蛋白激酶II（CaMKII）也被认为在CDF中起主要作用。本研究应用膜片钳技术研究了Ca^<2+>、CaM和CaMK Ⅱ在豚鼠心室肌细胞基础活性、CDF和CDI中的作用及其相互关系。在细胞附着模式下，CaM的抑制剂显着降低CDF和CDI，而CaMKII的抑制剂只调制CDF和CDI的时间过程。在由内向外的模式中，CaM（0.1-3μM）+ ATP（2.4-3 mM）（[Ca 2+] i <10 nM）产生剂量依赖性的通道活性，[CaM]与通道活性之间呈钟形关系，最大激活为对照的200-300%。增加[Ca^<2+>]_i（~500 nM）使[CaM]-通道活性曲线向左移动（即，降低[CaM]）。因此，在固定浓度的钙调素（如0.5 μM）下，Ca^<2+>表现出双相效应：[Ca^<2+>]_i <500 nM时易化，[Ca^<2+>]_i >500 nM时失活。CaM的这种Ca^<2+>依赖性作用被认为代表了CDF和CDI。基于这些数据，我们提出了一个简单的Ca_v1.2通道的Ca^<2+>和CaM依赖性调节模型，其中该通道有两个CaM结合位点，一个用于基础活性和CDF，另一个用于CDI。结果表明，CaM在维持通道的基础活性和产生CDF和CDI中起关键作用，CaMK Ⅱ和Ca^<2+>调节通道与CaM的相互作用。