Pannexin channels in cardiac arrhythmias

 描述（由申请人提供）：泛联蛋白（Px）通道，在2000年首次被描述，主要在脑中研究，据报道它们在突触功能、Ca波的传播和血管张力的调节中起作用。关于它们在心脏中的作用知之甚少。一般而言，已经描述了三种同种型（Px 1，2，3）。Px通道来自大孔，可渗透分子高达1 kDa，并可在生理细胞外Ca。在心室肌细胞中，Px通道以非常低的密度表达，但单通道电导如此之大（>300 pS），以至于可以在全细胞膜片钳中解析。该提案的目的是测试舒张期钙释放过程中Px通道激活促进触发心跳的新假设。肌浆网（SR）钙库舒张期钙释放和触发搏动是遗传性儿茶酚胺能多态性室性心动过速（CPVT）的标志，也被发现为心肌梗死（MI）后的获得性缺陷。舒张期Ca释放引起使细胞膜去极化的瞬时内向电流（Iti）（称为延迟后去极化，DAD），当其大到足以激活Na通道时，触发动作电位（AP）和传播性搏动。通常认为Iti主要是由Na Ca交换器（NCX）将Ca运输出细胞产生的。然而，目前尚不清楚NCX如何产生足够的电流来触发完整组织中的AP，其中心室肌细胞通过大电流“汇”电耦合并有效稳定。我们的初步数据表明，在舒张期Ca释放期间Px通道开放有助于来自CPVT小鼠模型（钙螯合蛋白KO小鼠（Casq 2-/-））的心肌细胞中的Iti。此外，Px通道的药理学抑制或Px 1的遗传KO有效地减少了Casq 2-/-小鼠中的触发性心律失常。因此，我们的中心假设是，Px通道重要地增加由SR Ca过早释放引起的DAD，并提高心律失常的易感性。提出的实验将研究Px通道激活和对分离的心肌细胞中Iti的贡献（Aim 1），并使用Casq 2-/-小鼠（Aim 2）和冠状动脉结扎小鼠（Aim 3）测试体内心律失常易感性。实现这些目标将提供心律失常中Px通道的新机制信息，并可能导致治疗常见心脏病（如MI）中钙触发心律失常的新范式。

 DESCRIPTION (provided by applicant): Pannexin (Px) channels, first been described in 2000, have been mostly studied in the brain where they reportedly play a role in synaptic function, propagation of Ca waves and regulation of vascular tone. Much less is known about their role in the heart. In general, three isoforms have been described (Px1, 2, 3). Px channels from large pores permeable to molecules up to 1 kDa and can open at physiological extracellular Ca. In ventricular cardiomyocytes Px channels are expressed in very low density, but single channel conductance is so large (>300 pS) that it can be resolved in whole cell patch clamp. The objective of the proposal is to test the novel hypothesis that Px channel activation during diastolic Ca release facilitates triggered heart beats. Diastolic Ca releases from the sarcoplasmic reticulum (SR) Ca store and triggered beats are hallmarks of inherited catecholaminergic polymorphic ventricular tachycardia (CPVT) and are also found as acquired defect after myocardial infarction (MI). Diastolic Ca releases cause a transient inward current (Iti) that depolarizes the cell membrane (termed delayed after depolarization's, DADs), which, when large enough to activate the Na channel, trigger an action potential (AP) and a propagated beat. It is generally accepted that Iti is foremost generated by the Na Ca exchanger (NCX) transporting Ca out of the cell. However, it remains unclear how NCX can generate enough current to trigger an AP in intact tissue, where ventricular myocytes are electrically coupled and effectively stabilized by a large current "sink". Our preliminary data indicate that Px channel opening during diastolic Ca release contributes to Iti in cardiomyocytes from a CPVT mouse model, the calsequestrin KO mice (Casq2-/-). Moreover, pharmacologic inhibition of Px channels or genetic KO of Px1 effectively reduced triggered arrhythmia in Casq2-/- mice. Thus, our central hypothesis is that Px channels importantly increase DADs caused by premature SR Ca release and elevate arrhythmia susceptibility. The proposed experiments will investigate Px channel activation and contribution to Iti in isolated cardiomyocytes (Aim 1) and test arrhythmia susceptibility in vivo using Casq2-/- mice (Aim 2) and mice with coronary artery ligation (Aim 3). Accomplishing these aims will provide new mechanistic information on Px channels in arrhythmia and may lead to a new paradigm for treating Ca triggered arrhythmia in common heart diseases such as MI.