Madhvani Roshni V, Angelini Marina, Xie Yuanfang, Pantazis Antonios, Suriany Silvie, Borgstrom Nils P, Garfinkel Alan, Qu Zhilin, Weiss James N, Olcese Riccardo
Division of Molecular Medicine, Department of Anesthesiology, Department of Medicine (Cardiology), Department of Physiology, Department of Integrative Biology and Physiology, Cardiovascular Research Laboratory, and Brain Research Institute, David Geffen School of Medicine at University of California, Los Angeles, Los Angeles, CA 90095.
Department of Pharmacology, University of California, Davis, Davis, CA 95616.
J Gen Physiol. 2015 May;145(5):395-404. doi: 10.1085/jgp.201411288.
Early afterdepolarizations (EADs) associated with prolongation of the cardiac action potential (AP) can create heterogeneity of repolarization and premature extrasystoles, triggering focal and reentrant arrhythmias. Because the L-type Ca(2+) current (ICa,L) plays a key role in both AP prolongation and EAD formation, L-type Ca(2+) channels (LTCCs) represent a promising therapeutic target to normalize AP duration (APD) and suppress EADs and their arrhythmogenic consequences. We used the dynamic-clamp technique to systematically explore how the biophysical properties of LTCCs could be modified to normalize APD and suppress EADs without impairing excitation-contraction coupling. Isolated rabbit ventricular myocytes were first exposed to H2O2 or moderate hypokalemia to induce EADs, after which their endogenous ICa,L was replaced by a virtual ICa,L with tunable parameters, in dynamic-clamp mode. We probed the sensitivity of EADs to changes in the (a) amplitude of the noninactivating pedestal current; (b) slope of voltage-dependent activation; (c) slope of voltage-dependent inactivation; (d) time constant of voltage-dependent activation; and (e) time constant of voltage-dependent inactivation. We found that reducing the amplitude of the noninactivating pedestal component of ICa,L effectively suppressed both H2O2- and hypokalemia-induced EADs and restored APD. These results, together with our previous work, demonstrate the potential of this hybrid experimental-computational approach to guide drug discovery or gene therapy strategies by identifying and targeting selective properties of LTCC.
与心脏动作电位（AP）延长相关的早期后去极化（EADs）可导致复极化异质性和早搏，引发局灶性和折返性心律失常。由于L型钙电流（ICa,L）在AP延长和EAD形成中均起关键作用，L型钙通道（LTCCs）是使AP持续时间（APD）正常化并抑制EADs及其致心律失常后果的一个有前景的治疗靶点。我们使用动态钳技术系统地探究如何改变LTCCs的生物物理特性，以在不损害兴奋 - 收缩偶联的情况下使APD正常化并抑制EADs。首先将分离的兔心室肌细胞暴露于H2O2或中度低钾血症以诱导EADs，然后在动态钳模式下用具有可调参数的虚拟ICa,L替代其内源性ICa,L。我们探究了EADs对以下方面变化的敏感性：（a）非失活平台电流的幅度；（b）电压依赖性激活的斜率；（c）电压依赖性失活的斜率；（d）电压依赖性激活的时间常数；以及（e）电压依赖性失活的时间常数。我们发现降低ICa,L的非失活平台成分的幅度可有效抑制H2O2和低钾血症诱导的EADs并恢复APD。这些结果与我们之前的工作一起，证明了这种混合实验 - 计算方法通过识别和靶向LTCC的选择性特性来指导药物发现或基因治疗策略的潜力。
