Non-selective Large Conductance Ion Channels In Triggered Arrhythmia - ABSTRACT Sudden cardiac death (SCD), a major public health problem in the US and worldwide, is often caused by a triggered arrhythmia, such as ventricular tachycardia (VT) evolving into ventricular fibrillation. VT is often preceded and maintained by spontaneous extra beats that do not originate from the sinus node. Those are thought to occur at the cellular level: Under pathologic conditions of excessive Ca load in the intracellular Ca stores (sarcoplasmic reticulum, SR) and/or malfunctioning ryanodine receptors (RyR2), Ca is released from the SR during diastole. This induces a transient inward current (Iti), consisting mainly of Na/Ca exchanger (NCX) current moving Ca out of the cell. Iti depolarizes the cell membrane, generating a delayed afterdepolarization (DAD). If the DAD is large enough to reach the Na channel activation threshold, an ectopic action potential (AP) occurs, which then can propagate and cause a triggered arrhythmia on the tissue level. Such diastolic Ca releases and associated triggered beats are hallmarks of inherited catecholaminergic polymorphic ventricular tachycardia (CPVT) and are also found as an acquired defect after myocardial infarction (MI). Yet, it remains unclear how NCX can generate enough current to trigger an AP in intact tissue, where ventricular myocytes are electrically coupled and thus effectively stabilized by a large current sink. Non-selective large-conductance (200-500 pS) ion channels (LCCs), activated by several mechanisms, including increase in intracellular Ca ([Ca]i) and/or extracellular ATP ([ATP]o), have been previously reported in the plasma membrane of ventricular and atrial cardiomyocytes. The molecular identity of these LCCs remains controversial, with number of candidates being proposed, including RyR, transient receptor potential (TRP), pannexin (Panx) channels, and connexin (Cx43) hemichannels. Our preliminary data from Casq2-/- mice (calsequestrin knock-out model of CPVT) strongly points toward Panx1 channels as the major candidate, although Cx43 hemichannels also may be contributing to LCC activity. Our central hypothesis to be tested in current application is that rapid simultaneous activation of Panx1 in certain pathophysiological conditions may serve as a significant amplifying and/or synchronizing factor for the Iti elicited by spontaneous diastolic Ca release, making AP triggering possible and in this way critically contributing to arrhythmia and SCD. The proposed experiments will investigate if activation of Panx1 by [ATP]o via paracrine signaling pathway increases the incidence of triggered AP in single cardiomyocytes from Casq2-/- and WT mice and in vivo (Aim 1), test if increased Panx1 expression in cardiac Purkinje cells contributes to their arrhythmogenicity (Aim 2) and study the effect of Panx modulation on arrhythmia risk using mouse model of acquired (MI, coronary ligation) triggered arrhythmia pathology. Accomplishing these aims may reveal a new mechanism of arrhythmia triggering in cardiac pathologies associated with diastolic Ca release and provide a new target for their therapeutic treatment.
