Calcium Leak Through Ryanodine Receptors Leads to Atrial Fibrillation in 3 Mouse Models of Catecholaminergic Polymorphic Ventricular Tachycardia

Abstract

Rationale Atrial fibrillation (AF) is the most common cardiac arrhythmia, however the mechanism(s) causing AF remain poorly understood and therapy is suboptimal. The ryanodine receptor (RyR2) is the major calcium (Ca2+) release channel on the sarcoplasmic reticulum (SR) required for excitation-contraction coupling in cardiac muscle. Objective In the present study we sought to determine whether intracellular diastolic SR Ca2+ leak via RyR2 plays a role in triggering AF and whether inhibiting this leak can prevent AF. Methods and Results We generated three knock-in mice with mutations introduced into RyR2 that result in leaky channels and cause exercise induced polymorphic ventricular tachycardia in humans [catecholaminergic polymorphic ventricular tachycardia (CPVT)]. We examined AF susceptibility in these three CPVT mouse models harboring RyR2 mutations to explore the role of diastolic SR Ca2+ leak in AF. AF was stimulated with an intra-esophageal burst pacing protocol in the three CPVT mouse models (RyR2-R2474S+/−, 70%; RyR2-N2386I+/−, 60%; RyR2-L433P+/−, 35.71%), but not in wild type (WT) mice (P<0.05). Consistent with these in vivo results, there was a significant diastolic SR Ca2+ leak in atrial myocytes isolated from the CPVT mouse models. Calstabin2 (FKBP12.6) is an RyR2 subunit that stabilizes the closed state of RyR2 and prevents a Ca2+ leak through the channel. Atrial RyR2 from RyR2-R2474S+/− mice were oxidized and the RyR2 macromolecular complex was depleted of calstabin2. The Rycal drug S107 stabilizes the closed state of RyR2 by inhibiting the oxidation/phosphorylation induced dissociation of calstabin2 from the channel. S107 reduced the diastolic SR Ca2+ leak in atrial myocytes and decreased burst pacing-induced AF in vivo. S107 did not reduce the increased prevalence of burst pacing-induced AF in calstabin2-deficient mice, confirming that calstabin2 is required for the mechanism of action of the drug. Conclusions The present study demonstrates that RyR2-mediated diastolic SR Ca2+ leak in atrial myocytes is associated with AF in CPVT mice. Moreover, the Rycal S107 inhibited diastolic SR Ca2+ leak through RyR2 and pacing-induced AF associated with CPVT mutations.

The Clinical Profile and Pathophysiology of Atrial Fibrillation

“…167 Catecholaminergic polymorphic ventricular tachycardia mutations alter cellular Ca 2+ handling and induce DADs; although they are typically associated with ventricular arrhythmias, they can also cause AF, 217 likely by inducing DAD-related rapid atrial ectopic firing. 218 Common gene variants, generally manifesting as single nucleotide polymorphisms, contribute to AF susceptibility in a quantitative fashion, with much lower penetrance than monogenic causes. It is likely that they predispose individuals to develop AF in association with acquired risk factors.…”

Could Plugging the Oxidation-Mediated Ca 2+ Leak Stem the Tide of the Atrial Fibrillation Epidemic?

“…Although several competing mechanisms have been proposed to account for the ␤-adrenergic-mediated ventricular tachycardia observed in CPVT, none incorporates RYR2 oxidation as a critical component, consistent with the absence of elevated RYR2 oxidation in the ventricular myocardium of the CPVT mice. 23 These results suggest that the mechanism for arrhythmia induction by CPVT-causing RYR2 mutations in atria could be distinct from the ventricle and could help explain why AF is not commonly observed in CPVT patients. 13 However, the conclusion that ␤-adrenergic activation and RYR2 phosphorylation do not play a role is at odds with the fact that 3 of the 5 case reports linking AF to CPVT found that AF was induced by exercise or emotional stress.…”

“…4 15 Moreover, several mouse studies demonstrated that AF can be prevented by inhibiting CaMKII-dependent phosphorylation of RYR2 at position S2814 in mice either expressing CPVT-causing RYR2 mutations or lacking FKBP12.6, 16 -19 as well as by treatment with drugs that stabilize calstabin2 binding to the RYR2 complex. 19 -22 In this issue of Circulation Research, Shan et al 23 report on 3 CPVT-causing RYR2 mutations (R2474S, N2386I, and L443P) that enhance SR Ca 2ϩ leak from atrial myocytes and increase susceptibility to AF induction by rapid atrial pacing. Moreover, despite the activation of CaMKII and phosphorylation of RYR2 at position S2814, the enhanced sensitivity of these CPVT mice to pacing-induced AF required RYR2 oxidation, leading to calstabin2 dissociation from RYR2 and enhanced spontaneous SR Ca 2ϩ release.…”

“…Accordingly, reducing agents eliminated differences in RYR2 oxidation and restored the calstabin2 association with RYR2 in atrial myocardium between the groups. The observation by Shan et al 23 that oxidation of RYR2 channels is central to AF induction in CPVT is intriguing, because dietary and pharmacological antioxidants have often been advocated for treatment of AF, with mixed results. 6,25,26 More important, the results of Shan et al 23 suggest that prevention of the dissociation…”

“…Shan et al 23 also concluded that neither protein kinase A nor CaMKII activation plays a notable role in the enhanced sensitivity of CPVT mice to pacing-induced AF susceptibility. Their conclusion is supported by the absence of differences between CVPT mice and wild-type mice in phosphorylation of RYR2 channels at either position S2808 (the protein kinase A/CaMKII site) under baseline conditions or position S2814 (the CaMKII site) after rapid pacing.…”