Abstract 2314: Stimulation of SERCA but Not RyR2 Phosphorylation is Critical for β-Adrenergically-Mediated Arrhythmogenesis in Exercise-Induced Sudden Cardiac Death
Episodes of catecholaminergic polymorphic ventricular tachycardia (CPVT) are associated with elevated levels of catecholamines. However, the specific molecular steps involved in the stimulatory effects of catecholamines in the genesis of CPVT remain to be defined. Two possible mechanisms for the pro-arrhythmic effect of β-adrenergic stimulation include:
acute phosphorylation of cardiac ryanodine receptors (RyR2) by PKA and/or CAMK2 exacerbating the primary genetic defects in the RyR2 complex thereby resulting in spontaneous sarcoplasmic reticulum (SR) Ca release and delayed afterdepolarizations (DADs); and
stimulation of the SR Ca pump (SERCA) causes cellular arrhythmia by enhancing the rate of refilling the SR with Ca also leading to abnormal Ca releases and DADs.
To examine the contribution of these two mechanisms to CPVT, we investigated Ca handling and predisposition to cellular arrhythmia in rat ventricular myocytes ectopically expressing the CPVT-linked calsequestrin (CASQ2) mutant CASQ2.G112+5X. As shown previously, exposure to isoproterenol (ISO, 1 μM) resulted in a marked increase in incidents of spontaneous Ca release and DADs in CASQ2.G112+5X myocytes vs control. To determine whether the arrhythmogenic effects of ISO were caused SERCA activation or phosphorylation of RyR2, we employed a viral construct for expression of a dominant negative phospholamban (PLB) mutant protein (K3E/R14E, PLB-dn) that increases SR Ca content in a manner similar to ISO but without the additional effects on the RyR2. Myocytes expressing PLB-dn did not show events of spontaneous Ca release and DADs vs control cells neither in the presence or absence of ISO. Simultaneous expression of CASQ2.G112+5X and PLB-dn did not change significantly the parameters of Ca handling, including the SR Ca content, compared to myocytes expressing PLB-dn alone. However, the myocytes expressing both mutants exhibited high levels of endogenous pro-arrhythmic activity which were not significantly further affected by exposure to ISO. These results indicate that the stimulatory effect of β-adrenergic stimulation on the genesis of arrhythmia in CPVT is mediated by stimulation of SERCA-mediated SR Ca uptake and does nor require modifications of RyR2 function.