CONICET | Buscador de Institutos y Recursos Humanos

To explore whether CaMKII-dependent phosphorylation events mediate reperfusion arrhythmias,Langendorff perfused hearts were submitted to global ischemia/reperfusion. Epicardialmonophasic or transmembrane action potentials and contractility were recorded. In rat hearts,reperfusion significantly increased the number of premature beats (PBs) relative to pre-ischemicvalues. This arrhythmic pattern was associated with a significant increase in CaMKII-dependentphosphorylation of Ser2814 on Ca2+-release channels (RyR2) and Thr17 on phospholamban(PLN) at the sarcoplasmic reticulum (SR). These phenomena could be prevented by the CaMKIIinhibitor KN-93. In transgenic mice with targeted inhibition of CaMKII at the SR membranes (SR-AIP), PBs were significantly decreased from 31 ± 6 to 5 ± 1 beats/3 min with a virtually complete disappearance of early-afterdepolarizations (EADs). In mice with genetic mutation of the CaMKII phosphorylation site on RyR2 (RyR2-S2814A), PBs decreased by 51.0 ± 14.7 %. In contrast, the number of PBs upon reperfusion did not change in transgenic mice with ablation of both PLN phosphorylation sites (PLN-DM). The experiments in SR-AIP mice, in which the CaMKII inhibitor peptide is anchored in the SR membrane but also inhibits CaMKII regulation of L-type Ca2+ channels, indicated a critical role of CaMKII-dependent phosphorylation of SR proteins and/or L-type Ca2+ channels in reperfusion arrhythmias. The experiments in RyR2-S2814A further indicate that up to 60% of PBs related to CaMKII are dependent on the phosphorylation of RyR2 Ser2814 site and could be ascribed to delayed-afterdepolarizations (DADs). Moreover, phosphorylation of PLN-Thr17 and L-type Ca2+ channels might contribute to reperfusion-induced PBs, by increasing SR Ca2+ content and Ca2+ influx.