CONICET | Buscador de Institutos y Recursos Humanos

Cardiac dysfunction is associated with an abnormal intracellular Ca2+ cycling, particularly during the setting of ischemia/reperfusion (I/R). Hearts submitted to ischemia suffer an increase in diastolic Ca2+, accompanied with an increase in sarcoplasmic reticulum (SR) Ca2+ load. At the onset of reperfusion there is a massive increase in cytosolic Ca2+, associated with an abrupt release of Ca2+ from the SR (Cardiovasc Res, 2006,2010). In a previous work we showed that arrhythmogenic Ca2+ waves are the subcellular events associated with this SR-Ca2+ release (Circulation, 2013). Moreover, Ca2+/calmodulin-dependent protein kinase II (CaMKII) has been associated with reperfusion arrhythmias, particularly associated with a CaMKII-dependent phosphorylation of Ser2814 site of ryanodine receptors (RyR2) (JMCC,2011). The aim of the present work was to evaluate the role of CaMKII on the abrupt SR-Ca2+ release and enhanced frequency of Ca2+ waves during reperfusion. The experiments were done in perfused mouse (C57BL/6) hearts loaded with the Ca2+ indicator Fluo-4, on an upright confocal microscope. Hearts were submitted to ischemia and reperfusion (15/30 min) at 32ºC, in the presence and absence of a CaMKII inhibitor (KN-93, 2.5 μM). We evaluated Ca2+ waves at the epicardial layer of the heart. Inhibition of CaMKII did not prevent the increase in Ca2+ wave frequency (0.15±0.05 pre-ischemia vs. 0.29±0.04 waves/100μm/s, ischemia) nor the velocity of the Ca2+ wave propagation (122.01±4.62 vs. 126.06±1.02 μm/s). During reperfusion in the presence of KN-93, we observed a reduction in Ca2+ waves frequency (0.28±0.06 vs. 0.14±0.02 waves/μm/s, control vs. KN-93), and also in the velocity of its propagation (114.99±3.02 vs. 94.75±1.45 μm/s, control vs. KN-93). The results indicate that the increase in the proarrhythmogenic Ca2+ waves at the onset of reperfusion are mainly mediated by a CaMKII-dependent phosphorylation.