Abstract 11235: Improvement of Defective Channel Function on RyR2 Mediated by CaMKII Activation Suppresses Arrhythmogenesis in TnT-related Hypertrophic Cardiomyopathy

Abstract

Backgrounds: In Familial Hypertrophic Cardiomyopathy (FHC) caused by cardiac troponin T (cTnT) mutations, Ca 2+ sensitivity in myofilaments is increased, results in leading to sudden cardiac death. The activation of calcium/calmodulin-dependent protein kinase II (CaMKII) is known to be one of the key elements for the alteration of calcium handling in heart failure, presumably mediated through defective channel function of cardiac ryanodine receptor (RyR2). However, how cTnT mutations lead to lethal arrhythmias and abnormal calcium handling through defective RyR2 remains elusive. Here, we investigated the pathogenic role of CaMKII on aberrant Ca 2+ release through RyR2 for arrhythmogenesis in FHC hearts and therapeutic effects of dantrolene, which was found to correct inter-domain interactions between N-terminal and central domain of RyR2 in FHC-related cTnT-mutated transgenic mouse (TG; cTnT-delta160E). TG have been reported myocardial disarray in histology and increased Ca 2+ sensitivity in myofilaments. Methods and results: On confocal Ca 2+ imaging by fluo4-AM exposed to isoproterenol (ISO; 10 nmol/L), the Ca 2+ spark frequency (SpF:s-1·100μm-1) was much higher in TG cardiomyocytes (ISO-TG: n=11: 7.8±0.6; p<0.05) than in non-TG (n=8: 3.4±0.4). It was largely reversed by CaMKII inhibitor {KN-93(1μM), n=7: 5.2±0.5; p<0.05}, but not by PKA inhibitor (PKI){H-89(1μM), n=7: 7.3±0.4}. The significant increase of spontaneous Ca 2+ transient (sCaT), which was measured after sequential pacing at 5 Hz was appeared in ISO-TG (42.1%; p<0.05 vs ISO-treated non-TG 14.0%), whereas it was again attenuated by CaMKII inhibitor (15.8%), but not by PKI (33.0%). These abnormal events in ISO-TG were reproduced by adding EGTA-AM into ISO-treated non-TG, suggesting that increased Ca 2+ buffering capacity, causing an increase in diastolic [Ca 2+ ], predisposes to aberrant Ca 2+ release events. Moreover, after the treatment of dantrolene (1μM), ISO-induced SpF (n=6: 5.2±0.3; p<0.05 vs ISO-TG) and sCaT (25.9%) were decreased in TG. Conclusions: CaMKII activation may play a role in the development of abnormal calcium handling in FHC-linked cTnT mutated hearts, possibly through defective inter-domain interaction between N-terminal and central domain of RyR2.