Proarrhythmic Defects in Timothy Syndrome Require Calmodulin Kinase II

Background-Timothy syndrome (TS) is a disease of excessive cellular Ca2+ entry and life-threatening arrhythmias caused by a mutation in the primary cardiac L-type Ca2+ channel (Ca(V)1.2). The TS mutation causes loss of normal voltage-dependent inactivation of Ca(V)1.2 current (I-Ca). During cellular Ca2+ overload, the calmodulin-dependent protein kinase II (CaMKII) causes arrhythmias. We hypothesized that CaMKII is a part of the proarrhythmic mechanism in TS. Methods and Results-We developed an adult rat ventricular myocyte model of TS (G406R) by lentivirus-mediated transfer of wild-type and TS Ca(V)1.2. The exogenous Ca(V)1.2 contained a mutation (T1066Y) conferring dihydropyridine resistance, so we could silence endogenous Ca(V)1.2 with nifedipine and maintain peak I-Ca at control levels in infected cells. TS Ca(V)1.2-infected ventricular myocytes exhibited the signature voltage-dependent inactivation loss under Ca2+ buffering conditions, not permissive for CaMKII activation. In physiological Ca2+ solutions, TS Ca(V)1.2-expressing ventricular myocytes exhibited increased CaMKII activity and a proarrhythmic phenotype that included action potential prolongation, increased I-Ca facilitation, and afterdepolarizations. Intracellular dialysis of a CaMKII inhibitory peptide, but not a control peptide, reversed increases in I-Ca facilitation, normalized the action potential, and prevented afterdepolarizations. We developed a revised mathematical model that accounts for CaMKII-dependent and CaMKII-independent effects of the TS mutation. Conclusion-In TS, the loss of voltage-dependent inactivation is an upstream initiating event for arrhythmia phenotypes that are ultimately dependent on CaMKII activation. (Circulation. 2008; 118: 2225-2234.)