Dmitry Terentyev1, Colin M Rees2, Weiyan Li2, Leroy L Cooper2, Hitesh K Jindal2, Xuwen Peng2, Yichun Lu2, Radmila Terentyeva2, Katja E Odening2, Jean Daley2, Kamana Bist2, Bum-Rak Choi2, Alain Karma2, Gideon Koren1. 1. From the Department of Medicine, Division of Cardiology, Cardiovascular Research Center, Rhode Island Hospital, Alpert Medical School of Brown University, Providence (D.T., W.L., L.L.C., H.K.J., Y.L., R.T., J.D., K.B., B.-R.C., G.K.); Physics Department, Center for Interdisciplinary Research on Complex Systems, Northeastern University, Boston, MA (C.M.R., A.K.); Department of Comparative Medicine, Pennsylvania State University College of Medicine, Hershey (X.P.); and Department of Cardiology and Angiology I, Heart Center Freiburg University, Freiburg, Germany (K.E.O.). dmitry_terentyev@brown.edu gideon_koren@brown.edu. 2. From the Department of Medicine, Division of Cardiology, Cardiovascular Research Center, Rhode Island Hospital, Alpert Medical School of Brown University, Providence (D.T., W.L., L.L.C., H.K.J., Y.L., R.T., J.D., K.B., B.-R.C., G.K.); Physics Department, Center for Interdisciplinary Research on Complex Systems, Northeastern University, Boston, MA (C.M.R., A.K.); Department of Comparative Medicine, Pennsylvania State University College of Medicine, Hershey (X.P.); and Department of Cardiology and Angiology I, Heart Center Freiburg University, Freiburg, Germany (K.E.O.).
Abstract
RATIONALE: Loss-of-function mutations in human ether go-go (HERG) potassium channels underlie long QT syndrome type 2 (LQT2) and are associated with fatal ventricular tachyarrhythmia. Previously, most studies focused on plasma membrane-related pathways involved in arrhythmogenesis in long QT syndrome, whereas proarrhythmic changes in intracellular Ca(2+) handling remained unexplored. OBJECTIVE: We investigated the remodeling of Ca(2+) homeostasis in ventricular cardiomyocytes derived from transgenic rabbit model of LQT2 to determine whether these changes contribute to triggered activity in the form of early after depolarizations (EADs). METHODS AND RESULTS: Confocal Ca(2+) imaging revealed decrease in amplitude of Ca(2+) transients and sarcoplasmic reticulum Ca(2+) content in LQT2 myocytes. Experiments using sarcoplasmic reticulum-entrapped Ca(2+) indicator demonstrated enhanced ryanodine receptor (RyR)-mediated sarcoplasmic reticulum Ca(2+) leak in LQT2 cells. Western blot analyses showed increased phosphorylation of RyR in LQT2 myocytes versus controls. Coimmunoprecipitation experiments demonstrated loss of protein phosphatases type 1 and type 2 from the RyR complex. Stimulation of LQT2 cells with β-adrenergic agonist isoproterenol resulted in prolongation of the plateau of action potentials accompanied by aberrant Ca(2+) releases and EADs, which were abolished by inhibition of Ca(2+)/calmodulin-dependent protein kinase type 2. Computer simulations showed that late aberrant Ca(2+) releases caused by RyR hyperactivity promote EADs and underlie the enhanced triggered activity through increased forward mode of Na(+)/Ca(2+) exchanger type 1. CONCLUSIONS: Hyperactive, hyperphosphorylated RyRs because of reduced local phosphatase activity enhance triggered activity in LQT2 syndrome. EADs are promoted by aberrant RyR-mediated Ca(2+) releases that are present despite a reduction of sarcoplasmic reticulum content. Those releases increase forward mode Na(+)/Ca(2+) exchanger type 1, thereby slowing repolarization and enabling L-type Ca(2+) current reactivation.
RATIONALE: Loss-of-function mutations in human ether go-go (HERG) potassium channels underlie long QT syndrome type 2 (LQT2) and are associated with fatal ventricular tachyarrhythmia. Previously, most studies focused on plasma membrane-related pathways involved in arrhythmogenesis in long QT syndrome, whereas proarrhythmic changes in intracellular Ca(2+) handling remained unexplored. OBJECTIVE: We investigated the remodeling of Ca(2+) homeostasis in ventricular cardiomyocytes derived from transgenic rabbit model of LQT2 to determine whether these changes contribute to triggered activity in the form of early after depolarizations (EADs). METHODS AND RESULTS: Confocal Ca(2+) imaging revealed decrease in amplitude of Ca(2+) transients and sarcoplasmic reticulum Ca(2+) content in LQT2 myocytes. Experiments using sarcoplasmic reticulum-entrapped Ca(2+) indicator demonstrated enhanced ryanodine receptor (RyR)-mediated sarcoplasmic reticulum Ca(2+) leak in LQT2 cells. Western blot analyses showed increased phosphorylation of RyR in LQT2 myocytes versus controls. Coimmunoprecipitation experiments demonstrated loss of protein phosphatases type 1 and type 2 from the RyR complex. Stimulation of LQT2 cells with β-adrenergic agonist isoproterenol resulted in prolongation of the plateau of action potentials accompanied by aberrant Ca(2+) releases and EADs, which were abolished by inhibition of Ca(2+)/calmodulin-dependent protein kinase type 2. Computer simulations showed that late aberrant Ca(2+) releases caused by RyRhyperactivity promote EADs and underlie the enhanced triggered activity through increased forward mode of Na(+)/Ca(2+) exchanger type 1. CONCLUSIONS: Hyperactive, hyperphosphorylated RyRs because of reduced local phosphatase activity enhance triggered activity in LQT2 syndrome. EADs are promoted by aberrant RyR-mediated Ca(2+) releases that are present despite a reduction of sarcoplasmic reticulum content. Those releases increase forward mode Na(+)/Ca(2+) exchanger type 1, thereby slowing repolarization and enabling L-type Ca(2+) current reactivation.
Authors: Alexander Kushnir; Jian Shan; Matthew J Betzenhauser; Steven Reiken; Andrew R Marks Journal: Proc Natl Acad Sci U S A Date: 2010-05-17 Impact factor: 11.205
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