RyR1 Mutation Associated with Malignant Hyperthermia Induces Cardiac Arrythmia via Mitochondrial Calcium Overload

Abstract

Introduction The “leaky” gain-of-function mutations of the ryanodine receptor type 1 (RyR1) are linked to skeletal muscle disorders, including malignant hyperthermia (MH), which occurs in response to particular medications used during general anesthesia. In addition to a severe reaction to particular anesthetic drugs, sudden cardiac death (SCD) in MH patients has been also reported to occur in non-anesthesia conditions. However, the molecular mechanism of SCD associated with MH remains unresolved. Although the major RyR isoform expressed in the heart is RyR2, we previously reported a low level of RyR1 expression in cardiac mitochondria but not in the sarcoplasmic reticulum (SR). Hypothesis Chronic mitochondrial Ca2+ overload through the ‘leaky’ RyR1 expressed in the cardiac mitochondria induces mitochondrial injury and oxidative stress leading to cardiac arrhythmia. Methods We utilized a knock-in mouse model carrying a human RyR1 MH mutation Y522S (YS) and assessed in vivo and ex vivo cardiac function. Cardiac mitochondria and ventricular myocytes were isolated from heterozygous YS and wild type (WT) mouse hearts for in situ assays. Results Both WT and heterozygous YS hearts possessed RyR1 expression in the mitochondria but not in SR, but only the heterozygous YS hearts showed disrupted mitochondrial morphology, damaged myofibrils, and high cellular oxidative state. In addition, isolated YS mitochondria showed higher basal mitochondrial Ca2+ concentration, and depolarized mitochondrial membrane potential. However, isolated YS ventricular myocytes possessed higher levels of mitochondrial reactive oxygen species and slower cytosolic Ca2+ clearance compared to controls. However, pretreating YS mitochondria/ ventricular myocytes with a clinically approved anti-MH drug and a RyR blocker, dantrolene, prevented these changes and normalized their Ca2+ handling profiles. Although the in vivo basal cardiac function was comparable to the WT, the YS heart was insensitive to isoproterenol-induced positive inotropy and chronotropy due to a higher level of catecholamine signaling at baseline. However, ex vivo YS (but not WT) hearts frequently developed multiple ventricular extrasystoles in response to isoproterenol bolus. Optical mapping of YS hearts showed increased early after-depolarization (EAD) and premature ventricular contractions (PVC) in response to isoproterenol, which was effectively abolished by dantrolene treatment. Conclusion: Chronic mitochondrial Ca2+ overload via ‘leaky’ RyR1 disrupts cardiac mitochondrial functions and structures and increases the vulnerability to ventricular arrhythmia by catecholamine.