MCU overexpression evokes disparate dose-dependent effects on mito-ROS and spontaneous Ca2 þ release in hypertrophic rat cardiomyocytes

Cardiac dysfunction in heart failure (HF) and diabetic cardiomyopathy (DCM) is associated with aberrant intracellular Ca2 þ handling and impaired mitochondrial function accompanied with reduced mitochondrial calcium concentration (mito-[Ca2 þ ]). Pharmacological or genetic facilitation of mito-Ca2 þ uptake was shown to restore Ca2 þ transient amplitude in DCM and HF, improving contractility. However, recent reports suggest that pharmacological enhancement of mito-Ca2 þ uptake can exacerbate ryanodine receptor-mediated spontaneous sarcoplasmic reticulum (SR) Ca2 þ release in ventricular myocytes (VMs) from diseased animals, increasing propensity to stress-induced ventricular tachyarrhythmia. To test whether chronic recovery of mito-[Ca2 þ ] restores systolic Ca2 þ release without adverse effects in diastole, we overexpressed mitochondrial Ca2 þ uniporter (MCU) in VMs from male rat hearts with hypertrophy induced by thoracic aortic banding (TAB). Measurement of mito-[Ca2 þ ] using genetic probe mtRCamp1h revealed that mito-[Ca2 þ ] in TAB VMs paced at 2 Hz under b-adrenergic stimulation is lower compared with shams. Adenoviral 2.5-fold MCU overexpression in TAB VMs fully restored mito-[Ca2 þ ]. However, it failed to improve cytosolic Ca2 þ handling and reduce proarrhythmic spontaneous Ca2 þ waves. Furthermore, mitochondrial-targeted genetic probes MLS-HyPer7 and OMM-HyPer revealed a significant increase in emission of reactive oxygen species (ROS) in TAB VMs with 2.5-fold MCU overexpression. Conversely, 1.5-fold MCU overexpression in TABs, that led to partial restoration of mito-[Ca2 þ ], reduced mitochondria-derived reactive oxygen species (mito-ROS) and spontaneous Ca2 þ waves. Our findings emphasize the key role of elevated mito-ROS in disease-related proarrhythmic Ca2 þ mishandling. These data establish nonlinear mito-[Ca2 þ ]/mito-ROS relationship, whereby partial restoration of mito-[Ca2 þ ] in diseased VMs is protective, whereas further enhancement of MCU-mediated Ca2 þ uptake exacerbates damaging mito-ROS emission.