The Role of Mitophagy in Cardiomyocyte Arrhythmogenesis

Abstract

Aging is associated with more than a ten-fold increase in the incidence of sudden cardiac death. Previously our laboratory published that redox modification of ryanodine receptors (RyR2) by mitochondria-derived reactive oxygen species (ROS) contributes to aberrant calcium handling in cardiomyocytes derived from aging rabbits. However to date, there are no studies that connect increased ROS, mitochondria turnover during aging, and arrhythmia. Here we report that several proteins involved in mitophagy such as p62, LC3II/I, and PINK 1 are down-regulated in the aging rabbit heart. Additionally, we show that the mitochondrial fission protein, DRP1, is increased in the aging rabbit heart. Analysis of the ultra-structure reveals smaller mitochondrion cross-sectional area, fewer mitochondria per square micron, abnormal internal structure, and greater disorganization of the network in aged cardiomyocytes. We therefore hypothesize that decreased mitophagy is a contributing factor that promotes pro-arrhythmic, aberrant calcium signaling through RyR2 activity in aging cardiomyocytes. Blocking mitophagy with chloroquine (1µM) in HL-1 cardiomyocytes under β-adrenergic stimulation (100nM isoproterenol for 5 minutes) resulted in an increase in mitochondria-derived ROS production, upregulation of fission proteins and downregulation of markers of mitophagy. Importantly, we observed increased calcium transient amplitude, increased fractional release, and decreased latency to spontaneous calcium wave formation after 20 seconds of 2.0hz pacing. MitoTEMPO (25μM) reduced ROS production and normalized calcium homeostasis. Our data demonstrate that inhibiting mitophagy in HL-1 cells increases mitochondria derived ROS thereby the hyperactivity of RyR2 and an arrhythmogenic phenotype suggesting that decreased mitophagy contributes to pro-arrhythmic changes in the aging cardiomyocyte.