Abstract
Cardiomyopathy is associated with cardiac Na+ channel downregulation that may contribute to arrhythmias. Previously, we have shown that elevated intracellular NADH causes a decrease in cardiac Na+ current (INa) signaled by an increase in mitochondrial reactive oxygen species (ROS). In this study, we tested whether the NADH–mitochondria ROS pathway was involved in the reduction of INa in a nonischemic cardiomyopathic model and correlated the findings with myopathic human hearts. Nonischemic cardiomyopathy was induced in C57BL/6 mice by hypertension after unilateral nephrectomy, deoxycorticosterone acetate (DOCA) pellet implantation, and salt water substitution. Sham operated mice were used as controls. After six weeks, heart tissue and ventricular myocytes isolated from mice were utilized for whole cell patch clamp recording, NADH/NAD+ level measurements, and mitochondrial ROS monitoring with confocal microscopy. Human explanted hearts were studied using optical mapping. Compared to the sham mice, the arterial blood pressure was higher, the left ventricular volume was significantly enlarged (104.7±3.9 vs. 87.9±6.1μL, P<0.05), and the ejection fraction was reduced (37.1±1.8% vs. 49.4±3.7%, P<0.05) in DOCA mice. Both the whole cell and cytosolic NADH level were increased (279±70% and 123±2% of sham, respectively, P<0.01), INa was decreased (60±10% of sham, P<0.01), and mitochondrial ROS overproduction was observed (2.9±0.3-fold of sham, P<0.01) in heart tissue and myocytes of myopathic mice vs. sham. Treatment of myocytes with NAD+ (500μM), mitoTEMPO (10μM), chelerythrine (50μM), or forskolin (5μM) restored INa back to the level of sham. Injection of NAD+ (100mg/kg) or mitoTEMPO (0.7mg/kg) twice (at 24h and 1h before myocyte isolation) to animals also restored INa. All treatments simultaneously reduced mitochondrial ROS levels to that of controls. CD38 was found to transduce the extracellular NAD+ signal. Correlating with the mouse model, failing human hearts showed a reduction in conduction velocity that improved with NAD+. Nonischemic cardiomyopathy was associated with elevated NADH level, PKC activation, mitochondrial ROS overproduction, and a concomitant decrease in INa. Reducing mitochondrial ROS by application of NAD+, mitoTEMPO, PKC inhibitors, or PKA activators, restored INa. NAD+ improved conduction velocity in human myopathic hearts.
Highlights
Despite extensive research and novel treatments, conditions associated with deranged cardiac metabolism, such as heart failure or ischemia, are accompanied still by a substantial risk of arrhythmic sudden death [1]
We evaluated the role of the β-adrenergic receptor (β-AR), CD38, and the purinergic receptor [13,14,15,16], to determine how extracellular NAD+, which is membrane impermeable, affected in cardiac Na+ current (INa)
In our previous studies on the mechanism by which mutations in glycerol-3-phosphate dehydrogenase 1 like (GPD1L) protein cause reduced INa and Brugada Syndrome, we have shown that increased cytosolic NADH can downregulate the cardiac Na+ channel through protein kinase C (PKC) activation and mitochondrial reactive oxygen species (ROS) overproduction [9,10]
Summary
Despite extensive research and novel treatments, conditions associated with deranged cardiac metabolism, such as heart failure or ischemia, are accompanied still by a substantial risk of arrhythmic sudden death [1]. While implanted cardiac defibrillators have decreased sudden death risk, they can cause physical and psychological complications They are expensive, and do not address the underlying pathology that leads to arrhythmic risk [2,3]. NADH is known to oscillate with myocardial ischemia, and mitochondrial injury is associated with increased NADH and ROS levels [11,12]. These changes could contribute to reduced INa, conduction block, and arrhythmic risk known to exist with reduced cardiac contractility. To show relevance of the findings, the effect of NAD+ on conduction velocity (CV) in human failing hearts was evaluated
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