A Central Role For Mitochondria in the Regulation of Cardiac Sodium Current

Abstract

Background: A mutant glycerol-3-phosphate dehydrogenase 1-like, A280V (A280V GPD1-L) reduces cardiac Na+ current (INa) and causes Brugada Syndrome. Recent data suggest that this effect is dependent on alterations in NADH, reactive oxygen species (ROS), and PKC activation. Since NADH and PKC can activate ROS production from mitochondria, we investigated the role of this organelle in mediating the effects of mutant GPD1-L and NADH on INa.Methods: HEK cells stably expressing the cardiac Na+ channel were used, and effects on INa were assessed by whole-cell patch clamp recording.Results: A280V GPD1-L caused a 2.48±0.17-fold increase of intracellular NADH level (n=3; P<0.001). Cytosolic NADH application (100 μM) or co-transfection with A280V GPD1-L resulted in significant decrease of INa (52±9% or 81±4%, respectively; P<0.01), which was reversed by 5-50 μM chelerythrine, 5 μM superoxide dismutase (SOD), 5-10 μM mitoTEMPO (a specific inhibitor to block mitochondrial superoxide generation), 1-5 μM rotenone (a complex I inhibitor), and 40-80 μM 4′-chlorodiazepam (an inhibitor of mitochondrial benzodiazepine receptor). The decreased INa induced by 30 nM PMA (60±7%, P<0.01) was prevented by SOD. Antimycin A (a complex III inhibitor known to produce ROS) at 20 μM decreased INa (51±4%, P<0.01). L-NAME (an inhibitor for uncoupled NOS), cyclosporin A (an inhibitor for mitochondrial permeability transition pore), and KN-93 (an inhibitor of CAMKII) had no effect on NADH reducing Na+ current.Conclusions: A280V GPD1-L appears to regulate Nav1.5 by altering the oxidized to reduced NAD(H) balance, which then activates mitochondrial ROS production through a PKC-dependent signaling mechanism. This ROS production leads to reduced INa. This signaling cascade may help explain the link between altered metabolism, conduction block, and arrhythmic risk.