Calcium-independent uncoupling activity of palmitic acid in liver mitochondria is regulated by the ion fluxes causing the interconversion of Δψ and ΔpH across the inner membrane

Abstract

The effect of ion fluxes across the inner membrane on calcium-independent uncoupling activity of palmitic acid was investigated in experiments on rat liver mitochondria energized by the oxidation of succinate. The following compounds were used as the inductors of ion fluxes: the K+/H+ antiporter nigericin causing transformation of ΔpH into electrical potential difference (Δψ) across the inner membrane; tetraphenylphosphonium (TPP+) that freely crosses phospholipid membranes; protonophore carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP) that induces a flow of H+ from the intermembrane space into the matrix and reduces Δψ and ΔpH. It was found that nigericin at a concentration of 20 nM, which causes an increase in maximal Δψ, partially inhibits the ability of palmitic acid to reduce Δψ and stimulates mitochondrial respiration. A specific inhibitor of the ATP/ADP antiporter (carboxyatractylate) and a substrate of the aspartate/glutamate antiporter (glutamate) increase Δψ and partially inhibit mitochondrial respiration in the presence of palmitic acid. Under these conditions, 10 μM cyclosporin A also inhibits respiration but has no effect on Δψ. The specific uncoupling activity of palmitic acid (VU) and its specific components that characterize participation of the ATP/ADP antiporter (VCatr), aspartate/glutamate antiporter (VGlu), and cyclosporin-A-sensitive system (VCsA) in the palmitic acid-induced uncoupling were estimated. It was shown that nigericin substantially reduces VU, VCatr and VGlu but increases VCsA. TPP+ at a concentration of 20 μM increases VU and VGlu, does not affect VCatr and reduces VCsA. FCCP at concentrations of 20 and 40 nM reduces Δψ by not more than 17% but does not affect VU, VCatr, VGlu and VCsA. The results suggest that the calcium-independent uncoupling effect of palmitic acid in liver mitochondria is caused by the return of protons to the matrix with participation of ADP/ATP and aspartate/glutamate antiporters and owing to activation of cyclosporin A-sensitive electron transport along the respiratory chain without affecting Δψ. The induced ion fluxes across the inner mitochondrial membrane can be considered as a factor of the calcium-independent regulation of uncoupling activity of palmitic acid in liver mitochondria with participation of the ADP/ATP and aspartate/glutamate antiporters and of the cyclosporin A-sensitive electron transport system.