Calcium- and ADP-Magnesium-induced respiratory uncoupling in isolated cardiac mitochondria: Influence of cycolsporin A

This study was designed to determine the effect of calcium and ADP-Mg on the oxidative phosphorylation in isolated cardiac mitochondria. The influence of cyclosporin A was also evaluated. The mitochondria were extracted from rat ventricles. Their oxidative phosphorylations were determined in two respiration media with different free Ca²⁺ concentrations. Respiration was determined with palmitoylcarnitine and either ADP- or ADP-Mg. With elevated free Ca²⁺concentrations and ADP-Mg, the transition state III to state IV respiration did not occurred. The ADP:O ratio was reduced. The phenomenon was not observed in the other experimental conditions (low free Ca²⁺ concentration with either ADP^- or ADP-Mg or elevated free Ca²⁺ concentration with ADP^-). Uncoupling was allied with a constant AMP production, which maintained an elevated ADP level in the respiration medium and prevented the return to state IV respiration. It was also observed in a respiration medium devoid of free Ca²⁺ when the mitochondria were pre-loaded with Ca²⁺. Uncoupling was inhibited by cyclosporin A. Furthermore, the Krebs cycle intermediates released from¹⁴C-palmitoylcarnitine oxidation revealed that succinate was increased by elevated free Ca²⁺ and ADP-Mg. Succinate is a FAD-linked substrate with low respiration efficiency. Its accumulation could account for the decreased ADP:O ratio. The Ca²⁺- and ADP-Mg-induced uncoupling might be partly responsible for the mechanical abnormalities observed during low-flow ischemia. (Mol Cell Biochem 000: 000-000, 1999)