Calmidazolium Inhibits Na+/ca2+ Antiporter In Mitochondria

Abstract

A major function of Ca2+ in mitochondria is the regulation of intramitochondrial enzyme activity. A rise of Ca2+ in mitochondria can stimulate the oxidative metabolism and ATP production through activating Ca2±sensitive matrix dehydrogenases and controlling the synthesis of ATP. The main mechanism of the Ca2+ efflux from mitochondria in the cells of heart, brain, skeletal muscles, pancreas, and the majority of tumor cells is the Na+/Ca2+ exchange (NCXmito). Application of the NCX inhibitors (tetraphenylphosphonium cation, CGP37157) increases the concentration of Ca2+ ([Ca2+]m) and NADH level in mitochondria, and stimulates the oxidative metabolism. Little is known about the regulation of NCXmito. In this work it is shown, that calmodulin (CaM) participates in the regulation of the antiporter activity. To study the NCXmito activity, we used the Ca2±signal generating purinoreceptors of Ehrlich ascites tumor cells (EATC) and an inhibitor of CaM, R24571. R24571- and ATP-induced changes in the [Ca2+]i, [Ca2+]m, NADH fluorescence, and the membrane potential of mitochondria were recorded. ATP produced a transitory increase in [Ca2+]i, [Ca2+]m and NADH fluorescence. A preliminary addition of Na+/Ca2+ exchange inhibitors slightly changed the cytosolic signal, but suppressed the efflux of Ca2+ from mitochondria. Addition of ionomycin (0.5μM) in the time of the plateau in the mitochondrial Ca2±signal caused a decrease in Rhod-2 and NADH fluorescence up to the initial level, which indicated the Ca2±efflux from mitochondria. R24571 (3-5μM) caused a transitory Ca2±signal in the cytosol and an irreversible increase of NADH and Rhod-2 fluorescence. Ionomycin caused NADH and Rhod-2 fluorescence to recover to the initial level. Thus, R24571 inhibits the NCXmito in EATC, which suggests that its activity is regulated by CaM.