O. B. Vadzyuk, Yu. Yu. Mazur, S. O. Kosterin

Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: vadzyuk@biochem.kiev.ua

In previous study we demonstrated the presence of ATP-sensitive potassium current in the inner mitochondrial membrane, which was sensitive to diazoxide and glybenclamide, in mitochondria isolated from the rat uterus. This current was supposed to be operated by mitochondrial ATP-sensitive potassium channel (mitoK_ATP). Regulation of the mitoK_ATP in uterus cells is not studied well enough yet.
It is well known that the reactive oxygen species (ROS) can play a dual role. They can dama­ge cells in high concentrations, but they can also act as messengers in cellular signaling, mediating survival of cells under stress conditions. ROS are known to activate mitoK_ATP during the oxidative stress in the brain and heart, conferring the protection of cells. The present study examined whether ROS mediate the mitoK_ATP activation in myometrium cells.
Oxidative stress was induced by rotenone. ROS generation was measured by 2′,7′-dichlorofluorescin diacetate. The massive induction of ROS production was demonstrated in the presence of rotenone. Hyperpolarization of the mitochondrial membrane was also detected with the use of the potential-sensitive dye DiOC₆ (3,3′-dihexyloxacarbocyanine iodide). Diazoxide, a selective activator of mitoK_ATP, depolarized mitochondrial membrane either under oxidative stress or under normal conditions, while mitoK_ATP blocker glybenclamide effectively restored mitochondrial potential in rat myocytes. Estimated <K_1/2> value for diazoxide to mitoK_ATP under normoxia was four times higher than under oxidative stress conditions: 5.01 ± 1.47∙10-6 М and 1.24 ± 0.21∙10-6 М respectively.
The ROS scavenger N-acetylcysteine (NAC) successfully eliminates depolarization of mitochondrial membrane by diazoxide under oxidative stress. These results suggest that elimination of ROS by NAC prevents the activation of mitoK_ATP under oxidative stress. Taking into account the higher affinity of diazoxide to mitoK_ATP under stress conditions than under normoxia, we conclude that the oxidative stress conditions are more favourable than normoxia for the activation of mitoK_ATP­. Thus we hypothesize that the ROS regulate the activity of the mitoK_ATP in myocytes.

Keywords: diazoxide, mitochondrial ATP-sensitive K(+)-channel, reactive oxygen species