Diazoxide affects mitochondrial bioenergetics by the opening of mKATP channel on submicromolar scale.

BACKGROUND: Cytoprotection afforded by mitochondrial ATP-sensitive K+-channel (mKATP-channel) opener diazoxide (DZ) largely depends on the activation of potassium cycle with eventual modulation of mitochondrial functions and ROS production. However, generally these effects were studied in the presence of Mg∙ATP known to block K+ transport. Thus, the purpose of our work was the estimation of DZ effects on K+ transport, K+ cycle and ROS production in rat liver mitochondria in the absence of Mg∙ATP. RESULTS: Without Mg·ATP, full activation of native mKATP-channel, accompanied by the increase in ATP-insensitive K+ uptake, activation of K+-cycle and respiratory uncoupling, was reached at ≤0.5 µM of DZ,. Higher diazoxide concentrations augmented ATP-insensitive K+ uptake, but not mKATP-channel activity. mKATP-channel was blocked by Mg·ATP, reactivated by DZ, and repeatedly blocked by mKATP-channel blockers glibenclamide and 5-hydroxydecanoate, whereas ATP-insensitive potassium transport was blocked by Mg2+ and was not restored by DZ. High sensitivity of potassium transport to DZ in native mitochondria resulted in suppression of mitochondrial ROS production caused by the activation of K+-cycle on sub-micromolar scale. Based on the oxygen consumption study, the share of mKATP-channel in respiratory uncoupling by DZ was found. CONCLUSIONS: The study of mKATP-channel activation by diazoxide in the absence of MgATP discloses novel, not described earlier, aspects of mKATP-channel interaction with this drug. High sensitivity of mKATP-channel to DZ results in the modulation of mitochondrial functions and ROS production by DZ on sub-micromolar concentration scale. Our experiments led us to the hypothesis that under the conditions marked by ATP deficiency affinity of mKATP-channel to DZ can increase, which might contribute to the high effectiveness of this drug in cardio- and neuroprotection.

The water-soluble vitamin E analogue Trolox protects against ischaemia/reperfusion damage in vitro and ex vivo. A comparison with vitamin E.

We investigated the activities, both in vitro and ex vivo, of the water-soluble vitamin analogue Trolox in a model of isolated heart ischaemia-reperfusion and we compared them with those of alpha -tocopherol. Isolated rat hearts were perfused with Krebs-Henseleit solution. For in vitro experiments, the hearts were perfused with Trolox (20 micromol l (-1)) and were subsequently subjected to 20 min of global ischaemia and 40 min of post-ischaemic reperfusion. For ex vivo experiments, either Trolox or alpha -tocopherol (10 mg kg(-1) ) were administered by gastric gavage 60 min before excision of the heart. Various parameters of cardiac function were evaluated and oxidative damage was assessed by TBARS production. Trolox significantly enhanced cardiac recovery after ischaemia/reperfusion, both when it was perfused in vitro and after its oral administration. Vitamin E also favourably affected cardiac recovery but did so less effectively than Trolox. Further, the production of TBARS was significantly inhibited by Trolox, suggesting that its beneficial effects are due to its antioxidant activities. In conclusion, perfusion of isolated rat hearts with low concentrations of the water-soluble vitamin E analogue Trolox effectively enhances cardiac recovery after a 20 min ischaemic period and decreases reperfusion-induced oxidative damage. Interestingly, Trolox retains its activities after oral administration. Vitamin E, when administered per os, also increases functional recovery but does so less potently than Trolox. These differential effects are likely due to the scavenging, by Trolox, of reactive oxygen species generated in the water phase.