Introduction: It has been reported that activation of the mitochondrial K channel attenuates mitochondrial potential without affecting ATP production in vitro. To elucidate the effects of diazoxide on mitochondrial potential and ATP production in vivo, mitochondrial potential, NADH fluorescence, DC potential and onset time of ischemic depolarization were examined in the rat cortex. Methods: Male Wistar rats (n=5) were anesthetized with 1% halothane and were artificially ventilated. Two cranial windows were made on the left parietal cortex. For the measurement of mitochondrial potential, a potentiometric dye, JC-1 (2 micro l), was injected into the parietal cortex through the two cranial windows. After a 30 minute equalization period, DC electrodes were inserted into the cortex, where JC-1 was injected. Then a mitochondrial ATP-sensitive K+ channel opener, diazoxide (1mmol/l in 1.5 mmol/l NaOH, 150 mmol/l NaCl), and its carrier (1.5 mmol/l NaOH, 150 mmol/l NaCl) were continuously infused at the rate of 0.5 micro l/min through the DC electrodes. Five minutes after the onset of infusion, 4vessel occlusion was performed for 10 minutes to observe the onset time of ischemic depolarization. Mitochondrial potential The cortex was illuminated with excitation light (485 nm). The intensity of red fluorescence (590 nm) and that of green fluorescence (530 nm) in an area adjacent to the DC electrode were measured every 20 seconds. The ratio of red:green fluorescence, indicative of mitochondrial potential, was normalized to that of the control value of the red:green ratio. NADH fluorescence The cortex was illuminated with excitation light (360 nm). The intensity of fluorescence (460 nm) in an area adjacent to the DC electrode was measured every 20 seconds. The intensity of fluorescence, indicative of aerobic glycolysis in mitochondria, was normalized to that of the control value. Results: Mitochondrial potential was decreased to 71·16 % of the control level with continuous infusion of diazoxide but was unchanged with infusion of the carrier (99·5%, p=0.006). There were no statistical differences in DC potential (diazoxide, -0.6·0.2 mV; carrier, -1.1·0.8 mV, p=0.18) and in intensity of NADH fluorescence (diazoxide, 101·1 %; carrier, 99·3 %, p=0.26) with infusion of diazoxide and its carrier. Onset times of DC depolarization after the initiation of 4-vessel occlusion were 3.3·1.3 min at the site of diazoxide infusion and 2.8·1.0 min (p=0.5) at the site of the carrier infusion. Conclusions In the present experiment, mitochondrial potential was decreased by 29% with activation of the mitochondrial ATP-sensitive K+ channel without affecting DC potential, intensity of NADH fluorescence and onset time of DC depolarization. This observation suggests that K+ gradient is an important factor making mitochondrial potential but is independent of energy production in mitochondria in vivo.
|Journal||Journal of Cerebral Blood Flow and Metabolism|
|Issue number||SUPPL. 1|
|Publication status||Published - Nov 13 2007|
ASJC Scopus subject areas
- Clinical Neurology
- Cardiology and Cardiovascular Medicine