Abstract 208: Augmented Cerebrovascular Responses to Diazoxide, an Opener of MitokAtp Channels, Following Middle Cerebral Artery (MCA) Occlusion in Rats

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Abstract

Previous research has not examined the effects of ischemia on cerebral vascular responses to mitochondrial activation in experimental strokes caused by occlusion of the MCA (MCAO). We investigated the role and mechanisms of mitochondrial derived vasoreactivity in the MCA of male SD rats following 90 min ischemia/48 h reperfusion injury. Ischemia was induced ipsilaterally (I) and the contralateral (C) side was non-ischemic. Electron microscopy showed disrupted mitochondrial morphology on the I side. Western blots for expression of mitochondrial proteins (Mean±SEM of immunoband intensity normalized to β-actin and as C vs. I): DRP-1 (1±0.1 vs. 3.1±0.2); VDAC (0.4±0.1 vs. 0.8±0.2); and complex-V (1.3±0.3 vs.2.0±0.3) as well as the non-mitochondrial proteins: phosphorylated (ph) ph-nNOS (0.4±0.1 vs. 0.7±0.1); ph-eNOS (0.2±0.06 vs. 0.7±0.2); COX-1 (0.5±0.1 vs. 0.8±0.1); and COX-2 (0.16±0.04 vs. 0.3±0.03) were elevated on I compared with C. The I mitochondrial membrane potential was greater (165±7 %) compared with C using TMRE fluorescence. Mitochondrial membrane depolarization decreased the TMRE intensity in both groups (100 to 80±3 vs. 165±7 to 70±4). Vascular responses of the MCA were characterized using the isolated, pressurized artery technique. Vasodilation in response to Ach (12±2 vs. 3±1), BK (42±9 vs. 32±6), SNP (75±7 vs. 38±4), and vasoconstriction to serotonin (65±8 vs. 33±7), were significantly decreased in I compared to C MCAs. On the other hand, 50 μM DZ induced vasodilation was enhanced in I arteries compared with C (5±1 vs. 17±2). Diazoxide induced dilation was decreased in the presence of the NOS inhibitor L-NAME (5±1 to 1±2 vs. 17±2 to 3±2) and the non-selective COX inhibitor indomethacin (5±1 to 1±2 vs.17±2 to 6±3). Our results indicate that experimental stoke has an major effect on mitochondria via inducing mitochondrial biogenesis leading to altered morphology, protein expression and function. Furthermore, the nitric-oxide and prostanoidpathways appear to be involved in enhanced diazoxide mediated vasodilation after MCAO. We speculate that targeting mitochondria may be useful therapy for improving outcome in stroke patients.

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