Hypoxia and Hyperoxia Both Increase Superoxide Production in Nucleus Tractus Solitarius (NTS) Neurons in Rat Brain Tissue Slices

Maintaining brain slices in 95%O ₂ produces hyperoxia, oxidative stress and increased cell death, whereas decreasing control O ₂ to 20-40% significantly decreases both oxidative stress and cell death ( J. Neurophysiol. 98:1030-41, 2007) and maintains neuronal excitability (Matott et al., this meeting). In this study, beginning at 20-40%O ₂ , we tested the hypothesis that acute hyperoxia and hypoxia both increase the rate of superoxide (·O ₂ ^- ) production in NTS neurons. Medullary slices (400µm, weaned rats, 36-37 ^o C) were maintained using 2-sided superfusion. Intracellular ·O ₂ ^- production was measured using dihydroethidium (DHE, 2.5µM) added to the superfusate. The rate of ·O ₂ ^- production (fluorescence intensity units/min, FIU/min) increased during hypoxia (40/20%(0%O ₂ ) and was blocked by myxothiazol (10µM; inhibits ·O ₂ ^- produced by mitochondrial Complex III). ·O ₂ ^- production also increased during hyperoxia (20(95%O ₂ ), but to a lesser extent than during hypoxia, and was not blocked by myxothiazol. We propose that NTS neurons undergo redox stimulation and/or stress via increased ·O ₂ ^- production during both hypoxia (Complex III) and hyperoxia (Complex I ?). We also hypothesize that the smaller increase in FIU _DHE /min during hyperoxia reflects increased production of ·NO during hyperoxia, which reacts with ·O ₂ ^- (and thus consumes ·O ₂ ^- ) to yield peroxynitrite. ONR N000140710890, NIH R01 HL 56683-10.