INFLUENCE OF POST-HYPOXIA REOXYGENATION CONDITIONS ON ENERGY-METABOLISM AND SUPEROXIDE PRODUCTION IN CULTURED NEURONS FROM THE RAT FOREBRAIN

Brain reperfusion and/or reoxygenation may be of particular importance in the etiology of neuronal damage after hypoxic-ischemic insult in n eonates, especially with reference to the generation of free radicals, To investigate this issue, the influence of either standard reoxygena tion or transient hyperoxia was studied on the consequences of severe hypoxia in a model of cultured neurons isolated from the fetal rat bra in. Culture dishes were exposed for 6 h to hypoxia (95% N-2/5% CO2). T hey were then placed under normoxia (95% air/5% CO2) or hyperoxia (95% O-2/5% CO2) for 3 h, and finally returned to normoxia. Control cultur es were kept under normoxic conditions. Cell morphology, protein conce ntrations, lactate dehydrogenase leakage, energy metabolism, as reflec ted by specific transport and incorporation of 2-D-[H-3]deoxyglucose, as well as superoxide radical formation were analyzed as a function of time. PO2 values in the cell incubating medium were decreased by 78% by hypoxia and increased by 221% by hyperoxia. No morphologic alterati on could be noticed before 72 h posthypoxia, when cell degeneration be came apparent, with a concomitant reduction in protein contents. Hypox ia-reoxygenation induced a transient cellular hypermetabolism, as show n by a 36% increase in 2-D-[H-3]deoxyglucose uptake 24 h after hypoxia , and then a 23% decrease below control values at 72 h. It also led to a sharp increase in the formation of superoxide radicals (+108%). Tra nsient hyperoxia during reoxygenation did not exacerbate these events, and thus would not enhance their detrimental effects on cell integrit y.