Increasing evidence implicates reactive oxygen species, particularly hydrogen peroxide (H(2)O(2)), as intracellular and intercellular messengers in the brain. This raises the question of how the antioxidant network in the brain can be sufficiently permissive to allow messages to be conveyed yet, at the same time, provide adequate protection against oxidative damage. Here we present evidence that this is accomplished in part by differential antioxidant compartmentalization between glia and neurons. Based on the rationale that the glia-to-neuron ratio is higher in guinea-pig brain than in rat brain, we examined the neuroprotective role of the glial antioxidant network by comparing the consequences of elevated H(2)O(2) in guinea-pig and rat brain slices. The effects of exogenously applied H(2)O(2) on evoked population spikes in hippocampal slices and on edema formation in forebrain slices were assessed. In contrast to the epileptiform activity observed in rat hippocampal slices after H(2)O(2) exposure, no pathophysiology was seen in guinea-pig hippocampal slices. Similarly, elevated H(2)O(2) caused edema in rat brain slices, whereas this did not occur in guinea-pig brain tissue. The resistance of guinea-pig brain tissue to H(2)O(2) challenge was lost, however, when glutathione (GSH) synthesis was inhibited (by buthionine sulfoximine), GSH peroxidase activity was inhibited (by mercaptosuccinate), or catalase was inhibited (by 3-amino-1,2,4,-triazole). Strikingly, exogenously applied ascorbate, a predominantly neuronal antioxidant, was able to compensate for loss of any other single component of the antioxidant network. Together, these data imply significant roles for glial antioxidants and neuronal ascorbate in the prevention of pathophysiological consequences of the endogenous neuromodulator, H(2)O(2).