We have measured the effects of hydrogen peroxide on the ability of the isolated rabbit cornea to maintain normal hydration during perfusion, on the structure of the endothelial cells, and on the redox state of glutathione and the activity of the hexose monophosphate shunt in these cells. Swelling of isolated corneas was immediate and severe upon exposure to 50μM H2O2in the absence of glucose, or to 200μM H2O2in the presence of glucose. The presence of glucose with 50μM H2O2delayed the onset of swelling for two hours and markedly decreased its severity. The concentration of glutathione in the endothelium, and its redox state, were unaltered by 50μM H2O2in the presence of glucose, but in its absence glutathione loss was significant and the fraction in the oxidized state was greatly increased. H2O2was removed from the medium by reaction with the cornea at a concentration-dependent rate. It was calculated that the reaction rate at 50μH H2O2was about twice that which could be accounted for by the increased yield of WADPH generated by the stimulation of the hexose monophosphate shunt. H2O2 may also react with endothelial cell membranes, which could account for the marked changes in cell structure seen in the scanning electron microscope.