Increased catalase levels and hypoxanthine-enhanced nitro-blue tetrazolium staining in rat retina after ischemia followed by recirculation

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Abstract
In the present study, using retinal ischemia as a model, we examined if different periods of ischemia and recirculation influenced the generation of reactive oxygen species, i.e. in hydrogen peroxide generation and nitroblue tetrazolium (NET) reduction. Ischemia was induced for 30 and 90 min by ligation of the optic nerve with the vessels and recirculation was established by removing the ligature. The rats were sacrificed after 15 min or 3 days of recirculation. The retinas were separated from the pigment epithelium for measurements of catalase activity and examination of NBT staining. Compared to controls, the catalase activity was increased after 30 and 90 min of ischemia followed by 15 min of recirculation, and after 90 min of ischemia followed by 3 days of recirculation. As in controls, NBT staining was observed, both after 30 and 90 min of ischemia followed by 15 min of recirculation, in photoreceptors, in both plexiform layers, in some ganglion and glial cells, and, occasionally, in cells in the inner nuclear layer. Opposite to controls, addition of hypoxanthine to the NBT solution resulted in an increased staining in vessels in the inner nuclear layer in retinas subjected to 30 min of ischemia followed by 3 days of recirculation. The increased catalase activity suggests an increased amount of this free radical scavenger after ischemia followed by short-term and long-term recirculation. The hypoxanthine-enhanced NBT staining of blood vessel walls after ischemia followed by long-term recirculation indicates an activation of xanthine oxidase and an increased production of NBT reductants, some of which may represent oxygen free radicals.