Hydroxyl Radical Generation in the Cat Retina During Reperfusion Following Ischemia

There is increasing evidence that oxygen-derived free radicals are generated during the early phase of reperfusion, and account for part of the damage caused by transient ischemia in various tissues. To study this in the retina, cats were injected intravenously with sodium salicylate (100 mg kg^-1), which reacts as a hydroxyl radical trap to form 2,3- and 2,5-dihydroxybenzoic acids (DHBA). Thirty minutes following injection, the retina of one eye of each animal was subjected to ischemia by intraocular pressure elevation via cannulation of the anterior chamber, while the fellow eye served as a sham-operated control. Ischemia was induced for 60 min (six eyes) and 90 min (eight eyes) followed by 5 min of reperfusion. In six other eyes, ischemia was induced for 90 min without reperfusion. After enucleation, the retinas were immediately removed, placed in ice-cold buffer and the retinal levels of 2,3- and 2,5-DHBA were quantitated by high pressure liquid chromatography, coupled with electrochemical detection. Results were normalized and expressed as ng DHBA μg^-1 salicylate mg^-1 retinal protein. After 60 min of ischemia followed by reperfusion the normalized levels of 2,3- and 2-5-DHBA were no different in the experimental and control retinas. However, the levels of both 2,3- and 2,5-DHBA were significantly higher in the retinas subjected to 90 min ischemia followed by reperfusion than in the control tissues (P = 0·012 and P = 0·036, n = 8 respectively). Following 90 min ischemia without reperfusion, the normalized dihydroxybenzoate levels in the retinas were no higher than in their controls. The relative increase of the normalized levels of the hydroxyl radical markers in the retinas between the 90 min ischemia and reperfusion and 90 min ischemia without reperfusion was three (for 2,5-DHBA) and eight-fold (for 2,3-DHBA) higher. These findings provide quantitative data and evidence that hydroxyl radicals are generated in the cat retina during the early phase of reperfusion following 90 min of ischemia.