Influences of glutathione and sulfhydryl containing compounds on aqueous humor outflow function

Several sulfhydryl reactive compounds have previously been shown to influence aqueous humor outflow facility. The purpose of the current study was to investigate the effect of glutathione depletion on certain of these sulfhydryl actions. Enucleated calf, monkey, and human eyes were perfused via the anterior chamber by the constant pressure (15 mmHg) technique. In calf eyes, perfusion of 10 m m cysteamine produced a small (−23%, P = 0·03) decrease in outflow facility that was also observed after hyaluronidase pretreatment. In contrast, following pretreatment with 1 m m BCNU [1,3 bis(2 chlorethyl)-1-nitrosourea], an inhibitor of glutathione reductase, and 10 m m diamide, a glutathione oxidant, which did not by themselves significantly affect outflow facility, perfusion of cysteamine resulted in an opposite effect—a remarkably large (+90%, P < 0·001) increase in outflow facility. Other reduced and oxidized sulfhydryl-containing compounds such as cysteine, β-mercaptoethanol, and glutathione, itself, as well as the non-sulfhydryl reducing agent, ascorbic acid, were substituted for cysteamine in this protocol and found to produce similar effects of varying magnitudes. In general, the reduced sulfhydryl containing compounds and ascorbic acid were the most effective. Pretreatment with BCNU alone without diamide did not produce this effect. Treatment with BCNU and diamide resulted in a greater than 75% decrease in reduced glutathione levels and a concomitant tenfold increase in glutathione mixed disulfide levels (0·229 vs. 0·030 μmol g −1 wet weight) in the calf trabecular meshwork. The subsequent perfusion with cysteamine reversed this mixed disulfide formation. In one pair of enucleated human eyes treated with BCNU, diamide and then cysteamine, loss of cell to cell attachments and breaks in the inner wall of Schlemm's canal were observed, a morphological effect similar to that of certain sulfhydryl binding compounds. Data from combination studies of BCNU and diamide with the sulfhydryl binding agents, ethacrynic acid and PCMBS ( p-chloromercuribenzene sulfonate), are presented which suggest that there are multiple sulfhydryl-sensitive sites in the outflow pathway that may be involved in the regulation of outflow resistance.