Organs of the digestive tract, including pancreas, small intestine, and colon, have mechanisms to modulate plasma thiol-disulfide balance. Because plasma glutathione disulfide (GSSG) concentration may be elevated from < 1 microM in control rats to over 25 microM during oxidative stress, we examined whether GSSG was cleared from rat mesenteric vasculature. When 100 microM GSSG was perfused through the gut via the superior mesenteric artery, an average of 45% was lost in a single pass. Results showed that gamma-glutamyltransferase (gamma-GT)-dependent and -independent mechanisms were involved in GSSG loss. Acivicin (AT125) treatment inhibited gamma-GT activity in the mesenteric vasculature by 94% and attenuated the loss of GSSG equivalents by 44%. These results supported a role for gamma-GT in GSSG loss from the mesenteric vasculature but indicated that still other mechanisms were involved in GSSG clearance. Elevations of portal levels of glutathione (GSH) and the mixed disulfide of cysteine and GSH (CySSG) also occurred with vascular GSSG perfusion and could account for about 40% of GSSG equivalents lost. Because portal elevations of GSH and CySSG were not inhibited by AT125, they were formed by a gamma-GT-independent mechanism(s). Given that cysteine was present in the mesenteric vasculature, the most likely mechanism to explain GSH and CySSG formation was via nonenzymatic thiol-disulfide exchange between GSSG and cysteine. Uptake of vascular GSSG by pancreas, small intestine (jejunum and ileum), or colon apparently did not occur as tissue contents of GSSG or GSH were not elevated, except for a small elevation of GSH in pancreas when mesenteric gamma-GT was inhibited with AT125. Additionally, GSSG was not transported from mesenteric vasculature into the small intestinal lumen because luminal levels of GSSG or GSH were not elevated. Further, total cysteine equivalents in lumen were unchanged indicating that GSSG was not transported to lumen and degraded to cystine by gamma-GT and dipeptidases localized to the intestinal brush-border. These results indicate that GSSG present in mesenteric vasculature is metabolized in the vascular compartment by gamma-GT-dependent and -independent reactions; together, these account for over 80% of lost GSSG equivalents. They also suggest that organs of the mesentery may play a quantitatively important role in plasma GSSG clearance and modulation of vascular thiol-disulfide balance.