Glutathione (GSH) plays a key role in defense against oxidative stress. The availability of GSH is ensured in tissues by systems devoted to its maintenance in the reduced state and by the flux of GSH and cysteine between sites of biosynthesis and sites of utilization. Little is known about the effect of oxidative stress on the distribution of low-molecular-mass thiols and their exchange rate between tissues. In this study, we found that a slow infusion of diamide (a specific thiol-oxidizing compound) evoked a dramatic increase in blood cysteine in rats. Our data suggest that inter-organ exchange of cysteine occurs, that cysteine derives from both glutathione via gamma-glutamyl transpeptidase and methionine via homocysteine and the trans-sulfuration pathway, and that these pathways are considerably influenced by oxidative stress.