1. A study has been made of the effect of neocuproine, a specific Cu(I) chelator, on vasodilator responses of rat isolated perfused tail artery to two nitrosothiols: S-nitroso-N-acetyl-D,L-penicillamine (SNAP) and S-nitroso-glutathione (GSNO). 2. Bolus injections (10 microl) of SNAP or GSNO (10(-7)-10(-3) M) were delivered into the lumen of perfused vessels pre-contracted with sufficient phenylephrine (1-7 microM) to develop pressures of 100-120 mmHg. Two kinds of experiment were made: SNAP and GSNO were either (a) pre-mixed with neocuproine (10(-4) M) and then injected into arteries; or (b) vessels were continuously perfused with neocuproine (10(-5) M) and then injected with either pure SNAP or GSNO. 3. In each case, neocuproine significantly attenuated vasodilator responses to both nitrosothiols, although the nature of the inhibitory effect differed in the two types of experiment. We conclude that the ability of exogenous nitrosothiols to relax vascular smooth muscle in our ex vivo model is dependent upon a Cu(I) catalyzed process. Evidence is presented which suggests that a similar Cu(I)-dependent mechanism is responsible for the release of NO from endogenous nitrosothiols and that this process may assist in maintaining vasodilator tone in vivo.