The present Study aims to examine whether resveratrol, a natural antioxidant present in red wine, restores the tolerance to nitroglycerin (GTN) on isolated human internal mammary artery (IMA), using an in vitro model of nitrate tolerance. IMA rings were obtained from 53 male patients undergoing coronary bypass operation. Nitrate tolerance was induced by incubating the artery ring with 100 mu M GTN for 90 minutes. Concentration-response curves to GTN (10(-9) to 10(-4) M) were obtained on IMA rings precontracted with noradrenaline. A low concentration (5 mu M) Of lucigenin was used as a tool to measure superoxide production in IMA segments. GTN produced concentration-dependent relaxation in isolated human IMA rings. Preexposure of artery rings to GTN reduced the relaxations to GTN [E-max values: 105 +/- 2%, and 76 +/- 3%, n = 10 to 12, P < 0.05; EC50 values (-log M): 6.72 +/- 0.05 and 4.95 +/- 0.06, P < 0.05, respectively]. Relaxation to sodium nitroprusside remained unchanged. Diminished relaxation to GTN is partially restored after removing endothelium or L-G-nitro-L-arginine (L-NOARG, 10-4 M) or superoxide dismutase (20 and 200 U/mL) or catalase (200 U/mL) pretreatments. Pretreatments with resveratrol (1, 10, and 20 mu M) for 20 minutes relatively improved the reduced relaxation to GTN in tolerant IMA rings. Coadministration of L-NOARG with resveratrol did not abolish the beneficial effect of resveratrol on nitrate tolerance. The inhibitory effect of resveratrol on GTN-induced tolerance was not abolished in arterial rings without endothelium. Exposure to GTN increased superoxide production in IMA segments with endothelium. Endothelium denudation, L-NOARG, or Superoxide dismutase pretreatments markedly inhibited the increased Superoxide production in tolerant arteries. Resveratrol (1 and 10 mu M) almost completely abolished basal or NAD(P)H-stimulated superoxide production in tolerant and nontolerant arteries. Vascular tolerance to GTN, in in vitro tolerant human IMA rings, can be induced by endothelial superoxide anions. Resveratrol partially restored the reduced relaxation to GTN by inhibiting NAD(P)-derived superoxide production in endothelium.