We studied the effects of L-NG-nitro-arginine (L-NOARG), which inhibits nitric oxide (NO) biosynthesis from L-arginine, on the non-adrenergic, non-cholinergic (NANC)-mediated relaxation induced by nicotine in isolated rat duodenum. L-NOARG reduced nicotine-induced relaxation, and L-arginine prevented the inhibitory effect of L-NOARG. However, L-NOARG did not inhibit the tetrodotoxin-insensitive relaxation induced by adenosine 5'-triphosphate, alpha, beta-methyleneadenosine 5'-triphosphate, thyrotropin-releasing hormone or the calcitonin gene-related peptide. Endogenous NO thus could possibly be involved in the NANC-mediated relaxation of rat duodenum induced by nicotine.