Previously we have demonstrated functional NOS 1 in large arteries. Since resistance arteries largely determine blood pressure, this study examined if functional NOS 1 also exists in resistance arteries. Phenylephrine contraction was measured in the absence and presence of the NOS 1 inhibitor N⁵-(1-imino-3-butenyl)-L-ornithine (VNIO) in isolated mesenteric resistance arteries (endothelium-intact and denuded) from Sprague-Dawley rats. For NOS 1 activity and expression, the mesenteric arterial bed was separated into cytosolic and particulate fractions. NOS activity was assayed measuring the conversion of ³H-arginine to ³H-citrulline inhibited by a nonselective NOS inhibitor or VNIO. VNIO increased phenylephrine sensitivity in endothelium-intact and denuded arteries. In cytosolic and particulate fractions of the arterial bed, ~40% of NOS activity was inhibited by VNIO. Immunoprecipitation and Western blot analysis revealed two NOS 1 immunoreactive bands. One band corresponded to the rat brain isoform, while the second was of a slightly lower molecular weight. The cytosolic fraction contained both isoforms while the particulate fraction had only the lower molecular weight form. These studies demonstrate the existence of functional NOS 1 in resistance arteries.