Segments of endothelium-denuded pulmonary arterial and aortic rings and strips of corpus cavernosum from rabbits were superfused with Krebs solution alone and then Krebs medium containing 0.1-0.5 mM N omega-Nitro-L-Arginine. Photorelaxation in response to ultraviolet light (366 nm) was significantly enhanced by 50 microM methylene blue in all preparations; 25 microM methylene blue also increased photorelaxation in corpus cavernosum and pulmonary artery. Enhanced photorelaxation was associated with increased tissue cGMP. This effect was significantly attenuated by 10 microM hemoglobin and was associated with decreased tissue cGMP but was unaffected by superoxide dismutase. We speculate that UV-generated free radicals convert the phenothiazine moiety of methylene blue to a phenyl radical which activates guanylate cyclase and thus enhances smooth muscle relaxation.