We investigated the effects of prolonged treatment with Escherichia coli lipopolysaccharide (LPS) on the responses to sodium nitroprusside (SNP) in endothelium-denuded rat aortic strips. Incubation of the aortic strips with LPS for 24 h dramatically attenuated relaxation and guanosine 3',5'-cyclic monophosphate (cGMP) formation by SNP, which were significantly restored by the inhibition of nitric oxide (NO) production with N omega-nitro-L-arginine. In the aorta coincubated with LPS and protein synthesis inhibitor (dexamethasone or cycloheximide, which prevents induction of endotoxin-inducible NO synthase), no attenuation of the relaxation was observed and the cGMP formation was significantly restored. Relaxation response to 8-bromo-cGMP or papaverine was not attenuated, even after 24 h of incubation. These results suggest that the attenuation of SNP responses is mainly associated with a decrease in the activation of guanylate cyclase (GC) as a consequence of the prolonged exposure to muscle-derived NO. Moreover SNP in the presence of methylene blue evoked a small but apparent relaxation of 24-h-incubated aorta without significant elevation of cGMP, suggesting the involvement of cGMP-independent pathways in the remaining relaxation produced by SNP.