The present study was undertaken to investigate the role of nitric oxide (NO) in erectile physiology by correlating its action with the existence and activity of nitric oxide synthase (NOS), which produces NO. We applied Western blot analysis in both human and rat penile tissue. In the rat, reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase staining and spectrophotometric assay were also performed, in addition to in vivo electroerection study with pharmacological manipulation. Western blot analysis identified a protein of 155 KDa identical to the neural form of NOS in the human and rat penis. The NOS blot densities in the two species were similar, and both were lower than that in the rat cerebellum. Histochemical staining localized NOS to neurons innervating the corpora cavernosa, including the pelvic plexus, the cavernosal nerves and their terminal fibers within the corporeal erectile tissue, and dorsal penile nerves. NOS activity was also found in the cerebellum, urethra, penis, and urinary bladder, in decreasing order of intensity. Intracavernous injections of NOS inhibitor (L-NOARG or L-NAME in concentrations from 10^-6 M to 10^-3 M suppressed electrostimulation-induced erection in a concentration-dependent manner. Subsequent intracavernous injection of L-Arginine (10^-2 M) partially restored the erection. The neural form of constitutive NOS in the corpora cavernosa synthesizes NO, which mediates penile erection. Determination of cavernosal NOS expression or activity may permit characterization of certain pathological conditions that cause impotence.