NITRIC-OXIDE SYNTHASE IN BOVINE SUPERIOR CERVICAL-GANGLION

We investigated the mechanism of increases in cyclic GMP levels in bov ine superior cervical ganglion (SCG) in response to muscarinic recepto r stimulation. Acetylcholine increased cyclic GMP levels in SCG. This increase was inhibited by N(G)-methyl-L-arginine (NMA), and the inhibi tion was reversed by L-arginine. Soluble nitric oxide (NO) synthase wa s partially purified from bovine SCG using 2',5-ADP Sepharose affinity chromatography. The resulting enzyme activity was Ca2+/calmodulin dep endent and required NADPH and tetrahydrobiopterin as cofactors. Supero xide dismutase protected and oxyhemoglobin blocked the effect of NO fo rmed by the enzyme. NMA inhibited the activity of the NO synthase. In western blots, an antibody generated against rat brain NO synthase spe cifically recognized the NO synthase from SCG as a 155-kDa protein ban d. Immunohistochemistry using the same antibody demonstrated that NO s ynthase was localized in postganglionic neuronal cell bodies of the SC G. Immunofluorescent labeling showed that some of the cells staining p ositive for dopamine-beta-hydroxylase also contained NO synthase. Thus , NO is synthesized in specific cells within bovine SCG, including sym pathetic neurons, and mediates the acetylcholine-induced stimulation o f soluble guanylyl cyclase.