Nitric oxide synthase (NOS) immunohistochemistry and nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry were used to investigate the distribution of nitroxergic, i.e., nitric oxide-synthesizing, neuronal perikarya and processes in the human ureterovesical junction (UVJ). Tissue specimens obtained from two cadaver kidney donors and two patients undergoing radical cystectomy for bladder cancer were examined. Clusters of NOS-immunoreactive neurons were localized in extramural ureterovesical ganglia. NOS-containing nerve fibers traveled within large extramural nerve trunks and marched among smooth muscle bundles. Extramural and intramural blood vessels were encircled by varicose NOS-positive axonal processes. The distribution of NOS immunoreactivity paralleled the staining pattern for NADPH-d activity. Urothelium stained strongly for NADPH-d activity but showed no NOS immunolabeling. Specimens from all four patients investigated showed similar staining patterns. Our results suggest that nitric oxide, a potent smooth-muscle-relaxing neurotransmitter in the autonomic nervous system, plays a physiologic role in opening the human UVJ.