Neurogenic-derived 6-nitrodopamine is the most potent endogenous modulator of the mouse urinary bladder relaxation.

6-Nitrodopamine (6-ND) modulates vas deferens, seminal vesicles, and corpus cavernosum contractility; however, its role on the lower urinary tract organs has not been evaluated. Investigations of isolated urinary bladders from wild-type (WT) mice revealed 6-ND release was comparable to that of dopamine and adrenaline, whereas noradrenaline was hardly detected, as assessed by liquid chromatography coupled to tandem mass spectrometry. In vitro, 6-ND induced concentration-dependent relaxations in carbachol pre-contracted bladders with high potency (pEC: 8.04 ± 0.86), independently of eNOS/sGC activity. Co-incubation of 6-ND (1-10 μM) antagonizes the contractile effects of acetylcholine (p < 0.05). Experiments using nitric oxide synthase (NOS) knockout mice demonstrated that 6-ND release from isolated urinary bladder was significantly reduced by neuronal NOS (nNOS) deletion and abolished by triple NOSs deletion (n/i/eNOS), while no significant changes were observed in endothelial (eNOS) or inducible (iNOS) knockout mice. Incubation with tetrodotoxin resulted in a significant decrease in 6-ND release in bladders obtained from WT, but not in nNOS mice. The bladders from nNOS and n/i/eNOS mice exhibited significantly higher contractile responses to electric field stimulation (EFS), compared to eNOS, iNOS, or WT bladders. The hyperreactivity observed in triple NOS knockouts was reversed by the incubation with bladder mucosal layer obtained from a donor WT mice, but not with the muscular layer. These findings clearly demonstrate 6-ND is the most potent endogenous relaxing agent of urinary bladder, and inhibition of its release is associated with bladder hyperreactivity.