Role Of Endogenous Hydrogen Sulfide In Vas Deferens Motility And Its Underlying Mechanisms

OBJECTIVE:To investigate the expression, localization, function and molecular targets of endogenous hydrogen sulfide (H2S) in mouse, rat and human vas deferens (VD).METHOD:Mature male Wistar rat, Balb/c mice (8 to 12 weeks old) and VD from patients undergoing surgery for monorchidism was used. Cystathionine-β-synthetase (CBS) and cystathionine-y-lyase (CSE) protein were assayed by western blotting and immunofluorescence techniques in rat, mouse and human VD. H2S release was measured bythe methylene blue assay method and smooth muscle contractility was studied by tension recording method in isolated VD strips. By in vivo studies, the role of H2S on seminal emission and litter size was identified. Furthermore, the target site of H2S in the vas deferens smooth muscle also was investigated by using isolated VD strips.RESULTS:The CBS and CSE protein were expressed in rat, mice and human VD. Immunohistochemical analysis revealed that the muscular tissue of VD stained strongly for CSE and CBS. Immunoreactivity for CBS and CSE was also observed in the luminal epithelium. VD tissue homogenates under basal conditions produced≈1nmol of H2S per mg of tissue and that addition of exogenous L-Cys boosted the production of H2S by 5-fold. The specificity of the assay and the involvement of both CBS and CSE were confirmed by the finding that H2S production was prevented by either PAG, an inhibitor of CSE, or AOAA, an inhibitor of CBS. Both NaHS and L-Cys produced dose-related relaxation of the rat VD norepinephrine (NE)-mediated contraction (IC50,9.8±2.4μM,67.9±18.0μM, respectively). NaHS also decrease the basal tone in the spontanous contraction of VD. L-Cys, an endogenous H2S donor, also caused a slowly developing decrease in the spontanous contraction of VD, in contrast, PAG, an inhibitor of CSE, caused a slowly developing increase. SAM, a CBS activator, produced a significant decrease in basal tone. These results confirm that endogenous H2S pathway involved in maintaining basal tone of the spontanous contraction of VD. In the in vivo studies, NaHS(28μmol-kg-1, ip) increased the sperm number of vas deferens after seminal emission. H2S relaxed the smooth muscle of vas deferens by opening the large conductance calcium-activated potassium channels(BKca) through targeting the-SH group via S-sulfhydration.CONCLUSIONS:Our findings demonstrated, for the first time, that CBS and CSE, both H2S synthesizing enzymes, were expressed in rat, mice and human VD, and that the L-Cys/H2S signaling pathway is involved in mediating VD smooth-muscle relaxation. The target site of H2S was the BKca by redox-mediated mechanism in the smooth muscle of vas deferens. Further studies are required to evaluate the pathophysiological role of H2S in functional abnormalities of the VD, such as emission and premature ejaculation.