Augmentation of cGMP/PKG pathway and colonic motility by hydrogen sulfide

Hydrogen sulfide (H₂S), like nitric oxide (NO), causes smooth muscle relaxation, but unlike NO, does not stimulate soluble guanylyl cyclase (sGC) activity and generate cyclic guanosine 5'-monophosphate (cGMP). The aim of this study was to investigate the interplay between NO and H₂S in colonic smooth muscle. In colonic smooth muscle from rabbit, mouse, and human, l-cysteine, substrate of cystathionine-γ-lyase (CSE), or NaHS, an H₂S donor, inhibited phosphodiesterase 5 (PDE5) activity and augmented the increase in cGMP levels, IP₃ receptor phosphorylation at Ser¹⁷⁵⁶ (measured as a proxy for PKG activation), and muscle relaxation in response to NO donor S-nitrosoglutathione (GSNO), suggesting augmentation of cGMP/PKG pathway by H₂S. The inhibitory effect of l-cysteine, but not NaHS, on PDE5 activity was blocked in cells transfected with CSE siRNA or treated with CSE inhibitor d,l-propargylglycine (dl-PPG), suggesting activation of CSE and generation of H₂S in response to l-cysteine. H₂S levels were increased in response to l-cysteine, and the effect of l-cysteine was augmented by GSNO in a cGMP-dependent protein kinase-sensitive manner, suggesting augmentation of CSE/H₂S by cGMP/PKG pathway. As a result, GSNO-induced relaxation was inhibited by dl-PPG. In flat-sheet preparation of colon, l-cysteine augmented calcitonin gene-related peptide release in response to mucosal stimulation, and in intact segments, l-cysteine increased the velocity of pellet propulsion. These results demonstrate that in colonic smooth muscle, there is a novel interplay between NO and H₂S. NO generates H₂S via cGMP/PKG pathway, and H₂S, in turn, inhibits PDE5 activity and augments NO-induced cGMP levels. In the intact colon, H₂S promotes colonic transit.NEW & NOTEWORTHY Hydrogen sulfide (H₂S) and nitric oxide (NO) are important regulators of gastrointestinal motility. The studies herein provide the cross talk between NO and H₂S signaling to mediate smooth muscle relaxation and colonic transit. H₂S inhibits phosphodiesterase 5 activity to augment cGMP levels in response to NO, which, in turn, via cGMP/PKG pathway, generates H₂S. These studies suggest that interventions targeted at restoring NO and H₂S homeostasis within the smooth muscle may provide novel therapeutic approaches to mitigate motility disorders.