Pulmonary, Gastrointestinal and Urogenital Pharmacology Myosin light chain phosphatase activation is involved in the hydrogen sulﬁde-induced relaxation in mouse gastric fundus Ingeborg Dhaese, Romain A. Lefebvre ⁎ Heymans Institute of Pharmacology, Ghent University, Ghent, Belgium abstract article info Article history: Received 25 September 2008 Received in revised form 24 November 2008 Accepted 9 January 2009 Available online 21 January 2009 Keywords: Hydrogen sulﬁde Sodium hydrogen sulﬁde Calcium desensitization Myosin light chain phosphatase Rho-kinase Calyculin-A Y-27632 Mouse gastric fundus The relaxant effect of hydrogen sulﬁde (H 2 S) in the vascular tree is well established but its inﬂuence and mechanism of action in gastrointestinal smooth muscle was hardly investigated. The inﬂuence of H 2 S on contractility in mouse gastric fundus was therefore examined. Sodium hydrogen sulﬁde (NaHS; H 2 S donor) was administered to prostaglandin F 2α (PGF 2α )-contracted circular muscle strips of mouse gastric fundus, before and after incubation with interfering drugs. NaHS caused a concentration-dependent relaxation of the pre-contracted mouse gastric fundus strips. The K + channels blockers glibenclamide, apamin, charybdotoxin, 4-aminopyridin and barium chloride had no inﬂuence on the NaHS-induced relaxation. The relaxation by NaHS was also not inﬂuenced by L-NAME, ODQ and SQ 22536, inhibitors of the cGMP and cAMP pathway, by nerve blockers capsazepine, ω-conotoxin and tetrodotoxin or by several channel and receptor blockers (ouabain, nifedipine, 2-aminoethyl diphenylborinate, ryanodine and thapsigargin). The myosin light chain phosphatase (MLCP) inhibitor calyculin-A reduced the NaHS-induced relaxation, but the Rho-kinase inhibitor Y-27632 had no inﬂuence. We show that NaHS is able to relax PGF 2α -contracted mouse gastric fundus strips. The results suggest that in the mouse gastric fundus, H 2 S causes relaxation at least partially via activation of MLCP. © 2009 Elsevier B.V. All rights reserved. 1. Introduction Hydrogen sulﬁde (H 2 S), a colourless gas with the smell of rotten eggs, has traditionally been considered to be toxic leading to brain intoxication and the inhibition of the respiratory system (Beauchamp et al., 1984). However, it was found that H 2 S can also be produced endogenously in mammalian tissues from L-cysteine by two enzymes, cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE) (Stipanuk and Beck, 1982). It was suggested that H 2 S may operate as an endogenous neurotransmitter based on its endogenous production and its biological effects at physiological concentrations. H 2 S is as such a possible third gaseous transmitter, besides nitric oxide (NO) and carbon monoxide (CO), and has been reported to play a role in hippocampal long-term potentiation (Abe and Kimura, 1996) and to exert antinociceptive (Distrutti et al., 2006), exocrine prosecretory (Schicho et al., 2006) and smooth muscle relaxing (Cheng et al., 2004; Hosoki et al., 1997; Kubo et al., 2007a; Teague et al., 2002; Wang et al., 2008; Zhao et al., 2001) effects. Smooth muscle relaxation by H 2 S has been shown in different parts of the vascular tree. In this system, the ATP-dependent K + channel (K ATP channel) blocker glibenclamide generally inhibits at least partially the vasorelaxation induced by H 2 S, indicating that H 2 S is an activator of K ATP channels, leading to hyperpolarization and subsequently relaxa- tion (Zhao et al., 2001; Cheng et al., 2004; Tang et al., 2005; Webb et al., 2008). Both H 2 S-synthetizing enzymes have been shown in human and guinea-pig myenteric neurons (Schicho et al., 2006). Recently, CSE but not CBS was shown to be present in mouse colon myenteric neurons, and the intact colonic muscle layer containing the myenteric plexus generated detectable levels of H 2 S(Linden et al., 2008). Still, there are only a few reports concerning the inﬂuence of H 2 S on gastrointestinal smooth muscle. H 2 S relaxed concentration-depen- dently acetylcholine-induced contraction in guinea-pig ileum (Hosoki et al., 1997). This was conﬁrmed by Teague et al. (2002); these authors also showed that H 2 S inhibited spontaneous contractions of rabbit ileum and electrically induced contractions of the guinea-pig ileum. The latter response was unaffected by glibenclamide, indicating that the mechanism by which H 2 S induces inhibition of gastrointestinal smooth muscle differs from that in the cardiovascular system. The gastric fundus contains a smooth muscle layer with tonic activity, essential in gastric receptive relaxation. Although a principal role for NO in nonadrenergic noncholinergic relaxation at this level was established, evidence for neuronal release of a relaxant agent that is not NO, vasoactive intestinal peptide (VIP), ATP or CO has been reported (Curro et al., 2004). The aim of this study was to investigate the inﬂuence of H 2 S on contractility in mouse gastric fundus, and to explore its mechanism of action. The data obtained in this study indicate that in mouse gastric fundus, H 2 S induces relaxation at least partially via activation of myosin light chain phosphatase; this seems not related to an inhibition of the RhoA/Rho-kinase pathway. European Journal of Pharmacology 606 (2009) 180–186 ⁎ Corresponding author. Heymans Institute of Pharmacology, De Pintelaan 185, B-9000 Ghent, Belgium. Tel.: +32 9 332 33 73; fax: +32 9 332 49 88. E-mail address: Romain.Lefebvre@UGent.be (R.A. Lefebvre). 0014-2999/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.ejphar.2009.01.011 Contents lists available at ScienceDirect European Journal of Pharmacology journal homepage: www.elsevier.com/locate/ejphar