Nitroxyl (HNO) as novel player in the regulation of gastrointestinal motility

一类新兴的胃肠功能调节剂是气体分子，即所谓的气体递质。在肠道中有两种公认的具有传递功能的气体：一氧化氮（NO）和硫化氢（H2S）。两者通过不同的机制诱导胃肠平滑肌松弛。NO激活可溶性鸟苷酸环化酶，产生cGMP，从而激活蛋白激酶G（或可能直接激活K+通道），H2S激活例如atp敏感和小电导Ca2+依赖性K+通道。第三种气体递质硝基（nitroxyl， HNO）最近受到越来越多的关注，本研究将研究其对胃肠运动的作用。关于HNO在胃肠道中的作用，我们所知道的是它发挥一种促分泌作用，这种作用强烈依赖于Ca2+，涉及Ca2+依赖性和atp敏感的K+通道，部分由环加氧酶代谢物介导（Pouokam et al. 2013）。初步实验表明，HNO供体安琪莉斯盐能唤起大鼠回肠和结肠纵向条带测量的肠道松弛。本研究将通过对分离的肌细胞进行膜片钳实验、对自发收缩的平滑肌细胞重新聚集进行成像实验和等长肌肉收缩测量来研究这种松弛的机制。将特别关注所涉及的离子电导的表征，巯基修饰可能的中介（由于HNO的高硫代性）以及cGMP信号通路的典型成分的参与。同时，初步实验显示，肌肠神经元的超极化伴随着电压依赖性Na+通道携带的内向电流的抑制。HNO诱导肌肠神经元（即肠神经系统中负责自主控制肠道运动的部分）膜电位变化的离子机制尚不清楚，本研究将对其进行研究。最近的文献研究表明NO、H2S和HNO信号通路之间存在串扰；例如，外源性H2S可能与内源性NO相互作用形成HNO，这就增加了H2S的某些作用确实是由HNO介导的可能性。这一假设将通过测试H2S松弛反应对hno刺激通路抑制剂的敏感性来验证。通过这些实验，希望能够更好地了解新兴气体递质的新成员对胃肠运动的调节。

An emerging class of regulators of gastrointestinal functions are gaseous molecules, the so-called gasotransmitters. There are two well-established gases with transmitter function in the gut: nitric oxide (NO) and hydrogen sulphide (H2S). Both induce gastrointestinal smooth muscle relaxation via different mechanisms. While NO activates soluble guanylate cyclase, the production of cGMP and consequently activates protein kinase G (or potentially directly activates K+ channels), H2S activates e.g. ATP-sensitive and small conductance Ca2+-dependent K+ channels.A third gasotransmitter, nitroxyl (HNO), whose actions on gastrointestinal motility shall be investigated in the present study, is getting more attention recently. All what is known about HNO regarding the gastrointestinal tract is that it exerts a prosecretory action, which is strongly dependent on Ca2+ and involves Ca2+-dependent and ATP-sensitive K+ channels and is in part mediated by cyclooxygenase metabolites (Pouokam et al. 2013). Preliminary experiments revealed that an HNO donor, Angelis salt, evokes intestinal relaxation measured at longitudinal strips from rat ileum and colon. The mechanisms underlying this relaxation shall be investigated in the present study by patch-clamp experiments at isolated myocytes, imaging experiments at spontaneously contracting smooth muscle cell reaggregates and isometric muscle contraction measurements. Special focus will be placed on the characterization of the involved ion conductances, the possible mediation by thiol group modification (due to the high thiophylic character of HNO) and the involvement of typical components of the cGMP signalling pathway. In parallel, preliminary experiments revealed a hyperpolarization of myenteric neurons concomitant with an inhibition of inward-currents carried by voltage-dependent Na+ channels. The ionic mechanisms underlying the change in membrane potential induced by HNO of myenteric neurons, i.e. the part of the enteric nervous system responsible for autonomous control of gut motility, are unknown and shall be identified in the present study.Recent studies in the literature showed a cross-talk between NO, H2S and HNO signalling pathways; for example exogenous H2S may interact with endogenous NO to form HNO, which rises the possibility that some actions of H2S may indeed be mediated by HNO. This hypothesis shall be investigated by testing the sensitivity of the relaxant response to H2S against identified inhibitors of the HNO-stimulated pathway. With these experiments it is hoped to get a better insight into the regulation of gastrointestinal motility by a new member of the emerging class of gasotransmitters.