Gastroprotective action of orexin-A against stress-induced gastric damage is mediated by endogenous prostaglandins, sensory afferent neuropeptides and nitric oxide

Abstract

Orexin-A, identified in the neurons and endocrine cells in the gut, has been implicated in control of food intake and sleep behavior but little is known about its influence on gastric secretion and mucosal integrity. The effects of orexin-A on gastric secretion and gastric lesions induced in rats by 3.5 h of water immersion and restraint stress (WRS) or 75% ethanol were determined. Orexin-A (5–80 μg/kg i.p.) increased gastric acid secretion and attenuated gastric lesions induced by WRS and this was accompanied by the significant rise in plasma orexin-A, CGRP and gastrin levels, the gastric mucosal blood flow (GBF), luminal NO concentration and an increase in mRNA for CGRP and overexpression of COX-2 protein and the generation of PGE₂ in the gastric mucosa. Orexin-A-induced protection was abolished by selective OX-1 receptor antagonist, vagotomy and attenuated by suppression of COX-1 and COX-2, deactivation of afferent nerves with neurotoxic dose of capsaicin, pretreatment with CCK₂/gastrin antagonist, CGRP_8–37 or capsazepine and by inhibition of NOS with l-NNA. This study shows for the first time that orexin-A exerts a potent protective action on the stomach of rats exposed to non-topical ulcerogens such as WRS or topical noxious agents such as ethanol and these effects depend upon hyperemia mediated by COX-PG and NOS-NO systems, activation of vagal nerves and sensory neuropeptides such as CGRP released from sensory nerves probably triggered by an increase in gastric acid secretion induced by this peptide.

Introduction

The orexins also called hypocretins are novel neuropeptides originally discovered in a small group of neurons in the lateral hypothalamic area (LHA) [1], [2]. The orexin peptides were originally identified as regulators of food intake and of sleep behavior and act at one or the other of two G protein-coupled receptors, the orexin-1 (OX-R1) and orexin-2 (OX-R2) receptors. The OX-1 receptor has a greater affinity for orexin-A over orexin-B, whereas the OX-2 receptor exhibits similar affinity for both ligands [3]. Intracerebroventricular administration or direct injection of orexins into the LHA has been shown to increase food intake in rodents [4]. Although the major function of the orexin system was the participation in the regulation of appetite behavior at the level of the hypothalamus, it recently became evident that the orexins are also produced in the neurons and endocrine cells in the gut [5], [6]. Besides the presence in the brain, orexin-like immunoreactivity was also found in the neurons of enteric nervous system (ENS) of different species including guinea pig, rat, mouse and humans. Moreover, the prepro-orexin, orexin A and orexin receptor mRNA expression were demonstrated in the myenteric plexus and afferent nerves colocalized with VIP-ergic nerves and in the gastrin-producing cells of the rat stomach [5]. Recent studies in humans revealed that the plasma orexin-A concentrations increase in fasting humans [7].

Previous studies revealed that the activation of sensory nerves in the upper gastrointestinal tract may influence the secretory functions in the stomach and play an important role in the mechanism of gastric mucosal integrity and gastric cytoprotection by increasing gastric mucosal blood flow via the release of CGRP from sensory afferent nerves [8], [9]. The presence of OX-R1 receptors in the vagal afferent nerves in the rat and humans has been recently confirmed [10], suggesting that binding of orexin-A, predominantly with OX-R1 receptors, might result in local release of sensory neuropeptides such as CGRP from the sensory afferent nerve endings. Moreover, Flemstrom et al. [11] revealed that orexin-A is a potent stimulant of duodenal bicarbonate secretion considered as a major line of defense against acid-induced mucosal injury. They demonstrated that orexin A caused a robust stimulation of duodenal HCO₃⁻ secretion in fed animals but did not affect this secretion in fasted animals while the response to VIP or melatonin in their study, were little affected by overnight fasting [11]. In another studies, fasting and glucopenia were considered as a potent stimulants for the orexin-A production and expression of OX-R1 in LHA [7], [10]. Studies in rats revealed that exogenous orexin-A stimulates gastric acid secretion [12] but the contribution of this peptide to the mechanism of gastric mucosal defense and gastroprotection against the damage induced by stress or topical necrotizng irritants such as ethanol, has not been extensively investigated.

This study was designed to compare the effects of intraperitoneal (i.p.) and intracerebroventricular (i.c.v.) administration of orexin-A on gastric acid secretion in rats equipped with chronic gastric fistula (GF) and gastric lesions induced by 3, 5 h of water immersion and restraint stress (WRS). For comparison, the effects of orexin-A on acute gastric lesions induced by intragastric application of 75% ethanol and the accompanying changes in the gastric blood flow (GBF) were assessed. An attempt was made to examine the involvement of OX-R1 and CCK₂/gastrin receptors, prostaglandins (PG), nitric oxide (NO), as well as vagal and sensory nerves releasing CGRP in orexin-A-induced gastroprotection against gastric mucosal damage induced by WRS.