928 Modulation By Muscarinic Acetylcholine Receptor Subtype 4 of Carbachol- Induced Pepsin Secretion in Mice: Relation to D Cells/Somatostatin

Abstract

Background/Aim: Muscarinic acetylcholine receptors (mAChRs) consist of five subtypes (M1~M5) and are widely expressed to mediate diverse autonomic functions in peripheral organs, including the gastrointestinal tract. Although both M1and M3receptors reportedly play a role in the regulation of pepsin secretion in response to acetylcholine, we recently found that carbachol (CCh)-induced pepsin secretion wasmarkedly decreased inM4 knockout (KO) mice. In the present study, we demonstrated, using M1~M5 KO mice, the importance of M4 receptors in the cholinergic regulation of pepsin secretion and investigated how this secretion is modulated by the activation of M4 receptors. Methods: C57BL/6J mice of wildtype (WT) and M1-, M2-, M3-, M4or M5-KO were used. Under urethane anesthesia, the abdomen was incised, the cardiac portion was ligated, and an acute fistula prepared with a polyethylene tube was provided in the stomach through a pylorus. Then, the stomach was instilled with saline (0.4 ml) through the fistula, and the solution was changed every 20 min. CCh (30 μg/kg) was given SC as a single injection. Atropine (0.3 mg/kg) or CYN154806 [somatostatin-2 receptor (SST2R) antagonist 0.1-3mg/kg] was given SC 20 min before CCh. Expressions of D cells and M4 receptors were examined immunohistochemically by double staining with anti-somatostatin and anti-M4 receptor antibodies. Results: CCh caused an increase of pepsin secretion in WT mice, and the effect was completely inhibited by prior administration of atropine. The stimulatory effect of CCh was similarly observed in the animals lacking M1-, M2or M5-receptors but significantly decreased in M3or M4KO mice, as compared to WT; especially, the response was all but completely abolished in M4-KOmice. CYN154806, the SST2R antagonist, significantly reversed the decreased pepsin response to CCh in M4but not M3-KO mice. The M4-KO mice showed basal pepsin secretion much less than WT mice, and these changes were also reversed by CYN154806. By contrast, somatostatin decreased pepsin secretion under basal and CCh-stimulated conditions. Furthermore, the immunohistochemical study showed the localization ofM4 receptors on D cells in the mouse stomach. Conclusion: These results suggest that under cholinergic stimulation the secretion of pepsin is mediated mainly by the activation of M4-receptors and partly through M3-receptors but does not involve other muscarinic receptor subtypes. Somatostatin has an inhibitory effect on pepsin secretion through SST2 receptors. It is assumed that the activation of M4-receptors inhibits the release of somatostatin from D cells and results in enhancement of pepsin response.