Protective role of M3 muscarinic acetylcholine receptor in indomethacin-induced small intestinal injury.

EP₄ prostanoid receptor (EP₄R) contributes to the intestinal epithelial Cl^- secretion, and inhibition of prostaglandin E (PGE) production by non-steroidal anti-inflammatory drugs (NSAIDs) plays a central role in NSAID-induced enteropathy. Although M₃ muscarinic acetylcholine receptor (M₃R) also contributes to the intestinal epithelial Cl^- secretion, it remains unclear whether M₃R is involved in NSAID-induced enteropathy due to a lack of selective agents. The present study explored how M₃R is involved in the regulation of the intestinal epithelial Cl^- secretion and its pathophysiological role in NSAID-induced enteropathy. Using the novel highly-selective M₃ positive allosteric modulator PAM-369 that we recently developed, we evaluated the role of M₃R in the intestinal epithelial secretion ex vivo by measuring the short circuit current (Isc) of intestinal epithelium with a Ussing chamber system and examined whether or not M₃R protects against small intestinal injury in indomethacin-treated mice. Both the PGE₁ derivative misoprostol and carbachol similarly increased the Isc in a concentration-dependent manner. The Isc increases were abolished either by receptor antagonists (an EP₄R antagonist and a M₃R antagonist, respectively) or by removal of extracellular Cl^-. PAM-369 enhanced the carbachol-induced Isc by potentiating M₃R, which could contribute to enhanced intestinal epithelial secretion. Treatment with PAM-369 ameliorated small intestinal injury in indomethacin-treated mice. Importantly, the M₃R expression was significantly up-regulated, and PAM-369 potentiation of M₃R was augmented in indomethacin-treated mice compared to untreated mice. These findings show that M₃R plays a role in maintaining the intestinal epithelial secretion, which could contribute to protection against indomethacin-induced small intestinal injury. M₃R is a promising target for treating or preventing NSAID-induced enteropathy. KEY MESSAGES: PAM-369, the M₃ positive allosteric modulator, was used to potentiate M₃R. PAM-369 enhanced carbachol-induced Isc in mouse ileum. PAM-369 ameliorated small intestinal injury in indomethacin-treated mice. M₃R is a promising target for treating or preventing NSAID-induced enteropathy.