Probes for CHRM1

Trihexyphenidyl (THP), a non-selective muscarinic receptor (mAChR) antagonist, is commonly used for the treatment of dystonia associated with TOR1A, otherwise known as DYT1 dystonia. A better understanding of the mechanism of action of THP is a critical step in the development of better therapeutics with fewer side effects. We previously found that THP normalizes the deficit in striatal dopamine (DA) release in a mouse model of TOR1A dystonia (Tor1a+/ΔE knockin (KI) mice), revealing a plausible mechanism of action for this compound, considering that abnormal DA neurotransmission is consistently associated with many forms of dystonia. However, the mAChR subtype(s) that mediate the rescue of striatal dopamine release remain unclear. In this study we used a combination of pharmacological challenges and cell-type specific mAChR conditional knockout mice of either sex to determine which mAChR subtypes mediate the DA release-enhancing effects of THP. We determined that THP acts in part at M4 mAChR on striatal cholinergic interneurons to enhance DA release in both Tor1a+/+ and Tor1a+/ΔE KI mice. Further, we found that the subtype selective M4 antagonist VU6021625 recapitulates the effects of THP. These data implicate a principal role for M4 mAChR located on striatal cholinergic interneurons in the mechanism of action of THP and suggest that subtype selective M4 mAChR antagonists may be effective therapeutics with fewer side effects than THP for the treatment of TOR1A dystonia.

Intestinal goblet cells maintain the protective epithelial barrier through mucus secretion and yet sample lumenal substances for immune processing through formation of goblet cell associated antigen passages (GAPs). The cellular biology of GAPs and how these divergent processes are balanced and regulated by goblet cells remains unknown. Using high-resolution light and electron microscopy, we found that in mice, GAPs were formed by an acetylcholine (ACh)-dependent endocytic event remarkable for delivery of fluid-phase cargo retrograde into the trans-golgi network and across the cell by transcytosis - in addition to the expected transport of fluid-phase cargo by endosomes to multi-vesicular bodies and lysosomes. While ACh also induced goblet cells to secrete mucins, ACh-induced GAP formation and mucin secretion were functionally independent and mediated by different receptors and signaling pathways, enabling goblet cells to differentially regulate these processes to accommodate the dynamically changing demands of the mucosal environment for barrier maintenance and sampling of lumenal substances.

BACKGROUND: Muscarinic receptor 1 positive allosteric modulators (M1PAMs) enhance colonic propulsive contractions and defecation through the facilitation of M1 receptor (M1R)-mediated signaling. We examined M1R expression in the colons of 5 species and compared colonic propulsion and defecation caused by the M1PAM, T440, the 5-HT4 agonist, prucalopride, and the cholinesterase inhibitor, neostigmine, in rats and dogs. METHODS: M1R expression was profiled by immunostaining and in situ hybridization. In vivo studies utilized male SD rats and beagle dogs. Colonic propulsive contractions were recorded by manometry in anesthetized rats. Gut contractions in dogs were assessed using implanted force transducers in the ileum, proximal, mid, and distal colons. KEY RESULTS: M1R was localized to neurons of myenteric and submucosal plexuses and the epithelium of the human colon. A similar receptor localization was observed in rat, dog, mouse, and pig. T440 enhanced normal defecation in rats in a dose-dependent manner. Prucalopride also enhanced defecation in rats, but the maximum effect was half that of T440. Neostigmine and T440 were similarly effective in enhancing defecation, but the effective dose of neostigmine was close to its lethal dose. In rats, all 3 compounds induced colonic contractions, but the associated propulsion was strongest with T440. In dogs, intestinal contractions elicited by T440 propagated from ileum to distal colon. Prucalopride and neostigmine also induced intestinal contractions, but these were less well coordinated. No loss of effectiveness of T440 on defecation occurred after 5 days of repeated dosing. CONCLUSION AND INFERENCES: These results suggest that M1PAMs produce highly coordinated propagating contraction by actions on the enteric nervous system of the colon. The localization of M1R to enteric neurons in both animals and humans suggests that the M1PAM effects would be translatable to human. M1PAMs provide a potential novel therapeutic option for constipation disorders