5-HT4 Receptor Ligand RS67333 Modulates Striatal Acetylcholine and Dopamine Release via the Inhibition of Acetylcholinesterase.

Overview

1. Centre for Cellular and Molecular Neuroscience, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT, United Kingdom
2. Oxford Parkinson’s Disease Centre, University of Oxford, Oxford OX1 3PT, United Kingdom
3. Aligning Science Across Parkinson’s (ASAP) Collaborative Research Network, Chevy Chase, Maryland 20815, United States

Abstract

Serotonin 5-HT4 receptors (5-HT4Rs) have emerged as potential therapeutic targets in neuropsychiatric and neurodegenerative disorders by modulating circuits that shape mood, cognition, and motor functions. Ligands for 5-HT4Rs can modify dopamine (DA) and acetylcholine (ACh) transmission, but mechanisms and circuits have not been fully resolved. Some 5-HT4R agonists have been suggested to have effects that include the inhibition of acetylcholinesterase (AChE), raising the speculation that 5-HT4R ligands might modulate ACh and DA through this action. Here, we investigated the impact of RS67333, a partial 5-HT4R agonist, on DA and ACh release dynamics in the striatum detected ex vivo in mouse brain slices using fast-scan cyclic voltammetry (FCV) and genetically encoded ACh sensor GRABACh3.0, respectively. We found that RS67333 significantly modulated electrically evoked DA release in the dorsolateral striatum (DLS) and nucleus accumbens core, effects that were abolished by a nicotinic receptor (nAChR) antagonist. In parallel, RS67333 altered evoked ACh signals by extending extracellular ACh lifetime, and correspondingly, RS67333 was found to inhibit striatal AChE enzymatic activity. By contrast, BIMU8, an alternative 5-HT4R ligand that did not inhibit striatal AChE, had no effect on the evoked striatal ACh or DA release. These findings indicate that RS67333 modulates striatal ACh transmission, which shapes downstream regulation of DA release by nAChRs, not through 5-HT4Rs but through AChE inhibition. These findings emphasize the caution due in attributing functions to 5-HT4Rs but also highlight an alternative pharmacological profile of some purported 5-HT4R ligands as AChE inhibitors of potential utility for treating ACh/DA disorders.

Reproduced under the paper's license (CC BY), from the paper cited above.

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Data

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• zenodo:17278002, at Zenodo; found in the availability statement

Data availability

The source data, as well as protocols, key lab materials, and code used and generated in this study are listed in a Key Resource Table and available alongside their persistent identifiers at https://doi.org/10.5281/zenodo.17278002.

Reproduced under the paper's license (CC BY), from the paper cited above.

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Version 3, 28 September 2026

• Publisher: n/a → American Chemical Society

Version 1, 27 September 2026: the first record

Recorded: type, language, journal, volume, issue, pages, dates, 3 authors, 10 keywords, 15 MeSH terms, 1 funder, 56 references.

Cite

This paper

Qiao, Q., Wu, W., & Cragg, S. J. (2026). 5-HT4 Receptor Ligand RS67333 Modulates Striatal Acetylcholine and Dopamine Release via the Inhibition of Acetylcholinesterase. ACS chemical neuroscience, 17(15), 3013-3022. https://doi.org/10.1021/acschemneuro.6c00342

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