Characterization of Selective M₅ Acetylcholine Muscarinic Receptor Modulators on Dopamine Signaling in the Striatum

The type-5 muscarinic acetylcholine receptor (mAChR, M₅) is almost exclusively expressed in dopamine (DA) neurons of the ventral tegmental area and substantia nigra pars compacta; therefore, they are ideally located to modulate DA signaling and underlying behaviors. However, the role of M₅ in shaping DA release is still poorly characterized. In this study, we first quantitatively mapped the expression of M₅ in different neurons of the mouse midbrain, then used voltammetry in mouse striatum to evaluate the effect of M₅-selective modulators on DA release. The M₅ negative allosteric modulator ML375 significantly decreased electrically evoked DA release and blocked the effect of Oxotremorine-M (Oxo-M; nonselective mAChR agonist) on DA release in the presence of an acetylcholine nicotinic receptor blocker. Conversely, the M₅ positive allosteric modulator VU 0365114 significantly increased electrically evoked DA release and the Oxo-M effect on DA release. We then assessed M₅'s impact on mesolimbic circuit function in vivo. Although psychostimulant-induced locomotor activity models in knockout mice have previously been used to characterize the role of M₅ in DA transmission, the results of these studies conflict, leading us to select a different in vivo model, namely a cocaine self-administration paradigm. In contrast to a previous study that also used this model, in the current study, administration of ML375 did not decrease cocaine self-administration in rats (using fixed and progressive ratio). These conflicting results illustrate the complexity of M₅ modulation and the need to further characterize its involvement in the regulation of dopamine signaling, central to multiple neuropsychiatric diseases. SIGNIFICANCE STATEMENT: This work describes the type-5 muscarinic receptor (M₅) pattern of expression within the midbrain as well as its physiological modulation by selective compounds at the axon terminal level in the striatum, where M₅ directly shapes dopamine transmission. It offers the first direct readout of mesolimbic dopamine release modulation by M₅, highlighting its role in regulating neurocircuits implicated in the pathophysiology of neuropsychiatric disorders such as substance use disorders, major depressive disorder, and schizophrenia.