The GPCR Smoothened on Cholinergic Interneurons Modulates Dopamine-associated Acetylcholine Dynamics and Affects Learning

The striatum is a hub for associative learning where fluctuations in dopamine and acetylcholine dynamically regulate behavior. Acetylcholine is released by cholinergic interneurons, which integrate diverse inputs to contextualize dopamine signals and shape behavior. We previously observed that the GPCR Smoothened on cholinergic interneurons suppresses L-DOPA-induced dyskinesias, a motor side-effect resulting from medication elevated dopamine in the Parkinsonian brain. Here, we examine whether Smoothened signaling modulates acetylcholine dynamics, its coordination with dopamine, and motor learning in the healthy brain. We find that cholinergic neuron-specific Smoothened activity bidirectionally modulates acetylcholine inhibition following dopaminergic or cholinergic neuron activity. These effects alter the temporal organization of acetylcholine in the dorsolateral striatum and its coupling to dopamine. Behaviorally, Smoothened ablation from cholinergic neurons promotes motor learning and altered adjustment to changes in the effort or time to obtain reward. These findings identify Smoothened as a bidirectional modulator of striatal dopamine-acetylcholine coordination and striatal learning. Highlights Sonic Hedgehog–Smoothened signaling in striatal cholinergic interneurons bidirectionally regulates dopamine-associated cholinergic pauses without altering dopamine release. Cholinergic Smoothened shapes the timing, duration, and coordination of endogenous dopamine–acetylcholine dynamics in the dorsolateral striatum. Cholinergic Smoothened modulates striatal learning by accelerating motor learning while constraining effort management in an instrumental task.