Impaired cholinergic modulation of striatal spiny projection neurons in repetitive behaviors associated with TSHZ3-linked autism spectrum disorder

Alterations in central cholinergic systems are implicated in autism spectrum disorder (ASD). We previously identified TSHZ3 as an ASD-linked gene and provided evidence, from transgenic mouse models, supporting the view that striatal cholinergic interneurons (SCINs) are key players in the core ASD trait represented by repetitive behaviors (RBs). Indeed, conditional deletion of Tshz3 in cholinergic neurons (Chat-cKO model) results in RBs associated with reduced SCIN firing activity and decreased striatal cholinergic tone. Here we tackled the resulting adaptations in the striatal circuitry underlying RBs in Chat-cKO, focusing on the two populations of striatal projection neurons (SPNs) that express dopamine D1 (D1R) or D2 (D2R) receptor. Using patch-clamp electrophysiological recordings, as well as behavioral, pharmacological and molecular approaches, we show reduced cholinergic transmission through the muscarinic M1 receptor onto SPNs, with no changes in the functionality or expression of this receptor. Notably, we demonstrate a loss of the differential excitability of D1R- vs. D2R-SPNs. Finally, acute in vivo treatment with the acetylcholinesterase inhibitor donepezil significantly alleviates RBs in mutant mice. These findings point to dysfunction of SCINs as a pathological and druggable mechanism underlying RBs in Tshz3-linked ASD, and to altered SPN excitability as a critical contributor in the resulting pathophysiological functioning of the striatum.