Quantitative Spatial Analysis of Neuroligin-3 mRNA Expression in the Enteric Nervous System Reveals a Potential Role in Neuronal–Glial Synapses and Reduced Expression in Nlgn3R451C Mice

Mutations in the Neuroligin-3 (Nlgn3) gene are implicated autism spectrum disorder (ASD) and gastrointestinal (GI) dysfunction, but cellular Nlgn3 expression enteric nervous system remains to be characterised. We combined RNAScope situ hybridization immunofluorescence measure mRNA cholinergic VIP-expressing submucosal neurons, nitrergic calretinin-containing myenteric neurons glial cells both WT Nlgn3R451C mutant mice. measured neuronal via quantitative three-dimensional image analysis. To validate dual RNAScope/immunofluorescence data, we interrogated available single-cell RNA sequencing (scRNASeq) data assess for Nlgn3, Nlgn1, Nlgn2 their binding partners, Nrxn1-3, MGDA1 MGDA2, neural subsets. Most expressed mRNA. In contrast other Nlgns was strongly glia, suggesting a role neuroligin-3 mediating neuron–glia interactions. The autism-associated R451C mutation reduces not VIPergic neurons. plexus, levels reduced calretinin, nNOS-labelled S100 β -labelled glia. provide comprehensive profile ileal subpopulations of mice expressing which may contribute understanding pathophysiology GI dysfunction ASD.