Camk2a-Cre and Tshz3 Expression in Mouse Striatal Cholinergic Interneurons: Implications for Autism Spectrum Disorder.

Xavier Caubit,Florence Desprez,Ahmed Fatmi,Bianca Habermann,Paolo Gubellini,Laurent Fasano,Elise Arbeille,Imane Messak,Dorian Chabbert

doi:10.3389/fgene.2021.683959

Abstract

Camk2a-Cre mice have been widely used to study the postnatal function of several genes in forebrain projection neurons, including cortical projection neurons (CPNs) and striatal medium-sized spiny neurons (MSNs). We linked heterozygous deletion of TSHZ3/Tshz3 gene to autism spectrum disorder (ASD) and used Camk2a-Cre mice to investigate the postnatal function of Tshz3, which is expressed by CPNs but not MSNs. Recently, single-cell transcriptomics of the adult mouse striatum revealed the expression of Camk2a in interneurons and showed Tshz3 expression in striatal cholinergic interneurons (SCINs), which are attracting increasing interest in the field of ASD. These data and the phenotypic similarity between the mice with Tshz3 haploinsufficiency and Camk2a-Cre-dependent conditional deletion of Tshz3 (Camk2a-cKO) prompted us to better characterize the expression of Tshz3 and the activity of Camk2a-Cre transgene in the striatum. Here, we show that the great majority of Tshz3-expressing cells are SCINs and that all SCINs express Tshz3. Using lineage tracing, we demonstrate that the Camk2a-Cre transgene is expressed in the SCIN lineage where it can efficiently elicit the deletion of the Tshz3-floxed allele. Moreover, transcriptomic and bioinformatic analysis in Camk2a-cKO mice showed dysregulated striatal expression of a number of genes, including genes whose human orthologues are associated with ASD and synaptic signaling. These findings identifying the expression of the Camk2a-Cre transgene in SCINs lineage lead to a reappraisal of the interpretation of experiments using Camk2a-Cre-dependent gene manipulations. They are also useful to decipher the cellular and molecular substrates of the ASD-related behavioral abnormalities observed in Tshz3 mouse models.

Highlights

The corticostriatal (CStr) circuitry is critically involved in functions ranging from motor control and habit formation to cognition
This study show that (i) all striatal cholinergic interneurons (SCINs) express TSHZ3 and the main striatal population expressing TSHZ3 are SCINs; (ii) the Camk2a-Cre transgene is expressed in the SCIN lineage; (iii) conditional deletion of Tshz3 using the Camk2a-Cre mediates efficient Tshz3 deletion in SCINs and drives changes in the expression of 725 genes in the striatum, among which 196 are differentially expressed in the cerebral cortex, suggesting profound alteration of striatal function
These results call to more caution in the interpretation of experiments using Camk2a-Credependent gene manipulations and point to SCINs as potential players in the autism spectrum disorder (ASD)-relevant behavioral abnormalities triggered by Tshz3 loss in both the heterozygous deletion model (Caubit et al, 2016) and the conditional model using the Camk2a-Cre transgene (Chabbert et al, 2019)

Summary

Introduction

The corticostriatal (CStr) circuitry is critically involved in functions ranging from motor control and habit formation to cognition. IT and PT neurons that project to the striatum reside mostly in the deep cortical layer (L) 5 (Reiner et al, 2010; Shepherd, 2013). There is growing evidence for a higher diversity of striatal interneurons, based on studies on their developmental origin, gene expression profile, and electrophysiological properties and connectivity. These data define at least 6–7 classes of GABAergic interneurons (Munoz-Manchado et al, 2018; Tepper et al, 2018) and reveal that SCINs are a heterogeneous population (Ahmed et al, 2019). Striatal interneurons, and SCINs in particular, play a key functional role by modulating striatal activity through distinct connections with MSNs and/or other interneurons (Tepper et al, 2018; Abudukeyoumu et al, 2019)

Methods

Results

Conclusion