Abstract
BackgroundOf the many genetic mutations known to increase the risk of autism spectrum disorder, a large proportion cluster upon synaptic proteins. One such family of presynaptic proteins are the neurexins (NRXN), and recent genetic and mouse evidence has suggested a causative role for NRXN2 in generating altered social behaviours. Autism has been conceptualised as a disorder of atypical connectivity, yet how single-gene mutations affect such connectivity remains under-explored. To attempt to address this, we have developed a quantitative analysis of microstructure and structural connectivity leveraging diffusion tensor MRI (DTI) with high-resolution 3D imaging in optically cleared (CLARITY) brain tissue in the same mouse, applied here to the Nrxn2α knockout (KO) model.MethodsFixed brains of Nrxn2α KO mice underwent DTI using 9.4 T MRI, and diffusion properties of socially relevant brain regions were quantified. The same tissue was then subjected to CLARITY to immunolabel axons and cell bodies, which were also quantified.ResultsDTI revealed increases in fractional anisotropy in the amygdala (including the basolateral nuclei), the anterior cingulate cortex, the orbitofrontal cortex and the hippocampus. Axial diffusivity of the anterior cingulate cortex and orbitofrontal cortex was significantly increased in Nrxn2α KO mice, as were tracts between the amygdala and the orbitofrontal cortex. Using CLARITY, we find significantly altered axonal orientation in the amygdala, orbitofrontal cortex and the anterior cingulate cortex, which was unrelated to cell density.ConclusionsOur findings demonstrate that deleting a single neurexin gene (Nrxn2α) induces atypical structural connectivity within socially relevant brain regions. More generally, our combined within-subject DTI and CLARITY approach presents a new, more sensitive method of revealing hitherto undetectable differences in the autistic brain.
Highlights
Of the many genetic mutations known to increase the risk of autism spectrum disorder, a large proportion cluster upon synaptic proteins
Cell density measurements are unlikely to reveal the true extent of changes within the autistic brain. We have addressed this by developing a dual imaging approach (DTI and Optically cleared brain tissue (CLARITY)) that quantifies the alignment and density of white matter, applied here to brain regions known to support social behaviour in a mouse model of autism
Given the social impairments found within Nrxn2α mice, for the current study, we identified the brain regions of interest (ROIs) most closely linked with social behaviour, using previously published reports of brain region involvement in social behaviour
Summary
Of the many genetic mutations known to increase the risk of autism spectrum disorder, a large proportion cluster upon synaptic proteins. One such family of presynaptic proteins are the neurexins (NRXN), and recent genetic and mouse evidence has suggested a causative role for NRXN2 in generating altered social behaviours. It is clear that autism is highly polygenic. Around ~ 400–1000 genes are involved in autism susceptibility [2,3,4,5]. Many of these genes cluster upon proteins relating to synaptic signalling [6]. NRXNs are encoded by three genes (NRXN1, NRXN2, NRXN3; note that CNTNAP1 and CNTNAP2 are sometimes referred to as NRXN4), of which two major isoforms exist: the longer α proteins with six laminin/neurexin/sex hormone (LNS) binding domains, and the shorter β proteins with one LNS binding domain [7, 8]
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