Dysfunction of NMDA receptors in neuronal models of an autism spectrum disorder patient with a DSCAM mutation and in Dscam-knockout mice

Chae Seok Lim ,Ro Un Lee,Ja Eun Choi,Jiah Lee,Min Jung Kim,Chul Hoon Kim,Jin A Lee ,Jung Eun Yang ,Ariful Islam ,Jae Hyung Lee ,Yinyi Xiong,Kyung-Min Lee ,Seung-Hyun Hwang ,Pojeong Park,Hyunhyo Seo,Kyuseok Shim ,Ji Hye Kwak ,Yong Seok Lee ,You Kyung Lee ,Bong Kiun Kaang

doi:10.1038/s41380-021-01216-9

Abstract

Heterogeneity in the etiopathology of autism spectrum disorders (ASD) limits the development of generic remedies, requires individualistic and patient-specific research. Recent progress in human-induced pluripotent stem cell (iPSC) technology provides a novel platform for modeling ASDs for studying complex neuronal phenotypes. In this study, we generated telencephalic induced neuronal (iN) cells from iPSCs derived from an ASD patient with a heterozygous point mutation in the DSCAM gene. The mRNA of DSCAM and the density of DSCAM in dendrites were significantly decreased in ASD compared to control iN cells. RNA sequencing analysis revealed that several synaptic function-related genes including NMDA receptor subunits were downregulated in ASD iN cells. Moreover, NMDA receptor (R)-mediated currents were significantly reduced in ASD compared to control iN cells. Normal NMDA-R-mediated current levels were rescued by expressing wild-type DSCAM in ASD iN cells, and reduced currents were observed by truncated DSCAM expression in control iN cells. shRNA-mediated DSCAM knockdown in control iN cells resulted in the downregulation of an NMDA-R subunit, which was rescued by the overexpression of shRNA-resistant DSCAM. Furthermore, DSCAM was co-localized with NMDA-R components in the dendritic spines of iN cells whereas their co-localizations were significantly reduced in ASD iN cells. Levels of phospho-ERK1/2 were significantly lower in ASD iN cells, suggesting a potential mechanism. A neural stem cell-specific Dscam heterozygous knockout mouse model, showing deficits in social interaction and social memory with reduced NMDA-R currents. These data suggest that DSCAM mutation causes pathological symptoms of ASD by dysregulating NMDA-R function.

Highlights

Autism spectrum disorder (ASD) refers to a group of complex neurodevelopmental disorders characterized by persistent difficulties with social communication and social interaction, restricted and repetitive behavior, activities, or interests, and early childhood manifestation of symptoms [1] and poses a serious threat to a patient’s ability to harmoniously adapt to society
Among the synaptic proteins associated with ASD, N-methyl-Daspartate receptor (NMDA) receptors (Rs) have garnered attention because many ASD models with mutations in distinct genes show NMDA-R dysfunctions [10,11,12,13] and NMDA-Rs are important in neuronal differentiation and synaptic plasticity in several brain regions including the hippocampus and cortex [14, 15]
To examine cellular phenotypes caused by the Down syndrome cell adhesion molecule (DSCAM) mutation, NMDA-R subunits and NMDA-R dysfunctions have been impliforebrain-like induced neuronal (iN) cells were generated from the above induced pluripotent stem cell (iPSC) using cated in the pathogenesis of ASD, we investigated whether previously described protocols [28] with minor modifications NMDA-R-mediated currents are altered in ASD iN cells

Summary

INTRODUCTION

Autism spectrum disorder (ASD) refers to a group of complex neurodevelopmental disorders characterized by persistent difficulties with social communication and social interaction, restricted and repetitive behavior (including sensory behaviors), activities, or interests, and early childhood manifestation of symptoms [1] and poses a serious threat to a patient’s ability to harmoniously adapt to society. To analyze gene expression changes between control and ASD iPSCs or iN cells, quantitative RT-PCR was performed using specific primers, which are presented in Supplementary Table 3. A mouse line with a spontaneous mutation in Dscam (Dscamdel17) showed gross morphological changes in brain size and shape [22], as well as a decrease in the thickness of cortical layers and altered dendritic morphology in pyramidal neurons [22]. These mice exhibit severe hydrocephalus, decreased motor function, and impaired motor learning ability [26]. PLA reactions were followed by immunocytochemistry with the addition of primary antibodies (PSD-95, NeuroMab, 75-028) and fluorophore-

MATERIALS AND METHODS

RESULTS

DISCUSSION