GABAergic deficits and schizophrenia-like behaviors in a mouse model carrying patient-derived neuroligin-2 R215H mutation

Abstract

Schizophrenia (SCZ) is a chronic neuropsychiatric disorder caused by both genetic and environmental factors. It is featured by long-standing delusion and hallucination (psychosis), and cognitive deficits (Freedman, 2003; Insel, 2010; Lewis and Lieberman, 2000). SCZ is a highly heritable disorder (Sullivan et al., 2003) with a complex genetic basis. Recent genomic studies identified a number of genetic variants associated with . CC-BY 4.0 International license It is made available under a (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint . http://dx.doi.org/10.1101/225524 doi: bioRxiv preprint first posted online Nov. 27, 2017; 3 SCZ, including a group of variants resided in the genes encoding synaptic adhesion molecules that promoting synaptic function and development such as IGSF9B, and NLGN4X (Schizophrenia Working Group of the Psychiatric Genomics, 2014).Neuroligins (NLGNs) are a family of synaptic adhesion molecules highly expressed in the brain and are ligands for another group of cell adhesion molecules neurexins (NRXNs) (Ichtchenko et al., 1995). There are five neuroligin genes (neuroligin-1, -2, -3, -4, and -5) in humans and four in mice (neuroligin 1-4). Neuroligin-1, -2, and -3 are close homologs between human and mice. Neuroligin-1 and neuroligin-2 differentially locate to excitatory and inhibitory synapses and are critical for the excitatory and inhibitory synapse formation and function, respectively (Chubykin et al., 2007;Dong et al., 2007; Levinson et al., 2005; Nam and Chen, 2005;Scheiffele et al., 2000;Song et al., 1999;Varoqueaux et al., 2004). Neuroligin-3 locates at both type of synapses and contributes to both neurotransmission (Budreck and Scheiffele, 2007; Etherton et al., 2011). In recent years, several genetic variants of neuroligin-3 and neuroligin-4 have been identified in autism patients (Chih et al., 2004; Jamain et al., 2003). Mutations in proteins interacting with neuroligins such as Neurexin1, SHANK and MDGA have also been associated with autism and schizophrenia patients (Bucan et al., 2009; Durand et al., 2007; Kim et al., 2008; Kirov et al., 2008). Genetic mouse models based on these findings recapitulate some aspects of patient symptoms as well (Baudouin et al., 2012;Connor et al., 2016; Etherton et al., 2011; Etherton et al., 2009; Jamain et al., 2008;Peça et al., 2011;Rothwell et al., 2014;Südhof, 2008;Tabuchi et al., 2007; Zhou et al., 2016).. CC-BY 4.0 International license It is made available under a (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint . http://dx.doi.org/10.1101/225524 doi: bioRxiv preprint first posted online Nov. 27, 2017; 4We have previously reported several novel mutations of NLGN2 from schizophrenia patients (Sun et al., 2011). Among the NL2 mutants, we found that the R215H mutant protein was retained in the endoplasmic...

Neuroligin-2 as a central organizer of inhibitory synapses in health and disease

“…In line with the molecular and cellular consequences of the R215H mutation in NLGN2, both KI mouse lines show behavioral pheno types that partially recapitulate the constitutive KO mice, including, most notably, the prominent increase in anxiety behaviors in the OF and EPM (93,97,98). They also display a far wider range of cognitive impairments than have previously been reported for the constitutive KO mice, including impaired spatial learning and memory in the Morris water maze and the novel location recogni tion test but not the novel object recognition test (97), as well as impairments in spontaneous alternation, contextual fear condition ing (93), and cued fear conditioning (98). Last, alterations in pre pulse inhibition and exaggerated stress responses after restraint stress were also reported (93,97), supporting the notion that these models recapitulate several core phenotypes of schizophrenia.…”

“…Despite this residual expression, the Jiang et al line showed the expected impairment in inhibitory synapse function but not excitatory synapse function, with decreased levels of the postsynaptic GABArelated proteins gephyrin and GABA A R2 as well as reduced mIPSC frequency and amplitude in both the dentate gyrus of the hippocampus (93) and the mPFC (98). Notably and in discrepancy with the constitutive Nlgn2 KO mice, levels of the presynaptic markers PV and VIAAT were also found to be reduced in both brain regions (93,98), indicating that the resid ual NLGN2R215H mutant protein may confer a gain of function with respect to presynaptic development rather than mimicking loss of function, as has previously been reported for an autism related mutation in NLGN3 (11). Moreover, electrophysiological approaches revealed abnormal GABAergic network activity in the mPFC, with a specific reduction in the power of fast gamma oscilla tions at 60 to 100 Hz (98).…”

“…Among these, an R215H point mutation was predicted to have damaging effects, albeit with in complete clinical penetrance. In cellbased assays, the R215H muta tion was found to encode a lossoffunction variant due to a failure in protein maturation and glycosylation, resulting in deficient trans location to the cell membrane and subsequent impairments in GABAergic synaptogenesis (92,93). Consistent with this notion, a subsequent genomic study screening subjects with schizophrenia detected an overall increased level of rare mutations in mostly non coding areas of the NLGN2 gene, although none of these mutations was identified as directly increasing the risk for this condition (94).…”

“…Animal models of NLGN2 mutations discovered in human patients provide useful tools to further investigate the precise role of aber rant NLGN2 in neuropsychiatric symptoms. Accordingly, on the basis of the R215H point mutation identified in a patient with schizophrenia (92), two homozygous R215H knockin (KI) mouse lines were generated (93,97). As expected on the basis of evidence from cellbased assays (92), these mouse lines expressed little or no NLGN2 protein, effectively rendering them constitutive KOs, although minor residual expression of immature nonglycosylated protein could be detected in the line reported by Jiang et al (93,98) but not the other reported by Chen et al (97).…”

“…Accordingly, on the basis of the R215H point mutation identified in a patient with schizophrenia (92), two homozygous R215H knockin (KI) mouse lines were generated (93,97). As expected on the basis of evidence from cellbased assays (92), these mouse lines expressed little or no NLGN2 protein, effectively rendering them constitutive KOs, although minor residual expression of immature nonglycosylated protein could be detected in the line reported by Jiang et al (93,98) but not the other reported by Chen et al (97). Despite this residual expression, the Jiang et al line showed the expected impairment in inhibitory synapse function but not excitatory synapse function, with decreased levels of the postsynaptic GABArelated proteins gephyrin and GABA A R2 as well as reduced mIPSC frequency and amplitude in both the dentate gyrus of the hippocampus (93) and the mPFC (98).…”