Aberrant expression of S-SCAM causes the loss of GABAergic synapses in hippocampal neurons

Samantha Skaar,Sang H Lee,Eric Danielson,Seung Min Shin

doi:10.1038/s41598-019-57053-y

Samantha Skaar, Sang H Lee + Show 2 more

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https://doi.org/10.1038/s41598-019-57053-y

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Journal: Scientific Reports	Publication Date: Jan 9, 2020
Citations: 11	License type: open-access

Affiliation: Medical College of Wisconsin

Abstract

The duplication and deletion mutations of the S-SCAM/MAGI-2 gene are associated with schizophrenia and infantile spasms, respectively. S-SCAM is a unique synaptic scaffolding protein that localizes to both excitatory and GABAergic synapses. However, consequences of aberrant S-SCAM expression on GABAergic synapses is little studied. Here we report the effect of S-SCAM knockdown and overexpression on GABAergic synapses. S-SCAM knockdown in cultured hippocampal neurons caused a drastic loss of both pre- and post-synaptic components of GABAergic synapses, indicating its essential role in GABAergic synapse formation and maintenance. Surprisingly, S-SCAM overexpression also attenuated GABAergic synapses, but the effect is mediated by the loss of postsynaptic GABAA receptors, gephyrin, and neuroligin 2 and does not involve presynaptic component vesicular GABA transporters. Overexpression studies using S-SCAM mutants with various domain deletions indicated that GABAergic synapse loss correlates with their ability to increase excitatory synaptic function. Consistently, AMPA receptor antagonist CNQX or calcineurin inhibitor FK506 abolished the S-SCAM overexpression-induced loss of GABAA receptors, supporting that GABAergic synapse loss by S-SCAM overexpression is due to the activity-induced dispersal of synaptic GABAA receptors. These results suggest that abnormal S-SCAM protein levels disrupt excitation/inhibition balance in neurons, which may explain the pathogenic nature of S-SCAM copy number variations.

Highlights

Maintaining proper balance in excitation and inhibition (E/I) is critical for circuit functionality and brain function[1]
Synaptic scaffolding molecule (S-SCAM) knockdown causes the loss of GABAergic synapses in cultured hippocampal neurons
S-SCAM RNAi reduced the number of co-localized GABAAR γ2 and vesicular GABA transporter (vGAT) puncta that represent GABAergic synapses (17.3 ± 0.9 vs 6.6 ± 0.7 per 100 μm; 38% of control)

Summary

Introduction

Maintaining proper balance in excitation and inhibition (E/I) is critical for circuit functionality and brain function[1]. Overexpression of S-SCAM in excitatory neurons enhances AMPA receptor (AMPAR)-mediated synaptic transmission through TARP11,12, without altering NMDA receptor (NMDAR) and presynaptic function[11]. S-SCAM transgenic mice showed reduced GABAA receptor α1 levels, in the synaptosomal fraction (biochemical correlates of synapses) without alterations in its total protein levels[13], indicating the possibility of defects in GABAergic synapses. We describe the effect of altering S-SCAM protein levels, mimicking the conditions in schizophrenia or infantile spasms, in cultured rat hippocampal neurons on the GABAergic synapses using immunocytochemistry combined with molecular genetic, pharmacological, and biochemical approaches. Our studies reveal the profound importance of S-SCAM in maintaining the proper balance of excitatory and inhibitory synapses in neurons and provide a clue to the pathogenic properties of S-SCAM copy number variations

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