Loss of Neuroligin3 specifically downregulates retinal GABAAα2 receptors without abolishing direction selectivity.

Mrinalini Hoon,Vidhyasankar Krishnamoorthy,Frederique Varoqueaux,Bjoern Falkenburger,Tim Gollisch,William D Phillips

doi:10.1371/journal.pone.0181011

Abstract

The postsynaptic adhesion proteins Neuroligins (NLs) are essential for proper synapse function, and their alterations are associated with a variety of neurodevelopmental disorders. It is increasingly clear that each NL isoform occupies specific subsets of synapses and is able to regulate the function of discrete networks. Studies of NL2 and NL4 in the retina in particular have contributed towards uncovering their role in inhibitory synapse function. In this study we show that NL3 is also predominantly expressed at inhibitory postsynapses in the retinal inner plexiform layer (IPL), where it colocalizes with both GABAA- and glycinergic receptor clusters in a 3:2 ratio. In the NL3 deletion-mutant (knockout or KO) mouse, we uncovered a dramatic reduction of the number of GABAAα2-subunit containing GABAA receptor clusters at the IPL. Retinal activity was thereafter assessed in KO and wild-type (WT) littermates by multi-electrode-array recordings of the output cells of retina, the retinal ganglion cells (RGCs). RGCs in the NL3 KO showed reduced spontaneous activity and an altered response to white noise stimulation. Moreover, upon application of light flashes, the proportion of cells firing at light offset (OFF RGCs) was significantly lower in the NL3 KO compared to WT littermates, whereas the relative number of cells firing at light onset (ON RGCs) increased. Interestingly, although GABAAα2-bearing receptors have been related to direction-selective circuits of the retina, features of direction selective-retinal ganglion cells recorded remained unperturbed in the NL3 KO. Together our data underscore the importance of NL3 for the integrity of specific GABAAergic retinal circuits and identifies NL3 as an important regulator of retinal activity.

Highlights

Neuroligins (NL1-4) are postsynaptic transmembrane adhesion molecules crucial for synapse maturation and function [1, 2]
About 40% of NL3 puncta colocalized with gephyrin, whereas ~55% associated with GABAAγ2 (Fig 2E)
We found a specific reduction in the number of GABAAα2 subunit-containing receptor clusters in the NL3 KO (WTmean = 131.07 ± 8.56 puncta/100 μm2 of inner plexiform layer (IPL); KOmean = 45.21 ± 1.05 puncta/100 μm2 of IPL; n = 4 WT-KO littermate pairs; p = 0.01) (Fig 4A and 4B), whereas the other GABAA receptor cluster densities remained comparable in both groups (Fig 4A and 4B)

Summary

Introduction

Neuroligins (NL1-4) are postsynaptic transmembrane adhesion molecules crucial for synapse maturation and function [1, 2]. The roles of NL3 and NL4 have been investigated in diverse neural networks using NL3 and NL4 deletion-mutant mice [1, 6,7,8]. NL3 is widely expressed in the CNS [1] and has been reported at both excitatory and inhibitory postsynapses of cultured hippocampal neurons [11] and in the cerebellum [8]. Initial studies failed to uncover significant changes in excitatory or inhibitory transmission in the cortex and the hippocampus of the NL3 deletion-mutant mouse [6], but more recently subtle alterations in mGluR1-mediated signaling and excitatory transmission have been reported in the cerebellum of the NL3 deletion-mutant mouse [8]. Alterations in tonic endocannabinoid signaling and inhibitory transmission have been uncovered in the hippocampus of the NL3 deletion-mutant mouse [12]

Methods

Results

Conclusion