Neuroligin-4 is localized to glycinergic postsynapses and regulates inhibition in the retina

Abstract

Neuroligins (NL1-NL4) are postsynaptic adhesion proteins that control the maturation and function of synapses in the central nervous system (CNS). Loss-of-function mutations in NL4 are linked to rare forms of monogenic heritable autism, but its localization and function are unknown. Using the retina as a model system, we show that NL4 is preferentially localized to glycinergic postsynapses and that the loss of NL4 is accompanied by a reduced number of glycine receptors mediating fast glycinergic transmission. Accordingly, NL4-deficient ganglion cells exhibit slower glycinergic miniature postsynaptic currents and subtle alterations in their stimuluscoding efficacy, and inhibition within the NL4-deficient retinal network is altered as assessed by electroretinogram recordings. These data indicate that NL4 shapes network activity and information processing in the retina by modulating glycinergic inhibition. Importantly, NL4 is also targeted to inhibitory synapses in other areas of the CNS, such as the thalamus, colliculi, brainstem, and spinal cord, and forms complexes with the inhibitory postsynapse proteins gephyrin and collybistin in vivo, indicating that NL4 is an important component of glycinergic postsynapses.synaptogenesis | inhibitory transmission | visual processing I n rodents, postsynaptic adhesion proteins of the neuroligin family (NL1-NL4) are expressed throughout the central nervous system (CNS) (1-3) and essential for synapse organization and function (2-5). In vivo, each NL isoform localizes to specific synapse subpopulations, with NL1, NL2, and NL3 predominantly associating with glutamatergic, GABAergic, or both types of postsynapses, respectively (1, 6-9).Thus far, the distribution and function of the fourth NL isoform has remained unclear, despite the wide interest triggered by the causal link of specific loss-of-function mutations in NL4 to cases of autism, which led to the notion that aberrant synaptic transmission may cause autism spectrum disorders (ASDs) (10).We examined the distribution of NL4 in the mouse retina, a well-characterized region of the CNS with distinct, topographically organized glutamatergic, GABAergic, and glycinergic synapses, which has recently allowed us to characterize crucial aspects of NL2 distribution and function (8). Additionally, we assessed NL4 function by studying synaptic activity and visual processing in the NL4-deficient (NL4-KO; ref.3) mouse retina. Finally, we studied NL4 localization in the rest of the CNS and identified some of its key binding partners at the synapse. Results NL4 Is Localized to Glycinergic Postsynapses in the Retina.We characterized the distribution of NL4 by immunohistochemistry by using an isoform-specific antibody (3) (Fig. 1). A punctate labeling was detected in the inner plexiform layer (IPL) of wildtype (WT) but not NL4-KO retinae (Fig. 1A). NL4-positive puncta were abundant in the outer IPL but sparse in the rest of the IPL (Fig. 1A), which is reminiscent of glycine receptor (GlyR) distribution in the retina (11,12). Indeed, upon co...

Molecular and functional heterogeneity of GABAergic synapses

“…As noted above in the section on NL2, GABAergic, but not glycinergic synapses, selectively contain this NL isoform, whereas NL3 and NL4 have been reported in glycinergic synapses, at least in the retina [37]. Since NL2 interacts with CB and gephyrin at GABAergic synapses, this difference in localization might explain why CB deletion does not affect the PSD of glycinergic synapses.…”

“…Apparently, NL2 is absent from glycinergic synapses, whereas NL3 and NL4 have been reported to be present in a subset of GABAergic and glycinergic synapses, respectively [36,37]. The significance of a segregated distribution of NLs to different types of synapses is not fully understood [38], because this apparent selectively is lost upon overexpression and because NL3 has been reported to be present in both glutamatergic and GABAergic synapses in vivo.…”

Proteomic Characterization of Inhibitory Synapses Using a Novel pHluorin-tagged γ-Aminobutyric Acid Receptor, Type A (GABAA), α2 Subunit Knock-in Mouse

“…However, in addition, the single step purification resulted in the isolation of ␥1, ␥3, and ␦ subunits (Table 2). Furthermore, a number of other previously verified interactions were confirmed, including binding of GABA A Rs or their closely associated proteins to gephyrin (Gphn), collybistin (Arhgef9), neuroligins 1-4 (Nlgn), PKC isoforms (Prkc), PKA (Prkacb), GABA B R2 (Gabbr2), and glycine receptor ␤ (Glrb) as described previously (12,22,(25)(26)(27)(28)(29)(30)(31). Crucially, a key component of excitatory synapses, the highly abundant PSD95 family of proteins (32), was absent from these purifications.…”

Neuroligin-1 controls synaptic abundance of NMDA-type glutamate receptors through extracellular coupling

“…NL1 is localized almost exclusively to glutamatergic synapses, NL2 is restricted to GABA and glycinergic synapses, NL3 is detected at both glutamatergic and GABAergic sites, and NL4 is detected at a subset of glycinergic synapses in the retina (18,25,(38)(39)(40). The mechanisms underlying this synaptic specificity of the NL isoforms and the selective association with neurotransmitter receptors at the respective synapses are poorly understood.…”