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Numerous rare variants that cause neurodevelopmental disorders (NDDs) occur within genes encoding synaptic proteins, including ionotropic glutamate receptors. However, in many cases, it remains unclear how damaging missense variants affect brain function. We determined the physiological consequences of an NDD causing missense mutation in the GRIK2 kainate receptor (KAR) gene, that results in a single amino acid change p.Ala657Thr in the GluK2 receptor subunit. We engineered this mutation in the mouse Grik2 gene, yielding a GluK2(A657T) mouse, and studied mice of both sexes to determine how hippocampal neuronal function is disrupted. Synaptic KAR currents in hippocampal CA3 pyramidal neurons from heterozygous A657T mice exhibited slow decay kinetics, consistent with incorporation of the mutant subunit into functional receptors. Unexpectedly, CA3 neurons demonstrated elevated action potential spiking because of downregulation of the small-conductance Ca2+ activated K+ channel (SK), which mediates the post-spike afterhyperpolarization. The reduction in SK activity resulted in increased CA3 dendritic excitability, increased EPSP-spike coupling, and lowered the threshold for the induction of LTP of the associational-commissural synapses in CA3 neurons. Pharmacological inhibition of SK channels in WT mice increased dendritic excitability and EPSP-spike coupling, mimicking the phenotype in A657T mice and suggesting a causative role for attenuated SK activity in aberrant excitability observed in the mutant mice. These findings demonstrate that a disease-associated missense mutation in GRIK2 leads to altered signaling through neuronal KARs, pleiotropic effects on neuronal and dendritic excitability, and implicate these processes in neuropathology in patients with genetic NDDs. [ABSTRACT FROM AUTHOR]

Perampanel (PMP) is a third-generation antiseizure drug reported to be a potent and selective noncompetitive negative allosteric modulator of one subfamily of ionotropic glutamate receptor (iGluR), the a-amino-3-hydroxy-S-methylisoxazole-4-propionic acid receptors (AMPARs). However, the recent structural resolution of AMPARs in complex with PMP revealed that its binding pocket is formed from residues that are largely conserved in two members of another family of iGluRs, the GluK4 and GluK5 kainate receptor (KAR) subunits. We show here that PMP inhibits both recombinant and neuronal KARs, contrary to the previous reports, and that the negative allosteric modulator (NAM) activity requires GluK5 subunits to be channel constituents. PMP inhibited heteromeric GluK1/GluK5 and GluK2/GluK5 KARs at IC50 values comparable to that for AMPA receptors but was much less potent on homomeric GluK1 or GluK2 KARs. The auxiliary subunits Neto1 or Neto2 also made GluK2-containing KARs more sensitive to inhibition. Finally, PMP inhibited mouse neuronal KARs containing GluK5 subunits and Neto proteins in nociceptive dorsal root ganglia neurons and hippocampal mossy fiber--CA3 pyramidal neuron synapses. These data suggest that clinical actions of PMP could arise from differential inhibition of AMPAR or KAR signaling and that more selective drugs might maintain antiseizure efficacy while reducing adverse effects. [ABSTRACT FROM AUTHOR]

Appropriate excitatory/inhibitory (E/l) balance is essential for normal cortical function and is altered in some psychiatric disorders, including autism spectrum disorders (ASDs). Cell-autonomous molecular mechanisms that control the balance of excitatory and inhibitory synapse function remain poorly understood; no proteins that regulate excitatory and inhibitory synapse strength in a coordinated reciprocal manner have been identified. Using super-resolution imaging, electrophysiology, and molecular manipulations, we show that cadherin-10, encoded by CDH10 within the ASD risk locus 5pl4.1, maintains both excitatory and inhibitory synaptic scaffold structure in cultured cortical neurons from rats of both sexes. Cadherin-10 localizes to both excitatory and inhibitory synapses in neocortex, where it is organized into nanoscale puncta that influence the size of their associated PSDs. Knockdown of cadherin-10 reduces excitatory but increases inhibitory synapse size and strength, altering the E/I ratio in cortical neurons. Furthermore, cadherin-10 exhibits differential participation in complexes with PSD-95 and gephyrin, which may underlie its role in maintaining the E/l ratio. Our data provide a new mechanism whereby a protein encoded by a common ASD risk factor controls E/I ratios by regulating excitatory and inhibitory synapses in opposing directions. [ABSTRACT FROM AUTHOR]

Peripheral sensory neurons in the dorsal root ganglia (DRG) are the initial transducers of sensory stimuli, including painful stimuli, from the periphery to central sensory and pain-processing centers. Small- to medium-diameter non-peptidergic neurons in the neonatal DRG express functional kainate receptors (KARs), one of three subfamilies of ionotropic glutamate receptors, as well as the putative KAR auxiliary subunit Neuropilin- and tolloid-like 2 (Neto2). Neto2 alters recombinant KAR function markedly but has yet to be confirmed as an auxiliary subunit that assembles with and alters the function of endogenous KARs. KARs in neonatal DRG require the GluK1 subunit as a necessary constituent, but it is unclear to what extent other KAR subunits contribute to the function and proposed roles of KARs in sensory ganglia,which include promotion of neurite outgrowth and modulation of glutamate release at the DRG-dorsal horn synapse. In addition, KARs containing the GluK1 subunit are implicated in modes of persistent but not acute pain signaling. We show here that the Neto2 protein is highly expressed in neonatal DRG and modifies KAR gating in DRG neurons in a developmentally regulated fashion in mice. Although normally at very low levels in adult DRG neurons, Neto2 protein expression can be upregulated via MEK/ERK signaling and after sciatic nerve crush and Neto2-/- neurons from adult mice have stunted neurite outgrowth. These data confirm that Neto2 is a bona fide KAR auxiliary subunit that is an important constituent of KARs early in sensory neuron development and suggest that Neto2 assembly is critical to KAR modulation of DRG neuron process outgrowth. [ABSTRACT FROM AUTHOR]

Mossy fiber synapses act as the critical mediators of highly dynamic communication between hippocampal granule cells in the dentate gyrus and CA3 pyramidal neurons. Excitatory synaptic strength at mossy fiber to CA3 pyramidal cell synapses is potentiated rapidly and reversibly by brief trains of low-frequency stimulation of mossy fiber axons. We show that slight modifications to the pattern of stimulation convert this short-term potentiation into prolonged synaptic strengthening lasting tens of minutes in rodent hippocampal slices. This low-frequency potentiation of mossy fiber EPSCs requires postsynaptic mGlu1 receptors for induction but is expressed presynaptically as an increased release probability and therefore impacts bothAMPAandNMDAcomponents of the mossy fiber EPSC. A nonconventional signaling pathway initiated by mGlu1 receptors contributes to induction of plasticity, because EPSC potentiation was prevented by a tyrosine kinase inhibitor and only partially reduced by guanosine 5=-O-(2-thiodiphosphate). A slowly reversible state of enhanced synaptic efficacy could serve as a mechanism for altering the integrative properties of this synapse within a relatively broad temporal window. [ABSTRACT FROM AUTHOR]

General or brain-region-specific decreases in spine number or morphology accompany major neuropsychiatric disorders. It is unclear, however, whether changes in spine density are specific for an individual mental process or disorder and, if so, which molecules confer such specificity. Here we identify the scaffolding protein IQGAP1 as a key regulator of dendritic spine number with a specific role in cognitive but not emotional or motivational processes. We show that IQGAP1 is an important component of NMDAR multiprotein complexes and functionally interacts with the NR2A subunits and the extracellular signal-regulated kinase 1 (ERK1) and ERK2 signaling pathway. Mice lacking the IQGAP1 gene exhibited significantly lower levels of surface NR2A and impaired ERK activity compared to their wild-type littermates. Accordingly, primary hippocampal cultures of IQGAP1-/-neurons exhibited reduced surface expression of NR2A and disrupted ERK signaling in response to NR2A-dependent NMDAR stimulation. These molecular changes were accompanied by region-specific reductions of dendritic spine density in key brain areas involved in cognition, emotion, and motivation. IQGAP1 knock-outs exhibited marked long-term memory deficits accompanied by impaired hippocampal long-term potentiation (LTP) in a weak cellular learning model; in contrast, LTP was unaffected when induced with stronger stimulation paradigms. Anxiety- and depression like behavior remained intact. On the basis of these findings, we propose that a dysfunctional IQGAP1 gene contributes to the cognitive deficits in brain disorders characterized by fewer dendritic spines. [ABSTRACT FROM AUTHOR]

Auxiliary proteins modify the biophysical function and pharmacological properties of ionotropic glutamate receptors and likely are important components of receptor signaling complexes in vivo. The neuropilin and tolloid-like proteins (NETO) 1 and NETO2, two closely related CUB domain-containing integral membrane proteins, were identified recently as auxiliary proteins that slowed GluK2a kainate receptor current kinetics without impacting receptor membrane localization. Here we demonstrate that NETO2 profoundly slows the desensitization rate of GluK1 kainate receptors, promotes plasma membrane localization of transfected receptors in heterologous cells and rat hippocampal neurons, and targets GluK1-containing receptors to synapses. Conversely, the closely related protein NETO1 increases the rate of GluK1 receptor desensitization. Incorporation ofNETOproteins into kainate receptor-signaling complexes therefore extends the temporal range of receptor gating by over an order of magnitude. The presence of these auxiliary proteins could underlie some of the unusual aspects of kainate receptor function in the mammalian CNS. [ABSTRACT FROM AUTHOR]

Kainate receptors (KARs) are neuronal proteins that exhibit a highly polarized distribution in the mammalian CNS. Assembly, intracellular trafficking, and synaptic targeting of KARs and other ionotropic glutamate receptors are processes controlled, in part, by various determinants within the constituent subunit proteins themselves. Here, we demonstrate that the linker region between the M3 and S2 domains, which in current structural models is thought to transduce ligand-binding energy into channel opening, additionally has an essential role in receptor biogenesis. Our results show that this gating-associated domain is engaged at two distinct critical stages of KAR biogenesis: first, during the transition from dimeric to tetrameric assembly states and, second, at a postassembly trafficking checkpoint within the endoplasmic reticulum. Alteration of a basic residue, arginine 663, altered the desensitization properties of the GluR6 kainate receptor in response to glutamate application, and these changes were weakly correlated with intracellular retention of the mutant receptors. Elimination of the positive charge also significantly attenuated oligomerization and stability of the intracellular subunit protein. Furthermore, charge swapping with an adjacent residue, glutamate 662, normalized the receptor physiological behavior and reversed the deficits in assembly and degradation, but only partially restored plasma membrane expression of the receptors. These results reveal a new role for this linker domain in glutamate receptor biogenesis and contribute to understanding the cellular controls of receptor assembly and trafficking, which will be important for relating receptor stoichiometry to their neuronal targeting and function. [ABSTRACT FROM AUTHOR]

Kainate receptor-mediated components of postsynaptic currents at hippocampal mossy fiber synapses have markedly slower kinetics than currents arising from AMPA receptors. Here, we demonstrate that other aspects of kainate and AMPA receptor function at this synapse are distinct; in particular, kainate receptor currents are less sensitive to short- and long-term increases in presynaptic strength. EPSCs arising predominantly from AMPA receptors exhibited well characterized paired-pulse facilitation, frequency facilitation, and NMDA receptor-independent long-term potentiation, whereas isolated kainate receptor synaptic currents (KA-EPSCs) exhibited attenuated facilitation and long-term potentiation. In addition, KA-EPSCs varied in their sensitivity to a low-affinity competitive antagonist, suggestive of a synaptic heterogeneity greater than that of EPSCs comprised predominantly of AMPA receptors. These data suggest that the proportional contribution of AMPA and kainate receptors to ensemble synaptic currents will vary depending on the firing frequency of mossy fiber afferents. These synaptic features may be a mechanism for limiting activation of kainate receptors at mossy fiber synapses, which has been shown to be involved in seizurogenic firing of the CA3 network. [ABSTRACT FROM AUTHOR]

Intracellular trafficking of ionotropic glutamate receptors is regulated predominantly by determinants in the cytoplasmic C-terminal domain of the subunit proteins. Although AMPA receptors are found at the vast majority of excitatory synapses, synaptic kainate receptors exhibit a much more restricted distribution, suggesting that specific mechanisms exist for selective trafficking of these receptor proteins. In this report, we define a critical forward trafficking motif that is necessary for surface expression of the glutamate receptor 6 (GluR6) kainate receptor as well as chimeric proteins containing only the GluR6 C-terminal domain. The trafficking determinant was identified by tracking surface expression of green fluorescent protein-tagged GluR6 receptors with confocal immunofluorescence in COS-7 cells and cultured neurons and patch-clamp electrophysiology in human embryonic kidney 293 cells. Serial truncation and alanine site mutagenesis of the GluR6 subunit C terminus localized the critical motif to a seven amino acid stretch of predominantly basic residues. Alanine mutation of the trafficking motif reduced kainate receptor current amplitudes by >90% and resulted in retention of the mutated receptors in the endoplasmic reticulum. This forward trafficking domain is the first such identified for kainate receptors. [ABSTRACT FROM AUTHOR]

Presents a study that investigated the distribution of kainate receptor subunits in mossy fiber synapses using subunit-selective antibodies and knock-out mice. Materials and methods; Characterization of subunit-specific antibodies; Ultrastructural localization of kainate receptor subunits in mossy fiber synapes; Details on the assembly of kainate receptor subunits in mouse hippocampus.

Presents a study which investigated the assembly-dependent trafficking of the kainate receptor subunit KA2. Review of related literature; Materials and methods used; Results and discussion.

Presents a study which examined the effect of genetic ablation of kainate (KA) receptors on synaptic transmission at mossy-fiber pyramidal cell synapse. Heteromeric combinations which form KA receptors; Generation and molecular characterization of KA2[sup-I-] mutant mice; Mossy-fiber long-term plasticity in KA2[sup-I-] mice.