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Developmental neurogenesis is a tightly regulated spatiotemporal process with its dysregulation implicated in neurodevelopmental disorders. NMDA receptors are glutamate-gated ion channels that are widely expressed in the early nervous system, yet their contribution to neurogenesis is poorly understood. Notably, a variety of mutations in genes encoding NMDA receptor subunits are associated with neurodevelopmental disorders. To rigorously define the role of NMDA receptors in developmental neurogenesis, we used a mutant zebrafish line (grin1 -/- ) that lacks all NMDA receptors yet survives to 10 days post-fertilization, offering the opportunity to study post-embryonic neurodevelopment in the absence of NMDA receptors. Focusing on the forebrain, we find that these fish have a progressive supernumerary neuron phenotype confined to the telencephalon at the end of embryonic neurogenesis, but which extends to all forebrain regions during postembryonic neurogenesis. This enhanced neuron population does not arise directly from increased numbers or mitotic activity of radial glia cells, the principal neural stem cells. Rather, it stems from a lack of timely maturation of transit-amplifying neuroblasts into post-mitotic neurons, as indicated by a decrease in expression of the ontogenetically-expressed chloride transporter, KCC2. Pharmacological blockade with MK-801 recapitulates the grin1 -/- supernumerary neuron phenotype, indicating a requirement for ionotropic signaling. Thus, NMDA receptors are required for suppression of indirect, transit amplifying cell-driven neurogenesis by promoting maturational termination of mitosis. Loss of suppression results in neuronal overpopulation that can fundamentally change brain circuitry and may be a key factor in pathogenesis of neurodevelopmental disorders caused by NMDA receptor dysfunction., (© 2024. The Author(s).)

Zebrafish are a powerful system to study brain development and to dissect the activity of complex circuits. One advantage is that they display complex behaviors, including prey capture, learning, responses to photic and acoustic stimuli, and social interaction (Dreosti et al., Front Neural Circuits 9:39, 2015; Bruckner et al., PLoS Biol 20:e3001838, 2022; Zoodsma et al., Mol Autism 13:38, 2022) that can be probed to assess brain function. Many of these behaviors are easily assayed at early larval stages, offering a noninvasive and high-throughput readout of nervous system function. Additionally, larval zebrafish readily uptake small molecules dissolved in water making them ideal for behavioral-based drug screens. Together, larval zebrafish and their behavioral repertoire offer a means to rapidly dissect brain circuitry and can serve as a template for high-throughput small molecule screens.NMDA receptor subunits are highly conserved in zebrafish compared to mammals (Zoodsma et al., Mol Autism 13:38, 2022; Cox et al., Dev Dyn 234:756-766, 2005; Zoodsma et al., J Neurosci 40:3631-3645, 2020). High amino acid and domain structure homology between humans and zebrafish underlie conserved functional similarities. Here we describe a set of behavioral assays that are useful to study the NMDA receptor activity in brain function., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

The NMDA receptor (NMDAR) is a ubiquitously expressed glutamate-gated ion channel that plays key roles in brain development and function. Not surprisingly, a variety of disease-associated variants have been identified in genes encoding NMDAR subunits. A critical first step to assess whether these variants contribute to their associated disorder is to characterize their effect on receptor function. However, the complexity of NMDAR function makes this challenging, with many variants typically altering multiple functional properties. At synapses, NMDARs encode pre- and postsynaptic activity to carry a charge transfer that alters membrane excitability and a Ca 2+ influx that has both short- and long-term signaling actions. Here, we characterized epilepsy-associated variants in GluN1 and GluN2A subunits with various phenotypic severity in HEK293 cells. To capture the complexity of NMDAR gating, we applied 10 glutamate pulses at 10 Hz to derive a charge integral. This assay is advantageous since it incorporates multiple gating parameters - activation, deactivation, and desensitization - into a single value. We then integrated this gating parameter with Mg 2+ block and Ca 2+ influx using fractional Ca 2+ currents to generate indices of charge transfer and Ca 2+ transfer over wide voltage ranges. This approach yields consolidated parameters that can be used as a reference to normalize channel block and allosteric modulation to better define potential patient treatment. This is especially true for variants in the transmembrane domain that affect not only receptor gating but also often Mg 2+ block and Ca 2+ permeation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) auxiliary subunits are specialized, nontransient binding partners of AMPARs that modulate AMPAR channel gating properties and pharmacology, as well as their biogenesis and trafficking. The most well-characterized families of auxiliary subunits are transmembrane AMPAR regulatory proteins (TARPs), cornichon homologs (CNIHs), and the more recently discovered GSG1-L. These auxiliary subunits can promote or reduce surface expression of AMPARs (composed of GluA1-4 subunits) in neurons, thereby impacting their functional role in membrane signaling. Here, we show that CNIH-2 enhances the tetramerization of WT and mutant AMPARs, presumably by increasing the overall stability of the tetrameric complex, an effect that is mainly mediated by interactions with the transmembrane domain of the receptor. We also find CNIH-2 and CNIH-3 show receptor subunit-specific actions in this regard with CNIH-2 enhancing both GluA1 and GluA2 tetramerization, whereas CNIH-3 only weakly enhances GluA1 tetramerization. These results are consistent with the proposed role of CNIHs as endoplasmic reticulum cargo transporters for AMPARs. In contrast, TARP γ-2, TARP γ-8, and GSG1-L have no or negligible effect on AMPAR tetramerization. On the other hand, TARP γ-2 can enhance receptor tetramerization but only when directly fused with the receptor at a maximal stoichiometry. Notably, surface expression of functional AMPARs was enhanced by CNIH-2 to a greater extent than TARP γ-2, suggesting that this distinction aids in maturation and membrane expression. These experiments define a functional distinction between CNIHs and other auxiliary subunits in the regulation of AMPAR biogenesis., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Kinetics of NMDA receptor (NMDAR) ion channel opening and closing contribute to their unique role in synaptic signaling. Agonist binding generates free energy to open a canonical gate at the M3 helix bundle crossing. Single channel activity is characterized by clusters, or periods of rapid opening and closing, that are separated by long silent periods. A conserved glycine in the outer most transmembrane helices, the M4 helices, regulates NMDAR function. Here we find that the GluN1 glycine mainly regulates single channel events within a cluster, whereas the GluN2 glycine mainly regulates entry and exit from clusters. Molecular dynamics simulations suggest that, whereas the GluN2 M4 (along with GluN2 pre-M1) regulates the gate at the M3 helix bundle crossing, the GluN1 glycine regulates a 'gate' at the M2 loop. Subsequent functional experiments support this interpretation. Thus, the distinct kinetics of NMDARs are mediated by two gates that are under subunit-specific regulation., (© 2023. The Author(s).)

Background: Autism spectrum disorder (ASD), like many neurodevelopmental disorders, has complex and varied etiologies. Advances in genome sequencing have identified multiple candidate genes associated with ASD, including dozens of missense and nonsense mutations in the NMDAR subunit GluN2B, encoded by GRIN2B. NMDARs are glutamate-gated ion channels with key synaptic functions in excitatory neurotransmission. How alterations in these proteins impact neurodevelopment is poorly understood, in part because knockouts of GluN2B in rodents are lethal., Methods: Here, we use CRISPR-Cas9 to generate zebrafish lacking GluN2B (grin2B -/- ). Using these fish, we run an array of behavioral tests and perform whole-brain larval imaging to assay developmental roles and functions of GluN2B., Results: We demonstrate that zebrafish GluN2B displays similar structural and functional properties to human GluN2B. Zebrafish lacking GluN2B (grin2B -/- ) surprisingly survive into adulthood. Given the prevalence of social deficits in ASD, we assayed social preference in the grin2B -/- fish. Wild-type fish develop a strong social preference by 3 weeks post fertilization. In contrast, grin2B -/- fish at this age exhibit significantly reduced social preference. Notably, the lack of GluN2B does not result in a broad disruption of neurodevelopment, as grin2B -/- larvae do not show alterations in spontaneous or photic-evoked movements, are capable of prey capture, and exhibit learning. Whole-brain imaging of grin2B -/- larvae revealed reduction of an inhibitory neuron marker in the subpallium, a region linked to ASD in humans, but showed that overall brain size and E/I balance in grin2B -/- is comparable to wild type., Limitations: Zebrafish lacking GluN2B, while useful in studying developmental roles of GluN2B, are unlikely to model nuanced functional alterations of human missense mutations that are not complete loss of function. Additionally, detailed mammalian homologies for larval zebrafish brain subdivisions at the age of whole-brain imaging are not fully resolved., Conclusions: We demonstrate that zebrafish completely lacking the GluN2B subunit of the NMDAR, unlike rodent models, are viable into adulthood. Notably, they exhibit a highly specific deficit in social behavior. As such, this zebrafish model affords a unique opportunity to study the roles of GluN2B in ASD etiologies and establish a disease-relevant in vivo model for future studies., (© 2022. The Author(s).)

Synaptotagmins belong to a large family of proteins. Although various synaptotagmins have been implicated as Ca 2+ sensors for vesicle replenishment and release at conventional synapses, their roles at retinal ribbon synapses remain incompletely understood. Zebrafish is a widely used experimental model for retinal research. We therefore investigated the homology between human, rat, mouse, and zebrafish synaptotagmins 1-10 using a bioinformatics approach. We also characterized the expression and distribution of various synaptotagmin (syt) genes in the zebrafish retina using RT-PCR, qPCR, and in situhybridization, focusing on the family members whose products likely underlie Ca 2+ -dependent exocytosis in the central nervous system (synaptotagmins 1, 2, 5, and 7). Most zebrafish synaptotagmins are well conserved and can be grouped in the same classes as mammalian synaptotagmins, based on crucial amino acid residues needed for coordinating Ca 2+ binding and determining phospholipid binding affinity. The only exception is synaptotagmin 1b, which lacks 34 amino acid residues in the C2B domain and is therefore unlikely to bind Ca 2+ there. Additionally, the products of zebrafish syt5a and syt5b genes share identity with mammalian class 1 and 5 synaptotagmins. Zebrafish syt1, syt2, syt5, and syt7 paralogues are found in the zebrafish brain, eye, and retina, excepting syt1b, which is only present in the brain. The complementary expression pattern of the remaining paralogues in the retina suggests that syt1a and syt5a may underlie synchronous release and syt7a and syt7b may mediate asynchronous release or other Ca 2+ -dependent processes in different retinal neurons., (© 2021 Wiley Periodicals LLC.)

Many physiologic effects of l-glutamate, the major excitatory neurotransmitter in the mammalian central nervous system, are mediated via signaling by ionotropic glutamate receptors (iGluRs). These ligand-gated ion channels are critical to brain function and are centrally implicated in numerous psychiatric and neurologic disorders. There are different classes of iGluRs with a variety of receptor subtypes in each class that play distinct roles in neuronal functions. The diversity in iGluR subtypes, with their unique functional properties and physiologic roles, has motivated a large number of studies. Our understanding of receptor subtypes has advanced considerably since the first iGluR subunit gene was cloned in 1989, and the research focus has expanded to encompass facets of biology that have been recently discovered and to exploit experimental paradigms made possible by technological advances. Here, we review insights from more than 3 decades of iGluR studies with an emphasis on the progress that has occurred in the past decade. We cover structure, function, pharmacology, roles in neurophysiology, and therapeutic implications for all classes of receptors assembled from the subunits encoded by the 18 ionotropic glutamate receptor genes. SIGNIFICANCE STATEMENT: Glutamate receptors play important roles in virtually all aspects of brain function and are either involved in mediating some clinical features of neurological disease or represent a therapeutic target for treatment. Therefore, understanding the structure, function, and pharmacology of this class of receptors will advance our understanding of many aspects of brain function at molecular, cellular, and system levels and provide new opportunities to treat patients., (U.S. Government work not protected by U.S. copyright.)

NMDA receptors are ligand-gated ion channels that are found throughout the brain and are required for both brain development and many higher order functions. A variety of human patients with diverse clinical phenotypes have been identified that carry autoantibodies directed against NMDA receptor subunits. Here we focus on two general classes of autoantibodies, anti-GluN1 antibodies associated with anti-NMDA receptor encephalitis and anti-GluN2 antibodies associated with systemic lupus erythematosus (SLE). These two general classes of anti-NMDA receptor autoantibodies display a wide range of pathophysiological mechanisms from altering synaptic composition to gating of NMDARs. While we have made progress in understanding how these autoantibodies work at the molecular and cellular level, many unanswered questions remain including their long-term actions on brain function, the significance of clonal variations, and their effects on different NMDA receptor-expressing cell types in local circuits. This information will be needed to define fully the transition from anti-NMDA receptor autoantibodies to a clinical phenotype., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

NMDA receptors (NMDARs) are glutamate-gated ion channels that mediate fast excitatory synaptic transmission in the nervous system. Applying glutamate to outside-out patches containing a single NMDAR, we find that agonist-bound receptors transition to the open state via two conformations, an "unconstrained pre-active" state that contributes to fast synaptic events and a "constrained pre-active" state that does not. To define how glutamate drives these conformations, we decoupled the ligand-binding domains from specific transmembrane segments for GluN1 and GluN2A. Displacements of the pore-forming M3 segments define the energy of fast opening. However, to enter the unconstrained conformation and contribute to fast signaling, the GluN2 pre-M1 helix must be displaced before the M3 segments move. This pre-M1 displacement is facilitated by the flexibility of the S2-M4 of GluN1 and GluN2A. Thus, outer structures-pre-M1 and S2-M4-work in concert to remove constraints and prime the channel for rapid opening, facilitating fast synaptic transmission., Competing Interests: Declaration Of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

NMDA receptors (NMDARs) are glutamate-gated ion channels that contribute to nearly all brain processes. Not surprisingly then, genetic variations in the genes encoding NMDAR subunits can be associated with neurodevelopmental, neurological and psychiatric disorders. These disease-associated variants (DAVs) present challenges, such as defining how DAV-induced alterations in receptor function contribute to disease progression and how to treat the affected individual clinically. As a starting point to overcome these challenges, we need to refine our understanding of the complexity of NMDAR structure function. In this regard, DAVs have expanded our knowledge of NMDARs because they do not just target well-known structure-function motifs, but rather give an unbiased view of structural elements that are important to the biology of NMDARs. Indeed, established NMDAR structure-function motifs have been validated by the appearance of disorders in patients where these motifs have been altered, and DAVs have identified novel structural features in NMDARs such as gating triads and hinges in the gating machinery. Still, the majority of DAVs remain unexplored and occur at sites in the protein with unidentified function or alter receptor properties in multiple and unanticipated ways. Detailed mechanistic and structural investigations are required of both established and novel motifs to develop a highly refined pathomechanistic model that accounts for the complex machinery that regulates NMDARs. Such a model would provide a template for rational drug design and a starting point for personalized medicine., (© 2020 The Authors. The Journal of Physiology © 2020 The Physiological Society.)

GABA A receptors (GABA A R) are inhibitory ion channels ubiquitously expressed in the central nervous system and play critical roles in brain development and function. Benzodiazepines are positive allosteric modulators of GABA A R, enhancing channel opening frequency when GABA is bound to the receptor. Midazolam is a commonly used benzodiazepine. It is frequently used for premature infants, but the long-term consequences of its use in this patient population are not well established. Here, we studied the acute effects of midazolam on immature synapses. Using a rodent organotypic hippocampal slice preparation, we evaluated how midazolam affects inhibitory synaptic transmission onto CA1 pyramidal neurons. We found that 1 μM midazolam enhances evoked inhibitory post synaptic currents (eIPSCs) at a holding potential of -60 mV. Similarly, 1 μM midazolam enhances miniature IPSCs (mIPSCs) in CA1 pyramidal neurons at holding potentials of -60 mV and -30 mV. At depolarized holding potentials, however, midazolam no longer enhances mIPSCs. Depolarization of the postsynaptic cell by itself increases mIPSC decay, which occludes the allosteric effects of midazolam. These results provide insight into how a benzodiazepine and membrane voltage may modulate GABA A R function in developing circuits., (Copyright © 2020. Published by Elsevier B.V.)

N -methyl-D-aspartate receptors (NMDARs) are glutamate-gated ion channels that play critical roles in neuronal development and nervous system function. Here, we developed a model to study NMDARs in early development in zebrafish, by generating CRISPR-mediated lesions in the NMDAR genes, grin1a and grin1b , which encode the obligatory GluN1 subunits. While receptors containing grin1a or grin1b show high Ca 2+ permeability, like their mammalian counterpart, grin1a is expressed earlier and more broadly in development than grin1b Both grin1a -/- and grin1b -/- zebrafish are viable. Unlike in rodents, where the grin1 knockout is embryonic lethal, grin1 double-mutant fish ( grin1a -/- ; grin1b -/- ), which lack all NMDAR-mediated synaptic transmission, survive until ∼10 d dpf (days post fertilization), providing a unique opportunity to explore NMDAR function during development and in generating behaviors. Many behavioral defects in the grin1 double-mutant larvae, including abnormal evoked responses to light and acoustic stimuli, prey-capture deficits, and a failure to habituate to acoustic stimuli, are replicated by short-term treatment with the NMDAR antagonist MK-801, suggesting that they arise from acute effects of compromised NMDAR-mediated transmission. Other defects, however, such as periods of hyperactivity and alterations in place preference, are not phenocopied by MK-801, suggesting a developmental origin. Together, we have developed a unique model to study NMDARs in the developing vertebrate nervous system. SIGNIFICANCE STATEMENT Rapid communication between cells in the nervous system depends on ion channels that are directly activated by chemical neurotransmitters. One such ligand-gated ion channel, the NMDAR, impacts nearly all forms of nervous system function. It has been challenging, however, to study the prolonged absence of NMDARs in vertebrates, and hence their role in nervous system development, due to experimental limitations. Here, we demonstrate that zebrafish lacking all NMDAR transmission are viable through early development and are capable of a wide range of stereotypic behaviors. As such, this zebrafish model provides a unique opportunity to study the role of NMDAR in the development of the early vertebrate nervous system., (Copyright © 2020 the authors.)

Patients with Systemic lupus erythematosus (SLE) experience various peripheral and central nervous system manifestations including spatial memory impairment. A subset of autoantibodies (DNRAbs) cross-react with the GluN2A and GluN2B subunits of the NMDA receptor (NMDAR). We find that these DNRAbs act as positive allosteric modulators on NMDARs with GluN2A-containing NMDARs, even those containing a single GluN2A subunit, exhibiting a much greater sensitivity to DNRAbs than those with exclusively GluN2B. Accordingly, GluN2A-specific antagonists provide greater protection from DNRAb-mediated neuronal cell death than GluN2B antagonists. Using transgenic mice to perturb expression of either GluN2A or GluN2B in vivo, we find that DNRAb-mediated disruption of spatial memory characterized by early neuronal cell death and subsequent microglia-dependent pathologies requires GluN2A-containing NMDARs. Our results indicate that GluN2A-specific antagonists or negative allosteric modulators are strong candidates to treat SLE patients with nervous system dysfunction.

N-methyl-D-aspartate receptors (NMDARs) mediate slow excitatory postsynaptic transmission in the central nervous system, thereby exerting a critical role in neuronal development and brain function. Rare genetic variants in the GRIN genes encoding NMDAR subunits segregated with neurological disorders. Here, we summarize the clinical presentations for 18 patients harboring 12 de novo missense variants in GRIN1, GRIN2A, and GRIN2B that alter residues in the M2 re-entrant loop, a region that lines the pore and is intolerant to missense variation. These de novo variants were identified in children with a set of neurological and neuropsychiatric conditions. Evaluation of the receptor cell surface expression, pharmacological properties, and biophysical characteristics show that these variants can have modest changes in agonist potency, proton inhibition, and surface expression. However, voltage-dependent magnesium inhibition is significantly reduced in all variants. The NMDARs hosting a single copy of a mutant subunit showed a dominant reduction in magnesium inhibition for some variants. These variant NMDARs also show reduced calcium permeability and single-channel conductance, as well as altered open probability. The data suggest that M2 missense variants increase NMDAR charge transfer in addition to varied and complex influences on NMDAR functional properties, which may underlie the patients' phenotypes., (© 2019 Wiley Periodicals, Inc.)

Clearance of synaptic vesicle proteins from active zones may be rate limiting for sustained neurotransmission. Issues of clearance are critical at ribbon synapses, which continually release neurotransmitters for prolonged periods of time. We used synaptophysin-pHluorin (SypHy) to visualize protein clearance from active zones in retinal bipolar cell ribbon synapses. Depolarizing voltage steps gave rise to small step-like changes in fluorescence likely indicating release of single SypHy molecules from fused synaptic vesicles near active zones. Temporal and spatial fluorescence profiles of individual responses were highly variable, but ensemble averages were well fit by clearance via free diffusion using Monte Carlo simulations. The rate of fluorescence decay of ensemble averages varied with the time and location of the fusion event, with clearance being most rapid at the onset of a stimulus when release rate is the highest., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

NMDA receptors (NMDARs) are glutamate-gated ion channels that are key mediators of excitatory neurotransmission and synaptic plasticity throughout the central nervous system. They form massive heterotetrameric complexes endowed with unique allosteric capacity provided by eight extracellular clamshell-like domains arranged as two superimposed layers. Despite an increasing number of full-length NMDAR structures, how these domains cooperate in an intact receptor to control its activity remains poorly understood. Here, combining single-molecule and macroscopic electrophysiological recordings, cysteine biochemistry, and in silico analysis, we identify a rolling motion at a yet unexplored interface between the two constitute dimers in the agonist-binding domain (ABD) layer as a key structural determinant in NMDAR activation and allosteric modulation. This rotation acts as a gating switch that tunes channel opening depending on the conformation of the membrane-distal N-terminal domain (NTD) layer. Remarkably, receptors locked in a rolled state display "super-activity" and resistance to NTD-mediated allosteric modulators. Our work unveils how NMDAR domains move in a concerted manner to transduce long-range conformational changes between layers and command receptor channel activity., (© 2018 The Authors.)

Parvalbumin-expressing (PV) GABAergic interneurons regulate local circuit dynamics. In terms of the excitation driving PV interneuron activity, the N-methyl-d-aspartate receptor (NMDAR)-mediated component onto PV interneurons tends to be smaller than that onto pyramidal neurons but makes a significant contribution to their physiology and development. In the visual cortex, PV interneurons mature during the critical period. We hypothesize that during the critical period, the NMDAR-mediated signaling and functional properties of glutamatergic synapses onto PV interneurons are developmentally regulated. We therefore compared the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR)- and NMDAR-mediated synaptic responses before (postnatal days 15-20, P15-P20), during (P25-P40), and after (P50-P60) the visual critical period. AMPAR miniature excitatory postsynaptic currents (mEPSCs) showed a developmental decrease in frequency, whereas NMDAR mEPSCs were absent or showed extremely low frequencies throughout development. For evoked responses, we consistently saw a NMDAR-mediated component, suggesting pre- or postsynaptic differences between evoked and spontaneous neurotransmission. Evoked responses showed input-specific developmental changes. For intralaminar inputs, the NMDAR-mediated component significantly decreased with development. This resulted in adult intralaminar inputs almost exclusively mediated by AMPARs, suited for the computation of synaptic inputs with precise timing, and likely having NMDAR-independent forms of plasticity. In contrast, interlaminar inputs maintained a stable NMDAR-mediated component throughout development but had a shift in the AMPAR paired-pulse ratio from depression to facilitation. Adult interlaminar inputs with facilitating AMPAR responses and a substantial NMDAR component would favor temporal integration of synaptic responses and could be modulated by NMDAR-dependent forms of plasticity. NEW & NOTEWORTHY We show for the first time input-specific developmental changes in the N-methyl-d-aspartate receptor component and short-term plasticity of the excitatory drive onto layers 2/3 parvalbumin-expressing (PV) interneurons in the visual cortex during the critical period. These developmental changes would lead to functionally distinct adult intralaminar and interlaminar glutamatergic inputs that would engage PV interneuron-mediated inhibition differently.

A variety of de novo and inherited missense mutations associated with neurological disorders are found in the NMDA receptor M4 transmembrane helices, which are peripheral to the pore domain in eukaryotic ionotropic glutamate receptors. Subsets of these mutations affect receptor gating with dramatic effects, including in one instance halting it, occurring at a conserved glycine near the extracellular end of M4. Functional experiments and molecular dynamic simulations of constructs with and without substitutions at this glycine indicate that it acts as a hinge, permitting the intracellular portion of the ion channel to laterally expand. This expansion stabilizes long-lived open states leading to slow deactivation and high Ca 2+ permeability. Our studies provide a functional and structural framework for the effect of missense mutations on NMDARs at central synapses and highlight how the M4 segment may represent a pathway for intracellular modulation of NMDA receptor function.

NMDA-type glutamate receptors are ligand-gated ion channels that mediate a Ca 2+ -permeable component of excitatory neurotransmission in the central nervous system (CNS). They are expressed throughout the CNS and play key physiological roles in synaptic function, such as synaptic plasticity, learning, and memory. NMDA receptors are also implicated in the pathophysiology of several CNS disorders and more recently have been identified as a locus for disease-associated genomic variation. NMDA receptors exist as a diverse array of subtypes formed by variation in assembly of seven subunits (GluN1, GluN2A-D, and GluN3A-B) into tetrameric receptor complexes. These NMDA receptor subtypes show unique structural features that account for their distinct functional and pharmacological properties allowing precise tuning of their physiological roles. Here, we review the relationship between NMDA receptor structure and function with an emphasis on emerging atomic resolution structures, which begin to explain unique features of this receptor., (© 2018 Hansen et al.)

A compound can change the activity of NMDA receptors in some regions of a synapse without affecting those in other regions., Competing Interests: JA, LW No competing interests declared, (© 2018, Amin et al.)

Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that mediate the vast majority of fast synaptic transmission in the nervous system. When the iGluR ion channel is in the open or conducting conformation, it is non-selective for monovalent cations, driving membrane excitation. Often the channel is also permeable to Ca 2+ . This process of Ca 2+ permeation and its physiological and pathological consequences depend strongly on the specific iGluR subtype as well as the specific subunits in the oligomeric complex. Recent evidence has highlighted additional levels of diversity to this process including a dependence on specific auxiliary subunits in non-NMDARs and post-translational modifications in NMDARs. Various de novo missense mutations associated with neurological disease in NMDAR subunits have been identified in regions critical to Ca 2+ influx. These features highlight the dynamics of Ca 2+ influx mediated by iGluRs and its critical role in synaptic physiology and pathology.

Ionotropic glutamate receptors (iGluRs), including AMPA receptor (AMPAR) and NMDA receptor (NMDAR) subtypes, are ligand-gated ion channels that mediate signaling at the majority of excitatory synapses in the nervous system. The iGluR pore domain is structurally and evolutionarily related to an inverted two-transmembrane K + channel. Peripheral to the pore domain in eukaryotic iGluRs is an additional transmembrane helix, the M4 segment, which interacts with the pore domain of a neighboring subunit. In AMPARs, the integrity of the alignment of a specific face of M4 with the adjacent pore domain is essential for receptor oligomerization. In contrast to AMPARs, NMDARs are obligate heterotetramers composed of two GluN1 and typically two GluN2 subunits. Here, to address the function of the M4 segments in NMDARs, we carry out a tryptophan scan of M4 in GluN1 and GluN2A subunits. Unlike AMPARs, the M4 segments in NMDAR subunits makes only a limited contribution to their biogenesis. However, the M4 segments in both NMDAR subunits are critical for receptor activation, with mutations at some positions, most notably at the extreme extracellular end, completely halting the gating process. Furthermore, although the AMPAR M4 makes a minimal contribution to receptor desensitization, the NMDAR M4 segments have robust and subunit-specific effects on desensitization. These findings reveal that the functional roles of the M4 segments in AMPARs and NMDARs have diverged in the course of their evolution and that the M4 segments in NMDARs may act as a transduction pathway for receptor modulation at synapses., (© 2017 Amin et al.)

NMDA receptors (NMDARs) are ion channels activated by the excitatory neurotransmitter glutamate and are essential to all aspects of brain function, including learning and memory formation. Missense mutations distributed throughout NMDAR subunits have been associated with an array of neurological disorders. Recent structural, functional, and computational studies have generated many insights into the activation process connecting glutamate binding to ion-channel opening, which is central to NMDAR physiology and pathophysiology. The field appears poised for breakthroughs, including the exciting prospect of resolving the conformations and energetics of elementary steps in the activation process, and atomic-level modeling of the effects of missense mutations on receptor function. The most promising strategy going forward is through strong integration of multiple approaches., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

AMPA receptors (AMPARs) mediate fast excitatory neurotransmission in the central nervous system. Functional AMPARs are tetrameric complexes with a highly modular structure, consisting of four evolutionarily distinct structural domains: an amino-terminal domain (ATD), a ligand-binding domain (LBD), a channel-forming transmembrane domain (TMD), and a carboxyl-terminal domain (CTD). Here we show that the isolated TMD of the GluA1 AMPAR is fully capable of tetramerization. Additionally, removal of the extracellular domains from the receptor did not affect membrane topology or surface delivery. Furthermore, whereas the ATD and CTD contribute positively to tetramerization, the LBD presents a barrier to the process by reducing the stability of the receptor complex. These experiments pinpoint the TMD as the "tetramerization domain" for AMPARs, with other domains playing modulatory roles. They also raise intriguing questions about the evolution of iGluRs as well as the mechanisms regulating the biogenesis of AMPAR complexes., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)

AMPA and NMDA receptors are glutamate-gated ion channels that mediate fast excitatory synaptic transmission throughout the nervous system. In the continual presence of glutamate, AMPA and NMDA receptors containing the GluN2A or GluN2B subunit enter into a nonconducting, desensitized state that can impact synaptic responses and glutamate-mediated excitotoxicity. The process of desensitization is dramatically different between subtypes, but the basis for these differences is unknown. We generated an extensive sequence alignment of ionotropic glutamate receptors (iGluRs) from diverse animal phyla and identified a highly conserved motif, which we termed the "hydrophobic box," located at the extracellular interface of transmembrane helices. A single position in the hydrophobic box differed between mammalian AMPA and NMDA receptors. Surprisingly, we find that an NMDAR-to-AMPAR exchange mutation at this position in the rat GluN2A or GluN2B subunit had a dramatic and highly specific effect on NMDAR desensitization, making it AMPAR-like. In contrast, a reverse exchange mutation in AMPARs had minimal effects on desensitization. These experiments highlight differences in desensitization between iGluR subtypes and the highly specific contribution of the GluN2 subunit to this process., Significance Statement: Rapid communication between cells in the nervous system depends on ion channels that are directly activated by neurotransmitter molecules. Here, we studied ionotropic glutamate receptors (iGluRs), which are ion channels activated by the neurotransmitter glutamate. By comparing the sequences of a vast number of iGluR proteins from diverse animal species, assisted by available structural information, we identified a highly conserved motif. We showed that a single amino acid difference in this motif between mammalian iGluR subtypes has dramatic effects on receptor function. These results have implications in both the evolution of synaptic function, as well as the role of iGluRs in health and disease., (Copyright © 2016 the authors 0270-6474/16/362617-06$15.00/0.)

We present a mathematical model for ionotropic glutamate receptors (iGluR's) that is built on mechanistic understanding and yields a number of thermodynamic and kinetic properties of channel gating. iGluR's are ligand-gated ion channels responsible for the vast majority of fast excitatory neurotransmission in the central nervous system. The effects of agonist-induced closure of the ligand-binding domain (LBD) are transmitted to the transmembrane channel (TMC) via interdomain linkers. Our model demonstrates that, relative to full agonists, partial agonists may reduce either the degree of LBD closure or the curvature of the LBD free energy basin, leading to less stabilization of the channel open state and hence lower channel open probability. A rigorous relation is derived between the channel closed-to-open free energy difference and the tension within the linker. Finally, by treating LBD closure and TMC opening as diffusive motions, we obtain gating trajectories that resemble stochastic current traces from single-channel recordings and calculate the rate constants for transitions between the channel open and closed states. Our model can be implemented by molecular dynamics simulations to realistically depict iGluR gating and may guide functional experiments in gaining deeper insight into this essential family of channel proteins.

AMPA receptors (AMPARs) play a critical role in excitatory glutamatergic neurotransmission. The number and subunit composition of AMPARs at synapses determines the dynamics of fast glutamatergic signalling. Functional AMPARs on the cell surface are tetramers. Thus tetrameric assembly of AMPARs represents a promising target for modulating AMPAR-mediated signalling in health and disease. Multiple structural domains within the receptor influence AMPAR assembly. In a proposed model for AMPAR assembly, the amino-terminal domain underlies the formation of a dimer pool. The transmembrane domain facilitates the formation and enhances the stability of the tetramer. The ligand-binding domain influences assembly through a process referred to as 'domain swapping'. We propose that this core AMPAR assembly process could be regulated by neuronal signals and speculate on possible mechanisms for such regulation., (© 2014 The Authors. The Journal of Physiology © 2014 The Physiological Society.)

The fidelity of integration of pre- and postsynaptic activity by NMDA receptors (NMDARs) requires a match between agonist binding and ion channel opening. To address how agonist binding is transduced into pore opening in NMDARs, we manipulated the coupling between the ligand-binding domain (LBD) and the ion channel by inserting residues in a linker between them. We found that a single residue insertion markedly attenuated the ability of NMDARs to convert a glutamate transient into a functional response. This was largely a result of a decreased likelihood of the channel opening and remaining open. Computational and thermodynamic analyses suggest that insertions prevent the agonist-bound LBD from effectively pulling on pore lining elements, thereby destabilizing pore opening. Furthermore, this pulling energy was more prominent in the GluN2 subunit. We conclude that an efficient NMDAR-mediated synaptic response relies on a mechanical coupling between the LBD and the ion channel.

The NMDA-sensitive glutamate receptor is a ligand-gated ion channel that mediates excitatory synaptic transmission in the nervous system. Extracellular zinc allosterically regulates the NMDA receptor by binding to the extracellular N-terminal domain, which inhibits channel gating. Phosphorylation of the intrinsically disordered intracellular C-terminal domain alleviates inhibition by extracellular zinc. The mechanism for this functional effect is largely unknown. Proline is a hallmark of intrinsic disorder, so we used proline mutagenesis to modulate disorder in the cytoplasmic domain. Proline depletion selectively uncoupled zinc inhibition with little effect on receptor biogenesis, surface trafficking, or ligand-activated gating. Proline depletion also reduced the affinity for a PDZ domain involved in synaptic trafficking and affected small molecule binding. To understand the origin of these phenomena, we used single molecule fluorescence and ensemble biophysical methods to characterize the structural effects of proline mutagenesis. Proline depletion did not eliminate intrinsic disorder, but the underlying conformational dynamics were changed. Thus, we altered the form of intrinsic disorder, which appears sufficient to affect the biological activity. These findings suggest that conformational dynamics within the intrinsically disordered cytoplasmic domain are important for the allosteric regulation of NMDA receptor gating.

Fast-spiking parvalbumin (PV)-positive interneurons in layers 2/3 of the visual cortex regulate gain control and tuning of visual processing. Synapse-associated protein 97 (SAP97) belongs to a family of proteins that have been implicated in regulating glutamatergic synaptic transmission at pyramidal-to-pyramidal connections in the nervous system. For PV interneurons in mouse visual cortex, the expression of SAP97 is developmentally regulated, being expressed in almost all juvenile but only a fraction, ~40%, of adult PV interneurons. Using whole-cell patch-clamping, single-cell RT-PCR to assay endogenous expression of SAP97 and exogenous expression of SAP97, we investigated the functional significance of SAP97 in PV interneurons in layers 2/3 of the visual cortex. PV interneurons expressing SAP97, either endogenously or via exogenous expression, showed distinct membrane properties from those not expressing SAP97. This included an overall decrease in membrane excitability, as indexed by a decrease in membrane resistance and an increase in the stimulus threshold for the first action potential firing. Additionally, SAP97-expressing PV interneurons fired action potentials more frequently and, at moderate stimulus intensities, showed irregular or stuttering firing patterns. Furthermore, SAP97-expressing PV interneurons showed increased glutamatergic input and more extensive dendritic branching when compared with non-expressing PV interneurons. These differences in membrane and synaptic properties would significantly alter how PV interneurons expressing SAP97 compared with those not expressing SAP97 would function in local networks. Thus, our results indicate that the scaffolding protein SAP97 is a critical molecular factor regulating the input-output relationships of cortical PV interneurons.

Glutamate-gated ion channels embedded within the neuronal membrane are the primary mediators of fast excitatory synaptic transmission in the CNS. The ion channel of these glutamate receptors contains a pore-lining transmembrane M3 helix surrounded by peripheral M1 and M4 helices. In the NMDA receptor subtype, opening of the ion channel pore, mediated by displacement of the M3 helices away from the central pore axis, occurs in a highly concerted fashion, but the associated temporal movements of the peripheral helices are unknown. To address the gating dynamics of the peripheral helices, we constrained the relative movements of the linkers that connect these helices to the ligand-binding domain using engineered cross-links, either within (intra-GluN1 or GluN2A) or between subunits. Constraining the peripheral linkers in any manner dramatically curtailed channel opening, highlighting the requirement for rearrangements of these peripheral structural elements for efficient gating to occur. However, the magnitude of this gating effect depended on the specific subunit being constrained, with the most dramatic effects occurring when the constraint was between subunits. Based on kinetic and thermodynamic analysis, our results suggest an asynchrony in the displacement of the peripheral linkers during the conformational and energetic changes leading to pore opening. Initially there are large-scale rearrangements occurring between the four subunits. Subsequently, rearrangements occur within individual subunits, mainly GluN2A, leading up to or in concert with pore opening. Thus, the conformational changes induced by agonist binding in NMDA receptors converge asynchronously to permit pore opening.

Most fast excitatory synaptic transmission in the nervous system is mediated by glutamate acting through ionotropic glutamate receptors (iGluRs). iGluRs (AMPA, kainate, and NMDA receptor subtypes) are tetrameric assemblies, formed as a dimer of dimers. Still, the mechanism underlying tetramerization--the necessary step for the formation of functional receptors that can be inserted into the plasma membrane--is unknown. All eukaryotic compared to prokaryotic iGluR subunits have an additional transmembrane segment, the M4 segment, which positions the physiologically critical C-terminal domain on the cytoplasmic side of the membrane. AMPA receptor (AMPAR) subunits lacking M4 do not express on the plasma membrane. Here, we show that these constructs are retained in the endoplasmic reticulum, the major cellular compartment mediating protein oligomerization. Using approaches to assay the native oligomeric state of AMPAR subunits, we find that subunits lacking M4 or containing single amino acid substitutions along an "interacting" face of the M4 helix that block surface expression no longer tetramerize in either homomeric or heteromeric assemblies. In contrast, subunit dimerization appears to be largely intact. These experiments define the M4 segment as a unique functional unit in AMPARs that is required for the critical dimer-to-tetramer transition.

The input, processing, and output characteristics of inhibitory interneurons help shape information flow through layers 2/3 of the visual cortex. Parvalbumin (PV)-positive interneurons modulate and synchronize the gain and dynamic responsiveness of pyramidal neurons. To define the diversity of PV interneurons in layers 2/3 of the developing visual cortex, we characterized their passive and active membrane properties. Using Ward's and k-means multidimensional clustering, we identified four PV interneuron subgroups. The most notable difference between the subgroups was their firing patterns in response to moderate stimuli just above rheobase. Two subgroups showed regular and continuous firing at all stimulus intensities above rheobase. The difference between these two continuously firing subgroups was that one fired at much higher frequencies and transitioned into this high-frequency firing rate at or near rheobase. The two other subgroups showed irregular, stuttering firing patterns just above rheobase. Both of these subgroups typically transitioned to regular and continuous firing at intense stimulations, but one of these subgroups, the strongly stuttering subgroup, showed irregular firing across a wider range of stimulus intensities and firing frequencies. The four subgroups also differed in excitatory synaptic input, providing independent support for the classification of subgroups. The subgroups of PV interneurons identified here would respond differently to inputs of varying intensity and frequency, generating diverse patterns of PV inhibition in the developing neural circuit.

Microglia are the immunocompetent cells of the central nervous system. In the physiological setting, their highly motile processes continually survey the local brain parenchyma and transiently contact synaptic elements. Although recent work has shown that the interaction of microglia with synapses contributes to synaptic remodeling during development, the role of microglia in synaptic physiology is just starting to get explored. To assess this question, we employed an electrophysiological approach using two methods to manipulate microglia in culture: organotypic hippocampal brain slices in which microglia were depleted using clodronate liposomes, and cultured hippocampal neurons to which microglia were added. We show here that the frequency of excitatory postsynaptic current increases in microglia-depleted brain slices, consistent with a higher synaptic density, and that this enhancement ensures from the loss of microglia since it is reversed when the microglia are replenished. Conversely, the addition of microglia to neuronal cultures decreases synaptic activity and reduces the density of synapses, spine numbers, surface expression of AMPA receptor (GluA1), and levels of synaptic adhesion molecules. Taken together, our findings demonstrate that non-activated microglia acutely modulate synaptic activity by regulating the number of functional synapses in the central nervous system.

Excitatory synapses that use the neurotransmitter glutamate are highly dynamic, constantly changing their character in an activity-dependent manner. In this issue of Neuron, Penn et al. (2012) describe a novel mechanism that changes the fidelity of glutamate signaling to maintain homeostatic synaptic plasticity., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Ionotropic glutamate receptors (GluRs) are ligand-gated ion channels with a modular structure. The ion channel itself shares structural similarity, albeit an inverted membrane topology, with P-loop channels. Like P-loop channels, prokaryotic GluR subunits (e.g. GluR0) have two transmembrane segments. In contrast, eukaryotic GluRs have an additional transmembrane segment (M4), located C-terminal to the ion channel core. However, the structural/functional significance of this additional transmembrane segment is poorly defined. Although topologically similar to GluR0, mammalian AMPA receptor (GluA1) subunits lacking the M4 segment do not display surface expression. This lack of expression is not due to the M4 segment serving as an anchor to the ligand-binding domain because insertion of an artificial polyleucine transmembrane segment does not rescue surface expression. Specific interactions between M4 and the ligand-binding domain are also unlikely because insertion of polyglycines into the linker connecting them has no deleterious effects on function or surface expression. However, tryptophan and cysteine scanning mutagenesis of the M4 segment, as well as recovery of function in the polyleucine background, defined a unique face of the M4 helix that is required for GluR surface expression. In the AMPA receptor structure, this face forms intersubunit contacts with the transmembrane helices of the ion channel core (M1 and M3) from another subunit within the homotetramer. Thus, our experiments show that a highly specific interaction of the M4 segment with an adjacent subunit is required for surface expression of AMPA receptors. This interaction may represent a mechanism for regulating AMPA receptor biogenesis.

NMDA receptors are glutamate-activated ion channel complexes central to the functioning of the mammalian nervous system. Opening of the NMDA receptor ion channel pore is initiated by agonist-induced conformational changes in the extracellular ligand-binding domain (LBD) but the dynamic mechanism of this process remains unresolved. We studied how a disulfide bond in the obligatory GluN1 subunit-the sole site of redox modulation in NMDA receptors-controls this activation gating mechanism. This disulfide bond is located in the hinge region of the LBD, and presumably constrains agonist-induced cleft closure of the clamshell-like LBD. Elimination of this bond, by either DTT-mediated reduction or mutagenesis, enhances gating efficiency such that pore opening now occurs with higher frequency and longer duration. The most prominent effect was to shift opening modes to long duration openings reminiscent of a high P(o) gating mode that the NMDA receptor exhibits under ambient oxidizing conditions. In terms of preopen gating steps, elimination of this bond has effects only on the fast gating step consistent with this step being GluN1-specific and reflecting GluN1 gating movements immediately before channel opening. Overall, our results suggest that the dynamics of the GluN1 LBD have strong effects on late pore opening steps including regulating the duration of pore opening. This redox-mediated gating modulation could be an underlying mechanism of NMDA receptor malfunction in redox-dependent disease states and presents a potential target of pharmacologic action., (Copyright © 2011 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

NMDA receptors are ligand-gated ion channels with a regulatory intracellular C-terminal domain (CTD). In GluN2B, the CTD is the largest domain in the protein but is intrinsically disordered. The GluN2B subunit is the major tyrosine-phosphorylated protein in synapses. Src kinase phosphorylates the GluN2B CTD, but it is unknown how this affects channel activity. In disordered proteins, phosphorylation can tip the balance between order and disorder. Transitions can occur in both directions, so it is not currently possible to predict the effects of phosphorylation. We used single molecule fluorescence to characterize the effects of Src phosphorylation on GluN2B. Scanning fluorescent labeling sites throughout the domain showed no positional dependence of the energy transfer. Instead, efficiency only scaled with the separation between labeling sites suggestive of a relatively featureless conformational energy landscape. Src phosphorylation led to a general expansion of the polypeptide, which would result in greater exposure of known protein-binding sites and increase the physical separation between contiguous sites. Phosphorylation makes the CTD more like a random coil leaving open the question of how Src exerts its effects on the NMDA receptor.

Ionotropic glutamate receptors (iGluRs), including the NMDA receptor subtype, are ligand-gated ion channels critical to fast signaling in the CNS. NMDA receptors are obligate heterotetramers composed of two GluN1 and typically two GluN2 subunits. However, the arrangement of GluN subunits in functional receptors-whether like subunits are adjacent to (N1/N1/N2/N2) or diagonal to (N1/N2/N1/N2) one another-remains unclear. Recently, a crystal structure of a homomeric AMPA receptor revealed that the four identical subunits adopt two distinct and subunit-specific conformations termed A/C and B/D with subunits of like conformations (e.g., A/C) diagonal to one another. In the structure, the two conformers were notable at the level of the linkers (S1-M1, M3-S2, and S2-M4) that join the ligand-binding domain to the transmembrane ion channel with the M3-S2 linker positioned more proximal to the central axis of the channel pore in the A/C conformation and S2-M4 more proximal in the B/D conformation. Using immunoblots and functional assays, we show that introduced cysteines in the M3/M3-S2 linker of GluN1, but not GluN2, show dimer formation and oxidation-induced changes in current amplitudes predictive of the A/C conformation. Conversely, introduced cysteines in the S2-M4 linker of GluN2, but not GluN1, showed similar functional effects, suggesting that the GluN2 subunit adopts the B/D conformation. Thus, we show that NMDA receptors, like AMPA receptors, possess distinct subunit-specific conformations with GluN1 approximating the A/C and GluN2 the B/D conformation. GluN subunits are therefore positioned in a N1/N2/N1/N2 arrangement in functional NMDA receptors.

The defining functional feature of N-methyl-d-aspartate (NMDA) receptors is activation gating, the energetic coupling of ligand binding into opening of the associated ion channel pore. NMDA receptors are obligate heterotetramers typically composed of glycine-binding GluN1 and glutamate-binding GluN2 subunits that gate in a concerted fashion, requiring all four ligands to bind for subsequent opening of the channel pore. In an individual subunit, the extracellular ligand-binding domain, composed of discontinuous polypeptide segments S1 and S2, and the transmembrane channel-forming domain, composed of M1-M4 segments, are connected by three linkers: S1-M1, M3-S2, and S2-M4. To study subunit-specific events during pore opening in NMDA receptors, we impaired activation gating via intrasubunit disulfide bonds connecting the M3-S2 and S2-M4 in either the GluN1 or GluN2A subunit, thereby interfering with the movement of the M3 segment, the major pore-lining and channel-gating element. NMDA receptors with gating impairments in either the GluN1 or GluN2A subunit were dramatically resistant to channel opening, but when they did open, they showed only a single-conductance level indistinguishable from wild type. Importantly, the late gating steps comprising pore opening to its main long-duration open state were equivalently affected regardless of which subunit was constrained. Thus, the NMDA receptor ion channel undergoes a pore-opening mechanism in which the intrasubunit conformational dynamics at the level of the ligand-binding/transmembrane domain (TMD) linkers are tightly coupled across the four subunits. Our results further indicate that conformational freedom of the linkers between the ligand-binding and TMDs is critical to the activation gating process.

GABAergic interneurons are key elements regulating the activity of local circuits, and abnormal inhibitory circuits are implicated in certain psychiatric and neurodevelopmental diseases. The glutamatergic input that interneurons receive is a key determinant of their activity, yet its molecular structure and development, which are often distinct from those of glutamatergic input to pyramidal cells, are poorly defined. The membrane-associated guanylate kinase (MAGUK) homologs PSD-95/SAP90, PSD-93/chapsyn110, SAP97, and SAP102 are central organizers of the postsynaptic density at excitatory synapses on pyramidal neurons. We therefore studied the cell-type-specific and developmental expression of MAGUKs in the nonoverlapping parvalbumin (PV)- and somatostatin (SOM)-positive interneurons in the visual cortex. These interneuron subtypes account for the vast majority of interneurons in the cortex and have different functional properties and postsynaptic structures, being either axodendritic (PV(+)) or axospinous (SOM(+)). To study cell-type-specific MAGUK expression, we used DIG-labeled riboprobes against each MAGUK along with antibodies against either PV or SOM and examined tissue from juvenile (P15) and adult mice. Both PV(+) and SOM(+) interneurons express mRNA for PSD-95, PSD-93, and SAP102 in P15 and adult tissue. In contrast, these interneuron subtypes express SAP97 at P15, but for adult visual cortex we found that most PV(+) and SOM(+) interneurons show low or no expression of SAP97. Given the importance of SAP97 in regulating AMPA receptor GluA1 subunit and NMDA receptor subunits at glutamatergic synapses, these results suggest a developmental shift in glutamate receptor subunit composition and regulation of glutamatergic synapses on PV(+) and SOM(+) interneurons., (© 2010 Wiley-Liss, Inc.)

The mammalian ionotropic glutamate receptor family encodes 18 gene products that coassemble to form ligand-gated ion channels containing an agonist recognition site, a transmembrane ion permeation pathway, and gating elements that couple agonist-induced conformational changes to the opening or closing of the permeation pore. Glutamate receptors mediate fast excitatory synaptic transmission in the central nervous system and are localized on neuronal and non-neuronal cells. These receptors regulate a broad spectrum of processes in the brain, spinal cord, retina, and peripheral nervous system. Glutamate receptors are postulated to play important roles in numerous neurological diseases and have attracted intense scrutiny. The description of glutamate receptor structure, including its transmembrane elements, reveals a complex assembly of multiple semiautonomous extracellular domains linked to a pore-forming element with striking resemblance to an inverted potassium channel. In this review we discuss International Union of Basic and Clinical Pharmacology glutamate receptor nomenclature, structure, assembly, accessory subunits, interacting proteins, gene expression and translation, post-translational modifications, agonist and antagonist pharmacology, allosteric modulation, mechanisms of gating and permeation, roles in normal physiological function, as well as the potential therapeutic use of pharmacological agents acting at glutamate receptors.