Topiramate Versus Naltrexone for Alcohol Use Disorder: A Genotype-Stratified Double-Blind Randomized Controlled Trial.

OBJECTIVE: There have been no well-controlled and well-powered comparative trials of topiramate with other pharmacotherapies for alcohol use disorder (AUD), such as naltrexone. Moreover, the literature is mixed on the effects of two polymorphisms-rs2832407 (in GRIK1) and rs1799971 (in OPRM1)-on response to topiramate and naltrexone, respectively. The authors sought to examine the comparative effectiveness of topiramate and naltrexone in improving outcomes in AUD and to examine the role of the rs2832407 and rs1799971 polymorphisms, respectively, on response to these medications. METHODS: In a 12-week, double-blind, placebo-controlled, randomized, multisite, genotype-stratified (rs2832407 and rs1799971) clinical trial comparing topiramate and naltrexone in treating AUD, 147 patients with AUD were randomly assigned to treatment with topiramate or naltrexone, stratified by genotype (rs2832407*CC and *AC/AA genotypes and rs1799971*AA and *AG/GG genotypes). The predefined primary outcome was number of heavy drinking days per week. Predefined secondary outcomes included standard drinks per drinking day per week, body mass index (BMI), craving, markers of liver injury, mood, and adverse events. RESULTS: For the number of heavy drinking days per week, there was a near-significant time-by-treatment interaction. For the number of standard drinks per drinking day per week, there was a significant time-by-treatment interaction, which favored topiramate. There were significant time-by-treatment effects, with greater reductions observed with topiramate than naltrexone for BMI, craving, and gamma-glutamyltransferase level. Withdrawal due to side effects occurred in 8% and 5% of the topiramate and naltrexone groups, respectively. Neither polymorphism showed an effect on treatment response. CONCLUSIONS: Topiramate is at least as effective and safe as the first-line medication, naltrexone, in reducing heavy alcohol consumption, and superior in reducing some clinical outcomes. Neither rs2832407 nor rs1799971 had effects on topiramate and naltrexone treatments, respectively.

The kainate receptor GluK2 mediates cold sensing in mice.

Thermosensors expressed in peripheral somatosensory neurons sense a wide range of environmental temperatures. While thermosensors detecting cool, warm and hot temperatures have all been extensively characterized, little is known about those sensing cold temperatures. Though several candidate cold sensors have been proposed, none has been demonstrated to mediate cold sensing in somatosensory neurons in vivo, leaving a knowledge gap in thermosensation. Here we characterized mice lacking the kainate-type glutamate receptor GluK2, a mammalian homolog of the Caenorhabditis elegans cold sensor GLR-3. While GluK2 knockout mice respond normally to heat and mechanical stimuli, they exhibit a specific deficit in sensing cold but not cool temperatures. Further analysis supports a key role for GluK2 in sensing cold temperatures in somatosensory DRG neurons in the periphery. Our results reveal that GluK2-a glutamate-sensing chemoreceptor mediating synaptic transmission in the central nervous system-is co-opted as a cold-sensing thermoreceptor in the periphery.

Tritium and deuterium labelling of a kainate receptor antagonist and evaluation as a radioligand.

Kainate receptors play a crucial role in mediating synaptic transmission within the central nervous system. However, the lack of selective pharmacological tool compounds for the GluK3 subunit represents a significant challenge in studying these receptors. Recently presented compound 1 stands out as a potent antagonist of GluK3 receptors, exhibiting nanomolar affinity at GluK3 receptors and strongly inhibiting glutamate-induced currents at homomeric GluK1 and GluK3 receptors in HEK293 cells with Kb values of 65 and 39 nM, respectively. This study presents the synthesis of two potent GluK3-preferring iodine derivatives of compound 1, serving as precursors for radiolabelling. Furthermore, we demonstrate the optimisation of dehalogenation conditions using hydrogen and deuterium, resulting in [2H]-1, and demonstrate the efficient synthesis of the radioligand [3H]-1 with a specific activity of 1.48 TBq/mmol (40.1 Ci/mmol). Radioligand binding studies conducted with [3H]-1 as a radiotracer at GluK1, GluK2, and GluK3 receptors expressed in Sf9 and rat P2 membranes demonstrated its potential applicability for selectively studying native GluK3 receptors in the presence of GluK1 and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor-blocking ligands.

The positive allosteric modulator BPAM344 and L-glutamate introduce an active-like structure of the ligand-binding domain of GluK2.

Kainate receptors belong to the family of ionotropic glutamate receptors and contribute to the majority of fast excitatory neurotransmission. Consequently, they also play a role in brain diseases. Therefore, understanding how these receptors can be modulated is of importance. Our study provides a crystal structure of the dimeric ligand-binding domain of the kainate receptor GluK2 in complex with L-glutamate and the small-molecule positive allosteric modulator, BPAM344, in an active-like conformation. The role of Thr535 and Gln786 in modulating GluK2 by BPAM344 was investigated using a calcium-sensitive fluorescence-based assay on transiently transfected cells expressing GluK2 and mutants hereof. This study may aid in the design of compounds targeting kainate receptors, expanding their potential as targets for the treatment of brain diseases.

Behavioral analysis of kainate receptor KO mice and the role of GluK3 subunit in anxiety.

Kainate receptors (KARs) are one of the ionotropic glutamate receptors in the central nervous system (CNS) comprised of five subunits, GluK1-GluK5. There is a growing interest in the association between KARs and psychiatric disorders, and there have been several studies investigating the behavioral phenotypes of KAR deficient mice, however, the difference in the genetic background has been found to affect phenotype in multiple mouse models of human diseases. Here, we examined GluK1-5 single KO mice in a pure C57BL/6N background and identified that GluK3 KO mice specifically express anxiolytic-like behavior with an alteration in dopamine D2 receptor (D2R)-induced anxiety, and reduced D2R expression in the striatum. Biochemical studies in the mouse cortex confirmed that GluK3 subunits do not assemble with GluK4 and GluK5 subunits, that can be activated by lower concentration of agonists. Overall, we found that GluK3-containing KARs function to express anxiety, which may represent promising anti-anxiety medication targets.

Reversing Aging: Decline in Complex Olfactory Learning Can be Rectified by Restoring Intrinsic Plasticity of Hippocampal CA1 Pyramidal Neurons.

The acquisition of complex rules requires modifications in intrinsic plasticity of excitatory neurons within relevant brain areas. Olfactory discrimination (OD) rule learning occludes slow calcium-dependent potassium current (sIAHP ) in piriform cortex (PC) pyramidal neurons, which increases their intrinsic neuronal excitability. Similar learning-induced sIAHP changes are demonstrated in hippocampal CA1. The shutdown of sIAHP is mediated by the metabotropic activation of the kainate subtype glutamatergic receptor, GluK2. Here, the duration of training required for OD rule learning increased significantly as the mice matured and aged is first shown, which appears earlier in 5xFAD mice. At the cellular biophysical level, aging is accompanied by reduction in the post-burst AHP in these neurons, while neuronal excitability remains stable. This is in contrast to aging CA1 neurons that exhibit enhanced post-burst AHPs in previous reports. Kainate reduces post-burst AHP in adults, but not in aged PC neurons, whereas it reduces post-burst AHPs in hippocampal CA1 pyramidal neurons of both young and aged mice. Overexpression of GluK2 in CA1 neurons restores OD learning capabilities in aged wild-type and 5xFAD mice, to a level comparable to young adults. Activation of GluK2 receptors in selectively vulnerable neurons can prevent aging-related cognitive decline is suggested.

Small-molecule positive allosteric modulation of homomeric kainate receptors GluK1-3: development of screening assays and insight into GluK3 structure.

The kainate receptors GluK1-3 (glutamate receptor ionotropic, kainate receptors 1-3) belong to the family of ionotropic glutamate receptors and are essential for fast excitatory neurotransmission in the brain, and are associated with neurological and psychiatric diseases. How these receptors can be modulated by small-molecule agents is not well understood, especially for GluK3. We show that the positive allosteric modulator BPAM344 can be used to establish robust calcium-sensitive fluorescence-based assays to test agonists, antagonists, and positive allosteric modulators of GluK1-3. The half-maximal effective concentration (EC50) of BPAM344 for potentiating the response of 100 µm kainate was determined to be 26.3 µm for GluK1, 75.4 µm for GluK2, and 639 µm for GluK3. Domoate was found to be a potent agonist for GluK1 and GluK2, with an EC50 of 0.77 and 1.33 µm, respectively, upon co-application of 150 µm BPAM344. At GluK3, domoate acts as a very weak agonist or antagonist with a half-maximal inhibitory concentration (IC50) of 14.5 µm, in presence of 500 µm BPAM344 and 100 µm kainate for competition binding. Using H523A-mutated GluK3, we determined the first dimeric structure of the ligand-binding domain by X-ray crystallography, allowing location of BPAM344, as well as zinc-, sodium-, and chloride-ion binding sites at the dimer interface. Molecular dynamics simulations support the stability of the ion sites as well as the involvement of Asp761, Asp790, and Glu797 in the binding of zinc ions. Using electron microscopy, we show that, in presence of glutamate and BPAM344, full-length GluK3 adopts a dimer-of-dimers arrangement.

Exploring thienothiadiazine dioxides as isosteric analogues of benzo- and pyridothiadiazine dioxides in the search of new AMPA and kainate receptor positive allosteric modulators.

The synthesis and biological evaluation on AMPA and kainate receptors of new examples of 3,4-dihydro-2H-1,2,4-thieno[3,2-e]-1,2,4-thiadiazine 1,1-dioxides is described. The introduction of a cyclopropyl chain instead of an ethyl chain at the 4-position of the thiadiazine ring was found to dramatically improve the potentiator activity on AMPA receptors, with compound 32 (BPAM395) expressing in vitro activity on AMPARs (EC2x = 0.24 µM) close to that of the reference 4-cyclopropyl-substituted benzothiadiazine dioxide 10 (BPAM344). Interestingly, the 4-allyl-substituted thienothiadiazine dioxide 27 (BPAM307) emerged as the most promising compound on kainate receptors being a more effective potentiator than the 4-cyclopropyl-substituted thienothiadiazine dioxide 32 and supporting the view that the 4-allyl substitution of the thiadiazine ring could be more favorable than the 4-cyclopropyl substitution to induce marked activity on kainate receptors versus AMPA receptors. The thieno-analogue 36 (BPAM279) of the clinically tested S18986 (11) was selected for in vivo evaluation in mice as a cognitive enhancer due to a safer profile than 32 after massive per os drug administration. Compound 36 was found to increase the cognition performance in mice at low doses (1 mg/kg) per os suggesting that the compound was well absorbed after oral administration and able to reach the central nervous system. Finally, compound 32 was selected for co-crystallization with the GluA2-LBD (L504Y,N775S) and glutamate to examine the binding mode of thienothiadiazine dioxides within the allosteric binding site of the AMPA receptor. At the allosteric site, this compound established similar interactions as the previously reported BTD-type AMPA receptor modulators.

Enhanced Membrane Incorporation of H289Y Mutant GluK1 Receptors from the Audiogenic Seizure-Prone GASH/Sal Model: Functional and Morphological Impacts on Xenopus Oocytes.

Epilepsy is a neurological disorder characterized by abnormal neuronal excitability, with glutamate playing a key role as the predominant excitatory neurotransmitter involved in seizures. Animal models of epilepsy are crucial in advancing epilepsy research by faithfully replicating the diverse symptoms of this disorder. In particular, the GASH/Sal (genetically audiogenic seizure-prone hamster from Salamanca) model exhibits seizures resembling human generalized tonic-clonic convulsions. A single nucleotide polymorphism (SNP; C9586732T, p.His289Tyr) in the Grik1 gene (which encodes the kainate receptor GluK1) has been previously identified in this strain. The H289Y mutation affects the amino-terminal domain of GluK1, which is related to the subunit assembly and trafficking. We used confocal microscopy in Xenopus oocytes to investigate how the H289Y mutation, compared to the wild type (WT), affects the expression and cell-surface trafficking of GluK1 receptors. Additionally, we employed the two-electrode voltage-clamp technique to examine the functional effects of the H289Y mutation. Our results indicate that this mutation increases the expression and incorporation of GluK1 receptors into an oocyte's membrane, enhancing kainate-evoked currents, without affecting their functional properties. Although further research is needed to fully understand the molecular mechanisms responsible for this epilepsy, the H289Y mutation in GluK1 may be part of the molecular basis underlying the seizure-prone circuitry in the GASH/Sal model.

A Pathogenic Missense Mutation in Kainate Receptors Elevates Dendritic Excitability and Synaptic Integration through Dysregulation of SK Channels.

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.SIGNIFICANCE STATEMENT Damaging mutations in genes encoding synaptic proteins have been identified in various neurodevelopmental disorders, but the functional consequences at the cellular and circuit level remain elusive. By generating a novel knock-in mutant mouse, this study examined the role of a pathogenic mutation in the GluK2 kainate receptor (KAR) subunit, a subclass of ionotropic glutamate receptors. Analyses of hippocampal CA3 pyramidal neurons determined elevated action potential firing because of an increase in dendritic excitability. Increased dendritic excitability was attributable to reduced activity of a Ca2+ activated K+ channel. These results indicate that a pathogenic KAR mutation results in dysregulation of dendritic K+ channels, which leads to an increase in synaptic integration and backpropagation of action potentials into distal dendrites.

Presynaptic Kainate Receptors onto Somatostatin Interneurons Are Recruited by Activity throughout Development and Contribute to Cortical Sensory Adaptation.

Somatostatin (SST) interneurons produce delayed inhibition because of the short-term facilitation of their excitatory inputs created by the expression of metabotropic glutamate receptor 7 (mGluR7) and presynaptic GluK2-containing kainate receptors (GluK2-KARs). Using mice of both sexes, we find that as synaptic facilitation at layer (L)2/3 SST cell inputs increases during the first few postnatal weeks, so does GluK2-KAR expression. Removal of sensory input by whisker trimming does not affect mGluR7 but prevents the emergence of presynaptic GluK2-KARs, which can be restored by allowing whisker regrowth or by acute calmodulin activation. Conversely, late trimming or acute inhibition of Ca2+/calmodulin-dependent protein kinase II is sufficient to reduce GluK2-KAR activity. This developmental and activity-dependent regulation also produces a specific reduction of L4 GluK2-KARs that advances in parallel with the maturation of sensory processing in L2/3. Finally, we find that removal of both GluK2-KARs and mGluR7 from the synapse eliminates short-term facilitation and reduces sensory adaptation to repetitive stimuli, first in L4 of somatosensory cortex, then later in development in L2/3. The dynamic regulation of presynaptic GluK2-KARs potentially allows for flexible scaling of late inhibition and sensory adaptation.SIGNIFICANCE STATEMENT Excitatory synapses onto somatostatin (SST) interneurons express presynaptic, calcium-permeable kainate receptors containing the GluK2 subunit (GluK2-KARs), activated by high-frequency activity. In this study we find that their presence on L2/3 SST synapses in the barrel cortex is not based on a hardwired genetic program but instead is regulated by sensory activity, in contrast to that of mGluR7. Thus, in addition to standard synaptic potentiation and depression mechanisms, excitatory synapses onto SST neurons undergo an activity-dependent presynaptic modulation that uses GluK2-KARs. Further, we present evidence that loss of the frequency-dependent synaptic components (both GluK2-KARs and mGluR7 via Elfn1 deletion) contributes to a decrease in the sensory adaptation commonly seen on repetitive stimulus presentation.

Progressive development of synchronous activity in the hippocampal neuronal network is modulated by GluK1 kainate receptors.

Kainate receptors are potent modulators of circuit excitability and have been repeatedly implicated in pathophysiological synchronization of limbic networks. While the role of aberrant GluK2 subunit containing KARs in generation of epileptiform hypersynchronous activity is well described, the contribution of other KAR subtypes, including GluK1 subunit containing KARs remain less well understood. To investigate the contribution of GluK1 KARs in developmental and pathological synchronization of the hippocampal neural network, we used multielectrode array recordings on organotypic hippocampal slices that display first multi-unit activity and later spontaneous population discharges resembling ictal-like epileptiform activity (IEA). Chronic blockage of GluK1 activity using selective antagonist ACET or lentivirally delivered shRNA significantly delayed developmental synchronization of the hippocampal CA3 network and generation of IEA. GluK1 overexpression, on the other hand, had no significant effect on occurrence of IEA, but enhanced the size of the neuron population participating in the population discharges. Correlation analysis indicated that local knockdown of GluK1 locally in the CA3 neurons reduced their functional connectivity, while GluK1 overexpression increased the connectivity to both CA1 and DG. These data suggest that GluK1 KARs regulate functional connectivity between the excitatory neurons, possibly via morphological changes in glutamatergic circuit, affecting synchronization of neuronal populations. The significant effects of GluK1 manipulations on network activity call for further research on GluK1 KAR as potential targets for antiepileptic treatments, particularly during the early postnatal development when GluK1 KARs are strongly expressed in the limbic neural networks.

Depressive-like Behaviors Induced by mGluR5 Reduction in 6xTg in Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is one representative dementia characterized by the accumulation of amyloid beta (Aß) plaques and neurofibrillary tangles (NFTs) in the brain, resulting in cognitive decline and memory loss. AD is associated with neuropsychiatric symptoms, including major depressive disorder (MDD). Recent studies showed a reduction in mGluR5 expression in the brains of stress-induced mice models and individuals with MDD compared to controls. In our study, we identified depressive-like behavior and memory impairment in a mouse model of AD, specifically in the 6xTg model with tau and Aß pathologies. In addition, we investigated the expression of mGluR5 in the brains of 6xTg mice using micro-positron emission tomography (micro-PET) imaging, histological analysis, and Western blot analysis, and we observed a decrease in mGluR5 levels in the brains of 6xTg mice compared to wild-type (WT) mice. Additionally, we identified alterations in the ERK/AKT/GSK-3ß signaling pathway in the brains of 6xTg mice. Notably, we identified a significant negative correlation between depressive-like behavior and the protein level of mGluR5 in 6xTg mice. Additionally, we also found a significant positive correlation between depressive-like behavior and AD pathologies, including phosphorylated tau and Aß. These findings suggested that abnormal mGluR5 expression and AD-related pathologies were involved in depressive-like behavior in the 6xTg mouse model. Further research is warranted to elucidate the underlying mechanisms and explore potential therapeutic targets in the intersection of AD and depressive-like symptoms.

Role of kainate receptors in pruriceptive processing in the mouse spinal cord.

Pruritus, including neuropathic and psychogenic pruritus, is an unpleasant feeling that causes a desire to scratch, which negatively impacts physical and psychological aspects of daily life. Nonetheless, little is known about the neural mechanisms involved in pruritus. Glutamate is a predominant excitatory neurotransmitter in the mammalian central nervous system and exerts its effects by binding to various glutamate receptors, including kainate (KA) receptors; however, the precise involvement of each glutamate receptor in pruriceptive processing remains unclear, particularly that of KA receptors. Therefore, the roles of KA receptors in histamine-dependent and -independent itch were investigated using CNQX, an AMPA/KA receptors antagonist, UBP310 and UBP302, antagonists of KA receptors, and small interfering (si)RNAs against KA receptor subunits in mice with acute and chronic pruritus. The effects of KA receptor antagonists on histamine-induced c-Fos expression in the spinal cord were also examined. The intrathecal administration of CNQX reduced the number of scratching events induced by histamine and chloroquine. On the other hand, UBP310 or UBP302 and the siRNAs of KA receptor subunits 1-3 significantly inhibited the induction of scratching events in mice treated with histamine, while no significant change was observed in the induction of spontaneous scratching events in mice with chronic pruritus. In addition, antagonists of KA receptors attenuated c-Fos expression in the superficial layers of the dorsal horn induced by histamine. These results indicate that KA receptors are involved in acute pruriceptive processing in the spinal cord induced by histamine, but not chloroquine or chronic itch.

Kainate receptors modulate the microstructure of synchrony during dentate gyrus epileptiform activity.

Temporal Lobe Epilepsy (TLE) is the most common form of epilepsy in adults. In TLE, recurrent mossy fiber (rMF) sprouting from dentate gyrus granule cells (DGCs) forms an aberrant epileptogenic network between dentate granule cells (DGCs) that operates via ectopically expressed kainate receptors (KARs). It was previously shown that KARs expressed at the rMF-DGC synapses play a prominent role in epileptiform network events in TLE. However, it is not well understood how KARs influence neuronal network dynamics and contribute to the generation of epileptiform network activity in the dentate gyrus. To address this question, we monitored the activity of DGCs using single-cell resolution calcium imaging performed in a reliable in vitro model of TLE. Under our experimental conditions, the most prominent DGC activity patterns were interictal-like epileptiform network events, which were correlated with high levels of neuronal synchronization. The pharmacological blockade of KARs reduced the frequency as well as the number of neurons involved in these events, without altering their spatiotemporal dynamics. Analysis of the microstructure of synchrony showed that blockade of KARs diminished the fraction of neurons forming the main functional cluster. Therefore, we propose that KARs act as modulators in the epileptic network by facilitating the recruitment of neurons into coactive cell assemblies, thereby contributing to the occurrence of epileptiform network events.

Structure-Activity Relationship and Solubility Studies of N1-Substituted Quinoxaline-2,3-diones as Kainate Receptor Antagonists.

Kainate receptors are a class of ionotropic glutamate receptors that respond to the excitatory neurotransmitter glutamate in the central nervous system and play an important role in the development of neurodegenerative disorders and the regulation of synaptic function. In the current study, we investigated the structure- activity relationship of the series of quinoxaline-2,3-diones substituted at N1, 6, and 7 positions, as ligands of kainate homomeric receptors GluK1-3 and GluK5. Pharmacological characterization showed that all derivatives obtained exhibited micromolar affinity at GluK3 receptors with Ki values in the range 0.1-4.4 µM range. The antagonistic properties of the selected analogues: N-(7-fluoro-6-iodo-2,3-dioxo-3,4-dihydroquinoxalin-1(2H)-yl)-3-sulfamoylbenzamide, N-(7-(1H-imidazol-1-yl)-6-iodo-2,3-dioxo-3,4-dihydroquinoxalin-1(2H)-yl)-3-sulfamoylbenzamide and N-(7-(1H-imidazol-1-yl)-2,3-dioxo-6-(phenylethynyl)-3,4-dihydroquinoxalin-1(2H)-yl)-3-sulfamoylbenzamide at GluK3 receptors, were confirmed by an intracellular calcium imaging assay. To correlate in vitro affinity data with structural features of the synthesized compounds and to understand the impact of the substituent in N1 position on ability to form additional protein-ligand interactions, molecular modeling and docking studies were carried out. Experimental solubility studies using UV spectroscopy detection have shown that 7-imidazolyl-6-iodo analogues with a sulfamoylbenzamide moiety at the N1 position are the best soluble compounds in the series, with molar solubility in TRISS buffer at pH 9 more than 3-fold higher compared to NBQX, a known AMPA/kainate antagonist.

Interneuronal GluK1 kainate receptors control maturation of GABAergic transmission and network synchrony in the hippocampus.

Kainate type glutamate receptors (KARs) are strongly expressed in GABAergic interneurons and have the capability of modulating their functions via ionotropic and G-protein coupled mechanisms. GABAergic interneurons are critical for generation of coordinated network activity in both neonatal and adult brain, yet the role of interneuronal KARs in network synchronization remains unclear. Here, we show that GABAergic neurotransmission and spontaneous network activity is perturbed in the hippocampus of neonatal mice lacking GluK1 KARs selectively in GABAergic neurons. Endogenous activity of interneuronal GluK1 KARs maintains the frequency and duration of spontaneous neonatal network bursts and restrains their propagation through the hippocampal network. In adult male mice, the absence of GluK1 in GABAergic neurons led to stronger hippocampal gamma oscillations and enhanced theta-gamma cross frequency coupling, coinciding with faster spatial relearning in the Barnes maze. In females, loss of interneuronal GluK1 resulted in shorter sharp wave ripple oscillations and slightly impaired abilities in flexible sequencing task. In addition, ablation of interneuronal GluK1 resulted in lower general activity and novel object avoidance, while causing only minor anxiety phenotype. These data indicate a critical role for GluK1 containing KARs in GABAergic interneurons in regulation of physiological network dynamics in the hippocampus at different stages of development.

Positive and negative allosteric modulation of GluK2 kainate receptors by BPAM344 and antiepileptic perampanel.

Kainate receptors (KARs) are a subtype of ionotropic glutamate receptors that control synaptic transmission in the central nervous system and are implicated in neurological, psychiatric, and neurodevelopmental disorders. Understanding the regulation of KAR function by small molecules is essential for exploring these receptors as drug targets. Here, we present cryoelectron microscopy (cryo-EM) structures of KAR GluK2 in complex with the positive allosteric modulator BPAM344, competitive antagonist DNQX, and negative allosteric modulator, antiepileptic drug perampanel. Our structures show that two BPAM344 molecules bind per ligand-binding domain dimer interface. In the absence of an agonist or in the presence of DNQX, BPAM344 stabilizes GluK2 in the closed state. The closed state is also stabilized by perampanel, which binds to the ion channel extracellular collar sites located in two out of four GluK2 subunits. The molecular mechanisms of positive and negative allosteric modulation of KAR provide a guide for developing new therapeutic strategies.

Kainate Receptor Antagonists: Recent Advances and Therapeutic Perspective.

Since the 1990s, ionotropic glutamate receptors have served as an outstanding target for drug discovery research aimed at the discovery of new neurotherapeutic agents. With the recent approval of perampanel, the first marketed non-competitive antagonist of AMPA receptors, particular interest has been directed toward 'non-NMDA' (AMPA and kainate) receptor inhibitors. Although the role of AMPA receptors in the development of neurological or psychiatric disorders has been well recognized and characterized, progress in understanding the function of kainate receptors (KARs) has been hampered, mainly due to the lack of specific and selective pharmacological tools. The latest findings in the biology of KA receptors indicate that they are involved in neurophysiological activity and play an important role in both health and disease, including conditions such as anxiety, schizophrenia, epilepsy, neuropathic pain, and migraine. Therefore, we reviewed recent advances in the field of competitive and non-competitive kainate receptor antagonists and their potential therapeutic applications. Due to the high level of structural divergence among the compounds described here, we decided to divide them into seven groups according to their overall structure, presenting a total of 72 active compounds.

Asymmetry and Ion Selectivity Properties of Bacterial Channel NaK Mutants Derived from Ionotropic Glutamate Receptors.

Ionotropic glutamate receptors are ligand-gated cation channels that play essential roles in the excitatory synaptic transmission throughout the central nervous system. A number of open-pore structures of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic-acid (AMPA)-type glutamate receptors became available recently by cryo-electron microscopy (cryo-EM). These structures provide valuable insights into the conformation of the selectivity filter (SF), the part of the ion channel that determines the ion selectivity. Nonetheless, due to the moderate resolution of the cryo-EM structures, detailed information such as ion occupancy of monovalent and divalent cations as well as positioning of the side-chains in the SF is still missing. Here, in an attempt to obtain high-resolution information about glutamate receptor SFs, we incorporated partial SF sequences of the AMPA and kainate receptors into the bacterial tetrameric cation channel NaK, which served as a structural scaffold. We determined a series of X-ray structures of NaK-CDI, NaK-SDI and NaK-SELM mutants at 1.42-2.10 Å resolution, showing distinct ion occupation of different monovalent cations. Molecular dynamics (MD) simulations of NaK-CDI indicated the channel to be conductive to monovalent cations, which agrees well with our electrophysiology recordings. Moreover, previously unobserved structural asymmetry of the SF was revealed by the X-ray structures and MD simulations, implying its importance in ion non-selectivity of tetrameric cation channels.