GluA1 Subunit Research Articles

Modulation of top-down influence affects trafficking of glutamatergic receptors in the primary visual cortex

Numerous studies indicate that top-down influence plays critical roles in visual perception by enhancing neuronal excitability in the primary visual cortex (V1). The underlying mechanisms are poorly understood. This study examined changes of excitatory glutamatergic markers in the V1 cortex of cat after top-down influence of cortical area 7 (A7) was modulated by transcranial direct current stimulation (tDCS). Our results showed that the content of glutamate as well as the total cellular protein of glutamatergic receptors, including the key subunit GluA1 of AMPA receptors and subunit NR1 of NMDA receptors, in the V1 cortex had no significant change after anode- and cathode-tDCS relative to sham-tDCS in A7. However, the plasma membrane protein content of GluA1 and NR1 in the V1 was significantly increased after anode-tDCS, but decreased after cathode-tDCS when compared with that after sham-tDCS in A7. Further, the abundance of phosphorylated GluA1 and NR1 in the V1 also elevated significantly after anode-tDCS, but lowered after cathode-tDCS compared with that after sham-tDCS. Additionally, the content of phosphorylated CaMKII (p-CaMKII), a protein kinase preferentially boosting phosphorylation of glutamatergic receptors, in the V1 improved after anode-tDCS although no significant alteration occurred after c-tDCS in A7. Taken together, our results indicate that feedback influence of A7 may facilitate the trafficking of glutamatergic receptors to postsynaptic membrane in the V1 cortex through receptors’ phosphorylation process, which could be an important mechanism of high-level cortex in modulating visual information processing in the V1 cortex.

Disrupted Maturation of Prefrontal Layer 5 Neuronal Circuits in an Alzheimer's Mouse Model of Amyloid Deposition.

Mutations in genes encoding amyloid precursor protein (APP) and presenilins (PSs) cause familial forms of Alzheimer's disease (AD), a neurodegenerative disorder strongly associated with aging. It is currently unknown whether and how AD risks affect early brain development, and to what extent subtle synaptic pathology may occur prior to overt hallmark AD pathology. Transgenic mutant APP/PS1 over-expression mouse lines are key tools for studying the molecular mechanisms of AD pathogenesis. Among these lines, the 5XFAD mice rapidly develop key features of AD pathology and have proven utility in studying amyloid plaque formation and amyloid β (Aβ)-induced neurodegeneration. We reasoned that transgenic mutant APP/PS1 over-expression in 5XFAD mice may lead to neurodevelopmental defects in early cortical neurons, and performed detailed synaptic physiological characterization of layer 5 (L5) neurons from the prefrontal cortex (PFC) of 5XFAD and wild-type littermate controls. L5 PFC neurons from 5XFAD mice show early APP/Aβ immunolabeling. Whole-cell patch-clamp recording at an early post-weaning age (P22-30) revealed functional impairments; although 5XFAD PFC-L5 neurons exhibited similar membrane properties, they were intrinsically less excitable. In addition, these neurons received smaller amplitude and frequency of miniature excitatory synaptic inputs. These functional disturbances were further corroborated by decreased dendritic spine density and spine head volumes that indicated impaired synapse maturation. Slice biotinylation followed by Western blot analysis of PFC-L5 tissue revealed that 5XFAD mice showed reduced synaptic AMPA receptor subunit GluA1 and decreased synaptic NMDA receptor subunit GluN2A. Consistent with this, patch-clamp recording of the evoked L23>L5 synaptic responses revealed a reduced AMPA/NMDA receptor current ratio, and an increased level of AMPAR-lacking silent synapses. These results suggest that transgenic mutant forms of APP/PS1 overexpression in 5XFAD mice leads to early developmental defects of cortical circuits, which could contribute to the age-dependent synaptic pathology and neurodegeneration later in life.

Sex- and age-dependent effects of chronic corticosterone exposure on depressive-like, anxiety-like, and fear-related behavior: Role of amygdala glutamate receptors in the rat.

Persistent glucocorticoid elevation consistent with chronic stress exposure can lead to psychopathology, including mood and anxiety disorders. Women and stress-exposed adolescents are more likely to be diagnosed with mood disorders, suggesting that sex and age are important factors in determining vulnerability, though much remains to be determined regarding the mechanisms underlying this risk. Thus, the aim of the present experiments was to use the chronic corticosterone (CORT) exposure paradigm, a model of depression-like behavior that has previously been established primarily in adult males, to determine the mood-related effects of CORT in female and adolescent rats. Depression- and anxiety-like effects in adulthood were determined using the sucrose preference (SPT), the forced swim test (FST), the elevated plus maze, and fear conditioning. Basolateral amygdala (BLA) and medial prefrontal cortex (mPFC) glutamate receptor subunit levels were then measured. In a subsequent experiment, adult male and female rats were tested for the effects of pharmacological activation (via AMPA) or inhibition (via NBQX) of AMPA receptors in the BLA on behavior in the FST. Overall, females showed reduced anxiety- and depressive-like behaviors relative to males. However, females treated with CORT in adolescence, but not adulthood, had increased immobility in the FST, indicative of depression-like behavior. In contrast, CORT did not alter behavior in adolescent-treated males, though the previously reported depression-like effect of adult CORT exposure was observed. Control females had higher expression of the AMPA receptor subunits GluA1 and GluA2/3 selectively in the BLA relative to males. Adolescent CORT treatment, however, decreased BLA GluA1 and GluA2/3 expression in females, but increased expression in males, consistent with the direction of depression-like behavioral effects. Male and female rats also demonstrated opposing patterns of response to BLA AMPA receptor modulation in the FST, with AMPA infusion magnifying the sex difference of decreased immobility in females. Overall, these experiments show that increased glutamate receptor function in the BLA may decrease the risk of developing depressive-like behavior, further supporting efforts to target glutamatergic receptors for the treatment of stress-related psychiatric disorders. These findings also support further focus on sex as a biological variable in neuropsychiatric research.

Increased Fear Generalization and Amygdala AMPA Receptor Proteins in Chronic Traumatic Brain Injury.

Cognitive impairments and emotional lability are common long-term consequences of traumatic brain injury (TBI). How TBI affects interactions between sensory, cognitive, and emotional systems may reveal mechanisms that underlie chronic mental health comorbidities. Previously, we reported changes in auditory-emotional network activity and enhanced fear learning early after TBI. In the current study, we asked whether TBI has long-term effects on fear learning and responses to novel stimuli. Four weeks following lateral fluid percussion injury (FPI) or sham surgery, adult male rats were fear conditioned to either white noise-shock or tone-shock pairing, or shock-only control and subsequently were tested for freezing to context and to the trained or novel auditory cues in a new context. FPI groups showed greater freezing to their trained auditory cue, indicating long-term TBI enhanced fear. Interestingly, FPI-Noise Shock animals displayed robust fear to the novel, untrained tone compared with Sham-Noise Shock across both experiments. Shock Only groups did not differ in freezing to either auditory stimulus. These findings suggest that TBI precipitates maladaptive associative fear generalization rather than non-associative sensitization. Basolateral amygdala (BLA) α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAr) subunits GluA1 and GluA2 levels were analyzed and the FPI-Noise Shock group had increased GluA1 (but not GluA2) levels that correlated with the level of tone fear generalization. This study illustrates a unique chronic TBI phenotype with both a cognitive impairment and increased fear and possibly altered synaptic transmission in the amygdala long after TBI, where stimulus generalization may underlie maladaptive fear and hyperarousal.

Familial ALS-associated SFPQ variants promote the formation of SFPQ cytoplasmic aggregates in primary neurons.

Splicing factor proline- and glutamine-rich (SFPQ) is a nuclear RNA-binding protein that is involved in a wide range of physiological processes including neuronal development and homeostasis. However, the mislocalization and cytoplasmic aggregation of SFPQ are associated with the pathophysiology of amyotrophic lateral sclerosis (ALS). We have previously reported that zinc mediates SFPQ polymerization and promotes the formation of cytoplasmic aggregates in neurons. Here we characterize two familial ALS (fALS)-associated SFPQ variants, which cause amino acid substitutions in the proximity of the SFPQ zinc-coordinating centre (N533H and L534I). Both mutants display increased zinc-binding affinities, which can be explained by the presence of a second zinc-binding site revealed by the 1.83 Å crystal structure of the human SFPQ L534I mutant. Overexpression of these fALS-associated mutants significantly increases the number of SFPQ cytoplasmic aggregates in primary neurons. Although they do not affect the density of dendritic spines, the presence of SFPQ cytoplasmic aggregates causes a marked reduction in the levels of the GluA1, but not the GluA2 subunit of AMPA-type glutamate receptors on the neuronal surface. Taken together, our data demonstrate that fALS-associated mutations enhance the propensity of SFPQ to bind zinc and form aggregates, leading to the dysregulation of AMPA receptor subunit composition, which may contribute to neuronal dysfunction in ALS.

Three Binding Conformations of BIO124 in the Pocket of the PICK1 PDZ Domain.

The PDZ family has drawn attention as possible drug targets because of the domains’ wide ranges of function and highly conserved binding pockets. The PICK1 PDZ domain has been proposed as a possible drug target because the interactions between the PICK1 PDZ domain and the GluA2 subunit of the AMPA receptor have been shown to progress neurodegenerative diseases. BIO124 has been identified as a sub µM inhibitor of the PICK1–GluA2 interaction. Here, we use all-atom molecular dynamics simulations to reveal the atomic-level interaction pattern between the PICK1 PDZ domain and BIO124. Our simulations reveal three unique binding conformations of BIO124 in the PICK1 PDZ binding pocket, referred to here as state 0, state 1, and state 2. Each conformation is defined by a unique hydrogen bonding network and a unique pattern of hydrophobic interactions between BIO124 and the PICK1 PDZ domain. Interestingly, each conformation of BIO124 results in different dynamic changes to the PICK1 PDZ domain. Unlike states 1 and 2, state 0 induces dynamic coupling between BIO124 and the αA helix. Notably, this dynamic coupling with the αA helix is similar to what has been observed in other PDZ–ligand complexes. Our analysis indicates that the interactions formed between BIO124 and I35 may be the key to inducing dynamic coupling with the αA helix. Lastly, we suspect that the conformational shifts observed in our simulations may affect the stability and thus the overall effectiveness of BIO124. We propose that a physically larger inhibitor may be necessary to ensure sufficient interactions that permit stable binding between a drug and the PICK1 PDZ domain.

MC4R Deficiency Causes Dysregulation of Postsynaptic Excitatory Synaptic Transmission as a Crucial Culprit for Obesity.

Melanocortin 4 receptor (MC4R) in the paraventricular nucleus of the hypothalamus (PVH) shows bidirectional characterization in modulating food intake and energy homeostasis. We demonstrate that MC4R knockdown (KD) in the PVH can attenuate AMPA receptor (AMPAR)-mediated postsynaptic responses by altering the phosphorylation of AMPAR GluA1 subunit through the protein kinase A (PKA)-dependent signaling cascade and simultaneously lead to rapid body weight gain. Furthermore, PKA KD in the PVH engendered similar electrophysiological and behavioral phenotypes as in MC4R KD mice. Importantly, we observed that the reduction of AMPAR GluA1 expression not only led to attenuated synaptic responses but also caused body weight gain, suggesting that the aberration of synaptic responses may be one of the crucial pathogeny of obesity. Our study provides the synaptic and molecular explanations of how body weight is regulated by MC4R in the PVH.

Upregulation of Netrin-1 in the hippocampus mediates the formation of visceral hypersensitivity induced by maternal separation

Early adverse life events (EALs), such as maternal separation (MS), can cause visceral hypersensitivity, which is thought to be a key pathophysiological mechanism of irritable bowel syndrome (IBS). Previous studies mainly focused on EALs-induced visceral hypersensitivity in adulthood but did not consider that it may have occurred in the preadult period. We previously found that rats who experienced MS suffered from visceral hypersensitivity starting from the post-weaning period. Moreover, the hippocampus is considered to be critical in regulating the formation of visceral hypersensitivity induced by MS. But the underlying mechanisms throughout different life periods are unclear. In this study, behavioral tests, RNA-seq, lentiviral interference, and molecular biology techniques were applied to investigate the molecular mechanism in the hippocampus underlying MS-induced long-lasting visceral hypersensitivity. It was found that both visceral sensitivity and anxiety-like behaviors were significantly increased in MS rats in post-weaning, prepubertal, and adult periods, especially in the prepubertal period. Subsequently, RNA-seq targeting the hippocampus identified that the expression level of Netrin-1 was significantly increased in all periods, which was further confirmed by quantitative real-time PCR and Western blot. Knocking-down hippocampal Netrin-1 in the post-weaning period by lentivirus interference alleviated visceral hypersensitivity and anxiety-like behaviors of MS rats in the later phase of life. In addition, deleted in colorectal cancer (DCC), instead of neogenin-1(Neo-1) or uncoordinated (UNC5), was proved to be the specific functional receptor of Netrin-1 in regulating visceral hypersensitivity, whose upregulation may result in the most severe symptoms in the prepubertal period. Furthermore, the activation of the Netrin-1/DCC pathway could enhance long-term potentiation (LTP) in the hippocampus, probably via recruitment of the AMPA receptor subunit GluA1, which finally resulted in the formation of visceral hypersensitivity. These novel findings suggest that long-lasting over-expression of Netrin-1 can mediate visceral hypersensitivity and anxiety disorder from the post-weaning period to adulthood by activating DCC/GluA1 pathway in the hippocampus. Moreover, early intervention of Netrin-1 in the post-weaning period could lead to significant symptom relief afterward, which provides evidence that the Netrin-1/DCC/GluA1 signaling pathway may be a potential therapeutic target for the treatment of visceral hypersensitivity in clinics.

All-trans Retinoic Acid has Limited Therapeutic Effects on Cognition and Hippocampal Protein Expression After Controlled Cortical Impact

Traumatic brain injury (TBI) is known to impair synaptic function, and subsequently contribute to observed cognitive deficits. Retinoic Acid (RA) signaling modulates expression of synaptic plasticity proteins and is involved in hippocampal learning and memory. All trans-retinoic acid (ATRA), a metabolite of Vitamin A, has been identified as a potential pharmacotherapeutic for other neurological disorders due to this role. This study conducted an ATRA dose response to determine its therapeutic effects on cognitive behaviors and expression of hippocampal markers of synaptic plasticity and RA signaling proteins after experimental TBI. Under isoflurane anesthesia, adult male Sprague Dawley rats received either controlled cortical impact (CCI, 2.5 mm deformation, 4 m/s) or control surgery. Animals received daily intraperitoneal injection of 0.5, 1, 5, or 10 mg/kg of ATRA or vehicle for 2 weeks. Animals underwent motor and spatial learning and memory testing. Hippocampal expression of synaptic plasticity proteins neurogranin (Ng), and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor GluA1 sub-unit, as well as RA signaling proteins STRA6, ADLH1a1, CYP26A1 and CYP26B1 were evaluated by western blot at 2-weeks post-injury. ATRA treatment significantly recovered Ng synaptic protein expression, while having no effect on motor performance, spatial learning, and memory, and GluA1 expression after TBI. RA signaling protein expression is unchanged 2 weeks after TBI.Overall, ATRA administration after TBI showed limited therapeutic benefits compared to the vehicle.

ADAR2 enzymes: efficient site-specific RNA editors with gene therapy aspirations.

The adenosine deaminase acting on RNA (ADAR) enzymes are essential for neuronal function and innate immune control. ADAR1 RNA editing prevents aberrant activation of antiviral dsRNA sensors through editing of long, double-stranded RNAs (dsRNAs). In this review, we focus on the ADAR2 proteins involved in the efficient, highly site-specific RNA editing to recode open reading frames first discovered in the GRIA2 transcript encoding the key GLUA2 subunit of AMPA receptors; ADAR1 proteins also edit many of these sites. We summarize the history of ADAR2 protein research and give an up-to-date review of ADAR2 structural studies, human ADARBI (ADAR2) mutants causing severe infant seizures, and mouse disease models. Structural studies on ADARs and their RNA substrates facilitate current efforts to develop ADAR RNA editing gene therapy to edit disease-causing single nucleotide polymorphisms (SNPs). Artificial ADAR guide RNAs are being developed to retarget ADAR RNA editing to new target transcripts in order to correct SNP mutations in them at the RNA level. Site-specific RNA editing has been expanded to recode hundreds of sites in CNS transcripts in Drosophila and cephalopods. In Drosophila and C. elegans, ADAR RNA editing also suppresses responses to self dsRNA.

Sciatic nerve block downregulates the BDNF pathway to alleviate the neonatal incision-induced exaggeration of incisional pain via decreasing microglial activation

Sciatic nerve block is under investigation as a possible therapeutic strategy for neonatal injury-induced exaggeration of pain responses to reinjury. Spinal microglial priming, brain-derived neurotrophic factor (BDNF) and Src homology-2 domain-containing protein tyrosine phosphatase-2 (SHP2) participate in exaggerated incisional pain induced by neonatal incision. However, effects of sciatic nerve block on exacerbated incisional pain and underlying mechanisms remain unclear. Here, we demonstrated that sciatic nerve block alleviates pain hypersensitivity and microglial activation in rats subjected to neonatal incision and adult incision (nIN-IN). Chemogenetic activation or inhibition of spinal microglia attenuates or mimics effects of sciatic nerve block on pain hypersensitivity, respectively. Moreover, α-amino-3-hydroxy- 5-methy- 4-isoxazole propionate (AMPA) receptor subunit GluA1 contributes to the exaggeration of incisional pain. The inhibition of BDNF or SHP2 blocks upregulations of downstream molecules in nIN-IN rats. Knockdown of SHP2 attenuates the increase of GluA1 induced by injection of BDNF in adult rats with only neonatal incision. The inhibition of microglia or ablation of microglial BDNF attenuates upregulations of SHP2 and GluA1. Additionally, sciatic nerve block downregulates the expression of these three molecules. Upregulation of BDNF, SHP2 or AMPA receptor attenuates sciatic nerve block-induced reductions of downstream molecules and pain hypersensitivity. Microglial activation abrogates reductions of these three molecules induced by sciatic nerve block. These results suggest that decreased activation of spinal microglia contributes to beneficial effects of sciatic nerve block on the neonatal incision-induced exaggeration of incisional pain via downregulating BDNF/SHP2/GluA1-containing AMPA receptor signaling. Thus, sciatic nerve block may be a promising therapy.

Opposing reduced VPAC1 and enhanced VPAC2 VIP receptors in the hippocampus of the Li2+-pilocarpine rat model of temporal lobe epilepsy

VIP binding sites are upregulated in mesial temporal lobe epilepsy (MTLE) patients, also suffering from severe cognitive deficits. Although altered VIP and VIP receptor levels were described in rodent models of epilepsy, the VIP receptor subtype(s) were never identified. We now investigated how VPAC1 and VPAC2 receptor levels change in the Li2+-pilocarpine rat model of MTLE. Cognitive decline and altered synaptic plasticity as estimated from phosphorylation of AMPA GluA1 subunit on Ser831 and Ser845 and AMPA GluA1/GluA2 ratio was also probed. Animals showing spontaneous recurrent seizures (SRSs) for at least 4 weeks showed impaired learning in the radial arm maze (RAM) and presented decreased VPAC1 and increased VPAC2 receptor levels. In addition, SRSs rats showed increased AMPA GluA1 phosphorylation in Ser831 and Ser845, marked decrease in GluA1 levels and a milder decrease in GluA2 levels. Consequently, the GluA1/GluA2 ratio was also decreased in SRSs rats.Altered VIP receptor levels may differentially prevent or contribute to MTLE pathology, since VPAC1 receptors promote the endogenous control of LTP, mediate endogenous VIP neuroprotection against altered synaptic plasticity following epileptiform activity, and mediate anti-inflammatory actions in microglia, while VPAC2 receptors mediate VIP endogenous neuroprotection against neonatal excitotoxicity and prevent reactive astrogliosis. This discovery imposes a different mindset for considering VIP receptors as therapeutic targets in MTLE, allowing a differential targeting of the cellular events contributing to epileptogenesis.

Extrahippocampal seizure and memory circuits overlap.

Seizures cause retrograde amnesia. We have previously demonstrated that seizures erode recently formed memories through shared ensembles and mechanisms in the CA1 region of the hippocampus. Here, we tested whether seizure circuits overlap spatial memory circuits outside of the CA. Spatial memory is consolidated by the hippocampal-cortical coupling that are connected via multiple pathways. We tested whether a seizure invades structures involved in memory consolidation by using the activity reporter TRAP2 mice. T-maze alternation learning activated neurons in the dentate gyrus (DG), mediodorsal thalamus (MD), retrosplenial cortex (RSC), and medial prefrontal cortex (mPFC). This spatial memory relies on the plasticity of the AMPA receptor GluA1 subunit. GluA1 knock-out (KO)/TRAP2 mice did not learn to alternate, and structures interposed between the hippocampus and the cortex were not active. A seizure prevented the recall of alternation memory and activated memory-labeled structures. There was a widespread overlap between learning-activated ensembles and seizure-activated neurons, which likely contributes to retrograde amnesia.

A Class I HDAC Inhibitor Rescues Synaptic Damage and Neuron Loss in APP-Transfected Cells and APP/PS1 Mice through the GRIP1/AMPA Pathway.

As a neurodegenerative disease, Alzheimer’s disease (AD) seriously affects the health of older people. Changes in synapses occur first over the course of the disease, perhaps even before the formation of Aβ plaques. Histone deacetylase (HDAC) mediates the damage of Aβ oligomers to dendritic spines. Therefore, we examined the relationship between HDAC activity and synaptic defects using an HDAC inhibitor (HDACI), BG45, in the human neuroblastoma SH-SY5Y cell line with stable overexpression of Swedish mutant APP (APPsw) and in APP/PS1 transgenic mice during this study. The cells were treated with 15 μM BG45 and the APP/PS1 mice were treated with 30 mg/kg BG45. We detected the levels of synapse-related proteins, HDACs, tau phosphorylation, and amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors using Western blotting and immunohistochemistry. We also measured the expression of cytoskeletal proteins in the cell model. The mRNA levels of the glutamate ion receptor alginate subunit 2 (GRIK2), sodium voltage-gated channel beta subunit (SCN3B), synaptophysin (SYP), Grm2 (the gene encoding glutamate receptor subunit 2 (GluR2)), Grid2IP, glutamate receptor interacting protein 1 (GRIP1), and GRIP2 were detected to explore the effects of the HDACI on regulating the expression of synaptic proteins and AMPA receptors. According to our studies, the expressions of HDAC1, HDAC2, and HDAC3 were increased, which were accompanied by the downregulation of the synapse-related proteins SYP, postsynaptic dendritic protein (PSD-95), and spinophilin as early as 24 h after transfection with the APPsw gene. BG45 upregulated the expression of synapse-related proteins and repaired cytoskeletal damage. In vivo, BG45 alleviated the apoptosis-mediated loss of hippocampal neurons, upregulated synapse-related proteins, reduced Aβ deposition and phosphorylation of tau, and increased the levels of the synapse-related genes GRIK2, SCN3B, SYP, Grm2, and Grid2IP. BG45 increased the expression of the AMPA receptor subunits GluA1, GluA2, and GluA3 on APPsw-transfected cells and increased GRIP1 and GRIP2 expression and AMPA receptor phosphorylation in vivo. Based on these results, HDACs are involved in the early process of synaptic defects in AD models, and BG45 may rescue synaptic damage and the loss of hippocampal neurons by specifically inhibiting HDAC1, HDAC2, and HDAC3, thereby modulating AMPA receptor transduction, increasing synapse-related gene expression, and finally enhancing the function of excitatory synapses. BG45 may be considered a potential drug for the treatment of early AD in further studies.

Autoimmunity to glutamate receptor channels

AbstractGlutamate and its receptors are involved in pleiotropic roles including experience‐dependent synaptic plasticity and brain development as the main fast excitatory neurotransmitters in the central nervous system. There are two main families of the glutamate receptors: (1) ionotropic glutamate receptors (iGluRs) that are ligand‐gated ion channel receptors accessible for fast synaptic transmission, and (2) metabotropic glutamate receptors (mGluRs) that are G‐protein‐coupled receptors modulating slow synaptic transmission through intracellular second messengers. Here, we summarize the current concepts of autoimmunity to these glutamate receptors in neurological disorders: anti‐GluN1 subunit of N‐methyl‐D‐aspartate receptor (NMDARs) for anti‐NMDAR encephalitis, anti‐GluA1 and GluA2 subunits of α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid receptor (AMPARs) for anti‐AMPAR encephalitis, and anti‐mGluR5 subunit for Ophelia syndrome. Most identified autoimmunity to glutamate receptors can lead to unique synaptopathies or neuronal antibody‐mediated encephalopathies. Evidence‐based optimal management guidelines and new disease‐modifying therapies need to be developed for patients with autoimmune encephalitis including anti‐glutamate receptor encephalitis.

Interfering With Contextual Fear Memories by Post-reactivation Administration of Propranolol in Mice: A Series of Null Findings.

Post-reactivation amnesia of contextual fear memories by blockade of noradrenergic signaling has been shown to have limited replicability in rodents. This is usually attributed to several boundary conditions that gate the destabilization of memory during its retrieval. How these boundary conditions can be overcome, and what neural mechanisms underlie post-reactivation changes in contextual fear memories remain largely unknown. Here, we report a series of experiments in a contextual fear-conditioning paradigm in mice, that were aimed at solving these issues. We first attempted to obtain a training paradigm that would consistently result in contextual fear memory that could be destabilized upon reactivation, enabling post-retrieval amnesia by the administration of propranolol. Unexpectedly, our attempts were unsuccessful to this end. Specifically, over a series of experiments in which we varied different parameters of the fear acquisition procedure, at best small and inconsistent effects were observed. Additionally, we found that propranolol did not alter retrieval-induced neural activity, as measured by the number of c-Fos+ cells in the hippocampal dentate gyrus. To determine whether propranolol was perhaps ineffective in interfering with reactivated contextual fear memories, we also included anisomycin (i.e., a potent and well-known amnesic drug) in several experiments, and measures of synaptic glutamate receptor subunit GluA2 (i.e., a marker of memory destabilization). No post-retrieval amnesia by anisomycin and no altered GluA2 expression by reactivation was observed, suggesting that the memories did not undergo destabilization. The null findings are surprising, given that the training paradigms we implemented were previously shown to result in memories that could be modified upon reactivation. Together, our observations illustrate the elusive nature of reactivation-dependent changes in non-human fear memory.

Retromer dysfunction in amyotrophic lateral sclerosis

Retromer is a heteropentameric complex that plays a specialized role in endosomal protein sorting and trafficking. Here, we report a reduction in the retromer proteins-vacuolar protein sorting 35 (VPS35), VPS26A, and VPS29-in patients with amyotrophic lateral sclerosis (ALS) and in the ALS model provided by transgenic (Tg) mice expressing the mutant superoxide dismutase-1 G93A. These changes are accompanied by a reduction of levels of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunit GluA1, a proxy of retromer function, in spinal cords from Tg SOD1G93A mice. Correction of the retromer deficit by a viral vector expressing VPS35 exacerbates the paralytic phenotype in Tg SOD1G93A mice. Conversely, lowering Vps35 levels in Tg SOD1G93A mice ameliorates the disease phenotype. In light of these findings, we propose that mild alterations in retromer inversely modulate neurodegeneration propensity in ALS.

Phosphorylation of GluA1-Ser831 by CaMKII Activation in the Caudate and Putamen Is Required for Behavioral Sensitization After Challenge Nicotine in Rats.

BackgroundPhosphorylation of the glutamate receptor (GluA1) subunit of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor plays a crucial role in behavioral sensitization after exposure to psychostimulants. The present study determined the potential role of serine 831 (Ser831) phosphorylation in the GluA1 subunit of the caudate and putamen (CPu) in behavioral sensitization after challenge nicotine.MethodsChallenge nicotine (0.4 mg/kg) was administered subcutaneously (s.c.) after 7 days of repeated exposure to nicotine (0.4 mg/kg, s.c.) followed by 3 days of withdrawal in rats. Bilateral intra-CPu infusions of drugs were mainly performed to test this hypothesis.ResultsChallenge nicotine increased both phosphorylated (p)Ser831 immunoreactivity (IR) and pCa2+/calmodulin-dependentprotein kinases II (pCaMKII)-IR in the medium spiny neurons (MSNs) of the CPu. These increases were prevented by bilateral intra-CPu infusion of the metabotropic glutamate receptor 5 (mGluR5) antagonist MPEP (0.5 nmol/side) and the N-methyl-D-aspartate (NMDA) receptor antagonist MK801 (2 nmol/side). However, the dopamine D1 receptor (D1R) antagonist SCH23390 (7.5 nmol/side) prevented only pSer831-IR alone. Bilateral intra-CPu infusion of the Tat-GluA1D peptide (25 pmol/side), which interferes with the binding of pCaMKII to GluA1-Ser831, decreased the challenge nicotine–induced increase in locomotor activity.ConclusionsThese findings suggest that the GluA1-Ser831 phosphorylation in the MSNs of the CPu is required for the challenge nicotine–induced behavioral sensitization in rats. CaMKII activation linked to mGluR5 and NMDA receptors, but not to D1R, is essential for inducing the CaMKII-Ser831 interaction.

Identification and functional evaluation of GRIA1 missense and truncation variants in individuals with ID: An emerging neurodevelopmental syndrome

SummaryGRIA1 encodes the GluA1 subunit of α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptors, which are ligand-gated ion channels that act as excitatory receptors for the neurotransmitter L-glutamate (Glu). AMPA receptors (AMPARs) are homo- or heteromeric protein complexes with four subunits, each encoded by different genes, GRIA1 to GRIA4. Although GluA1-containing AMPARs have a crucial role in brain function, the human phenotype associated with deleterious GRIA1 sequence variants has not been established. Subjects with de novo missense and nonsense GRIA1 variants were identified through international collaboration. Detailed phenotypic and genetic assessments of the subjects were carried out and the pathogenicity of the variants was evaluated in vitro to characterize changes in AMPAR function and expression. In addition, two Xenopus gria1 CRISPR-Cas9 F0 models were established to characterize the in vivo consequences. Seven unrelated individuals with rare GRIA1 variants were identified. One individual carried a homozygous nonsense variant (p.Arg377Ter), and six had heterozygous missense variations (p.Arg345Gln, p.Ala636Thr, p.Ile627Thr, and p.Gly745Asp), of which the p.Ala636Thr variant was recurrent in three individuals. The cohort revealed subjects to have a recurrent neurodevelopmental disorder mostly affecting cognition and speech. Functional evaluation of major GluA1-containing AMPAR subtypes carrying the GRIA1 variant mutations showed that three of the four missense variants profoundly perturb receptor function. The homozygous stop-gain variant completely destroys the expression of GluA1-containing AMPARs. The Xenopus gria1 models show transient motor deficits, an intermittent seizure phenotype, and a significant impairment to working memory in mutants. These data support a developmental disorder caused by both heterozygous and homozygous variants in GRIA1 affecting AMPAR function.

Vasopressin acts as a synapse organizer in limbic regions by boosting PSD95 and GluA1 expression.

Hypothalamic arginine vasopressin (AVP)-containing magnocellular neurosecretory neurons (AVPMNN) emit collaterals to synaptically innervate limbic regions influencing learning, motivational behaviour, and fear responses. Here, we characterize the dynamics of expression changes of two key determinants for synaptic strength, the postsynaptic density (PSD) proteins AMPAR subunit GluA1 and PSD scaffolding protein 95 (PSD95), in response to in vivo manipulations of AVPMNN neuronal activation state, or exposure to exogenous AVP ex vivo. Both long-term water deprivation in vivo, which powerfully upregulates AVPMNN metabolic activity, and exogenous AVP application ex vivo, in brain slices, significantly increased GluA1 and PSD95 expression as measured by western blotting, in brain regions reportedly receiving direct ascending innervations from AVPMNN (i.e., ventral hippocampus, amygdala and lateral habenula). By contrast, the visual cortex, a region not observed to receive AVPMNN projections, showed no such changes. Ex vivo application of V1a and V1b antagonists to ventral hippocampal slices ablated the AVP stimulated increase in postsynaptic protein expression measured by western blotting. Using a modified expansion microscopy technique, we were able to quantitatively assess the significant augmentation of PSD95 and GLUA1 densities in subcellular compartments in locus coeruleus tyrosine hydroxylase immunopositive fibres, adjacent to AVP axon terminals. Our data strongly suggest that the AVPMNN ascending system plays a role in the regulation of the excitability of targeted neuronal circuits through upregulation of key postsynaptic density proteins corresponding to excitatory synapses.