Stimulating ß-adrenergic receptors promotes synaptic potentiation by switching CaMKII movement from LTD to LTP mode.

Learning, memory, and cognition are thought to require synaptic plasticity, specifically including hippocampal long-term potentiation and depression (LTP and LTD). LTP versus LTD is induced by high-frequency stimulation versus low-frequency, but stimulating ß-adrenergic receptors (ßARs) enables LTP induction also by low-frequency stimulation (1 Hz) or theta frequencies (∼5 Hz) that do not cause plasticity by themselves. In contrast to high-frequency stimulation-LTP, such ßAR-LTP requires Ca2+-flux through L-type voltage-gated Ca2+-channels, not N-methyl-D-aspartate-type glutamate receptors. Surprisingly, we found that ßAR-LTP still required a nonionotropic scaffolding function of the N-methyl-D-aspartate-type glutamate receptor: the stimulus-induced binding of the Ca2+/calmodulin-dependent protein kinase II (CaMKII) to its GluN2B subunit that mediates CaMKII movement to excitatory synapses. In hippocampal neurons, ß-adrenergic stimulation with isoproterenol (Iso) transformed LTD-type CaMKII movement to LTP-type movement, resulting in CaMKII movement to excitatory instead of inhibitory synapses. Additionally, Iso enabled induction of a major cell-biological feature of LTP in response to LTD stimuli: increased surface expression of GluA1 fused with super-ecliptic pHluorein. Like for ßAR-LTP in hippocampal slices, the Iso effects on CaMKII movement and surface expression of GluA1 fused with super-ecliptic pHluorein involved L-type Ca2+-channels and specifically required ß2-ARs. Taken together, these results indicate that Iso transforms LTD stimuli to LTP signals by switching CaMKII movement and GluN2B binding to LTP mode.

Long-Term Outcome of Pediatric Patients with Anti-NMDA Receptor Encephalitis in a Single Center.

BACKGROUND: Anti-N-methyl-D-aspartate (NMDA) receptor encephalitis is the most common autoimmune encephalitis in children. There is a high probability of recovery if treated promptly. We aimed to analyze the clinical features and long-term outcomes of pediatric patients with anti-NMDA receptor encephalitis. METHOD: We conducted a retrospective study with definite diagnoses of anti-NMDA receptor encephalitis in 11 children treated in a tertiary referral center between March 2012 and March 2022. Clinical features, ancillary tests, treatment, and outcomes were reviewed. RESULTS: The median age at disease onset was 7.9 years. There were eight females (72.7%) and three males (27.3%). Three (27.3%) patients initially presented with focal and/or generalized seizures and eight (72.7%) with behavioral change. Seven patients (63.6%) revealed normal brain MRI scans. Seven (63.6%) had abnormal EEG results. Ten patients (90.1%) received intravenous immunoglobulin, corticosteroid, and/or plasmapheresis. After a median follow-up duration of 3.5 years, one patient was lost to follow-up at the acute stage, nine (90%) had an mRS ≤ 2, and only one had an mRS of 3. CONCLUSIONS: With the early recognition of anti-NMDA receptor encephalitis based on its clinical features and ancillary tests, we were able to treat patients promptly with first-line treatment and achieve favorable neurological outcomes.

Systematic analysis to identify novel disease indications and plausible potential chemical leads of glutamate ionotropic receptor NMDA type subunit 1, GRIN1.

Schizophrenia is a mental illness affecting the normal lifestyle of adults and early adolescents incurring major symptoms as jumbled speech, involvement in everyday activities eventually got reduced, patients always struggle with attention and memory, reason being both the genetic and environmental factors responsible for altered brain chemistry and structure, resulting in schizophrenia and associated orphan diseases. The network biology describes the interactions among genes/proteins encoding molecular mechanisms of biological processes, development, and diseases. Besides, all the molecular networks, protein-protein Interaction Networks have been significant in distinguishing the pathogenesis of diseases and thereby drug discovery. The present meta-analysis prioritizes novel disease indications viz. rare and orphan diseases associated with target Glutamate Ionotropic Receptor NMDA Type Subunit 1, GRIN1 using text mining knowledge-based tools. Furthermore, ZINC database was virtually screened, and binding conformation of selected compounds was performed and resulted in the identification of Narciclasine (ZINC04097652) and Alvespimycin (ZINC73138787) as potential inhibitors. Furthermore, docked complexes were subjected to MD simulation studies which suggests that the identified leads could be a better potential drug to recuperate schizophrenia.

CaMKII T286 phosphorylation has distinct essential functions in three forms of long-term plasticity.

The Ca2+/calmodulin-dependent protein kinase II (CaMKII) mediates long-term potentiation or depression (LTP or LTD) after distinct stimuli of hippocampal NMDA-type glutamate receptors (NMDARs). NMDAR-dependent LTD prevails in juvenile mice, but a mechanistically different form of LTD can be readily induced in adults by instead stimulating metabotropic glutamate receptors (mGluRs). However, the role that CaMKII plays in the mGluR-dependent form of LTD is not clear. Here we show that mGluR-dependent LTD also requires CaMKII and its T286 autophosphorylation (pT286), which induces Ca2+-independent autonomous kinase activity. In addition, we compared the role of pT286 among three forms of long-term plasticity (NMDAR-dependent LTP and LTD, and mGluR-dependent LTD) using simultaneous live imaging of endogenous CaMKII together with synaptic marker proteins. We determined that after LTP stimuli, pT286 autophosphorylation accelerated CaMKII movement to excitatory synapses. After NMDAR-LTD stimuli, pT286 was strictly required for any movement to inhibitory synapses. Similar to NMDAR-LTD, we found the mGluR-LTD stimuli did not induce CaMKII movement to excitatory synapses. However, in contrast to NMDAR-LTD, we demonstrate that the mGluR-LTD did not involve CaMKII movement to inhibitory synapses and did not require additional T305/306 autophosphorylation. Thus, despite its prominent role in LTP, we conclude that CaMKII T286 autophosphorylation is also required for both major forms of hippocampal LTD, albeit with differential requirements for the heterosynaptic communication of excitatory signals to inhibitory synapses.

Integrative RNA profiling of TBEV-infected neurons and astrocytes reveals potential pathogenic effectors.

Tick-borne encephalitis virus (TBEV), the most medically relevant tick-transmitted flavivirus in Eurasia, targets the host central nervous system and frequently causes severe encephalitis. The severity of TBEV-induced neuropathogenesis is highly cell-type specific and the exact mechanism responsible for such differences has not been fully described yet. Thus, we performed a comprehensive analysis of alterations in host poly-(A)/miRNA/lncRNA expression upon TBEV infection in vitro in human primary neurons (high cytopathic effect) and astrocytes (low cytopathic effect). Infection with severe but not mild TBEV strain resulted in a high neuronal death rate. In comparison, infection with either of TBEV strains in human astrocytes did not. Differential expression and splicing analyses with an in silico prediction of miRNA/mRNA/lncRNA/vd-sRNA networks found significant changes in inflammatory and immune response pathways, nervous system development and regulation of mitosis in TBEV Hypr-infected neurons. Candidate mechanisms responsible for the aforementioned phenomena include specific regulation of host mRNA levels via differentially expressed miRNAs/lncRNAs or vd-sRNAs mimicking endogenous miRNAs and virus-driven modulation of host pre-mRNA splicing. We suggest that these factors are responsible for the observed differences in the virulence manifestation of both TBEV strains in different cell lines. This work brings the first complex overview of alterations in the transcriptome of human astrocytes and neurons during the infection by two TBEV strains of different virulence. The resulting data could serve as a starting point for further studies dealing with the mechanism of TBEV-host interactions and the related processes of TBEV pathogenesis.

Short Tandem Repeat Variation in the CNR1 Gene Associated With Analgesic Requirements of Opioids in Postoperative Pain Management.

Short tandem repeats (STRs) and variable number of tandem repeats (VNTRs) that have been identified at approximately 0.7 and 0.5 million loci in the human genome, respectively, are highly multi-allelic variations rather than single-nucleotide polymorphisms. The number of repeats of more than a few thousand STRs was associated with the expression of nearby genes, indicating that STRs are influential genetic variations in human traits. Analgesics act on the central nervous system via their intrinsic receptors to produce analgesic effects. In the present study, we focused on STRs and VNTRs in the CNR1, GRIN2A, PENK, and PDYN genes and analyzed two peripheral pain sensation-related traits and seven analgesia-related traits in postoperative pain management. A total of 192 volunteers who underwent the peripheral pain sensation tests and 139 and 252 patients who underwent open abdominal and orthognathic cosmetic surgeries, respectively, were included in the study. None of the four STRs or VNTRs were associated with peripheral pain sensation. Short tandem repeats in the CNR1, GRIN2A, and PENK genes were associated with the frequency of fentanyl use, fentanyl dose, and visual analog scale pain scores 3 h after orthognathic cosmetic surgery (Spearman's rank correlation coefficient ρ = 0.199, p = 0.002, ρ = 0.174, p = 0.006, and ρ = 0.135, p = 0.033, respectively), analgesic dose, including epidural analgesics after open abdominal surgery (ρ = -0.200, p = 0.018), and visual analog scale pain scores 24 h after orthognathic cosmetic surgery (ρ = 0.143, p = 0.023), respectively. The associations between STRs in the CNR1 gene and the frequency of fentanyl use and fentanyl dose after orthognathic cosmetic surgery were confirmed by Holm's multiple-testing correction. These findings indicate that STRs in the CNR1 gene influence analgesia in the orofacial region.

Treatment of Viral Encephalitis.

Viral encephalitis is difficult to treat. Herpes simplex encephalitis has been successfully treated with acyclovir, but is still a cause for significant morbidity even with that treatment. A rare form of autoimmune encephalitis related to NMDA receptor antibody after infection by herpes simplex can be treated with corticosteroid therapy. Arthropod-borne encephalitides, such as West Nile virus encephalitis and Eastern equine encephalitis, are primarily treated with supportive measures. Attempts have been made to use immunoglobulin therapy with limited effects. Progressive multifocal leukoencephalopathy has been treated with an emerging immune activation therapy in a limited number of patients with incomplete success.

A soluble derivative of PrPC activates cell-signaling and regulates cell physiology through LRP1 and the NMDA receptor.

Cellular prion protein (PrPC) is a widely expressed glycosylphosphatidylinositol-anchored membrane protein. Scrapie prion protein is a misfolded and aggregated form of PrPC responsible for prion-induced neurodegenerative diseases. Understanding the function of the nonpathogenic PrPC monomer is an important objective. PrPC may be shed from the cell surface to generate soluble derivatives. Herein, we studied a recombinant derivative of PrPC (soluble cellular prion protein, S-PrP) that corresponds closely in sequence to a soluble form of PrPC shed from the cell surface by proteases in the A Disintegrin And Metalloprotease (ADAM) family. S-PrP activated cell-signaling in PC12 and N2a cells. TrkA was transactivated by Src family kinases and extracellular signal-regulated kinase 1/2 was activated downstream of Trk receptors. These cell-signaling events were dependent on the N-methyl-d-aspartate receptor (NMDA-R) and low-density lipoprotein receptor-related protein-1 (LRP1), which functioned as a cell-signaling receptor system in lipid rafts. Membrane-anchored PrPC and neural cell adhesion molecule were not required for S-PrP-initiated cell-signaling. S-PrP promoted PC12 cell neurite outgrowth. This response required the NMDA-R, LRP1, Src family kinases, and Trk receptors. In Schwann cells, S-PrP interacted with the LRP1/NMDA-R system to activate extracellular signal-regulated kinase 1/2 and promote cell migration. The effects of S-PrP on PC12 cell neurite outgrowth and Schwann cell migration were similar to those caused by other proteins that engage the LRP1/NMDA-R system, including activated α2-macroglobulin and tissue-type plasminogen activator. Collectively, these results demonstrate that shed forms of PrPC may exhibit important biological activities in the central nervous system and the peripheral nervous system by serving as ligands for the LRP1/NMDA-R system.

The NMDA receptor subunit GluN3A regulates synaptic activity-induced and myocyte enhancer factor 2C (MEF2C)-dependent transcription.

N-Methyl-d-aspartate type glutamate receptors (NMDARs) are key mediators of synaptic activity-regulated gene transcription in neurons, both during development and in the adult brain. Developmental differences in the glutamate receptor ionotropic NMDA 2 (GluN2) subunit composition of NMDARs determines whether they activate the transcription factor cAMP-responsive element-binding protein 1 (CREB). However, whether the developmentally regulated GluN3A subunit also modulates NMDAR-induced transcription is unknown. Here, using an array of techniques, including quantitative real-time PCR, immunostaining, reporter gene assays, RNA-Seq, and two-photon glutamate uncaging with calcium imaging, we show that knocking down GluN3A in rat hippocampal neurons promotes the inducible transcription of a subset of NMDAR-sensitive genes. We found that this enhancement is mediated by the accumulation of phosphorylated p38 mitogen-activated protein kinase in the nucleus, which drives the activation of the transcription factor myocyte enhancer factor 2C (MEF2C) and promotes the transcription of a subset of synaptic activity-induced genes, including brain-derived neurotrophic factor (Bdnf) and activity-regulated cytoskeleton-associated protein (Arc). Our evidence that GluN3A regulates MEF2C-dependent transcription reveals a novel mechanism by which NMDAR subunit composition confers specificity to the program of synaptic activity-regulated gene transcription in developing neurons.

GluN2A-NMDA receptor-mediated sustained Ca2+ influx leads to homocysteine-induced neuronal cell death.

Homocysteine, a metabolite of the methionine cycle, is a known agonist of N-methyl-d-aspartate receptor (NMDAR), a glutamate receptor subtype and is involved in NMDAR-mediated neurotoxicity. Our previous findings have shown that homocysteine-induced, NMDAR-mediated neurotoxicity is facilitated by a sustained increase in phosphorylation and activation of extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK MAPK). In the current study, we investigated the role GluN1/GluN2A-containing functional NMDAR (GluN2A-NMDAR) and GluN1/GluN2B-containing functional NMDAR (GluN2B-NMDAR) in homocysteine-induced neurotoxicity. Our findings revealed that exposing primary cortical neuronal cultures to homocysteine leads to a sustained low-level increase in intracellular Ca2+ We also showed that pharmacological inhibition of GluN2A-NMDAR or genetic deletion of the GluN2A subunit attenuates homocysteine-induced increase in intracellular Ca2+ Our results further established the role of GluN2A-NMDAR in homocysteine-mediated sustained ERK MAPK phosphorylation and neuronal cell death. Of note, the preferential role of GluN2A-NMDAR in homocysteine-induced neurotoxicity was distinctly different from glutamate-NMDAR-induced excitotoxic cell death that involves overactivation of GluN2B-NMDAR and is independent of ERK MAPK activation. These findings indicate a critical role of GluN2A-NMDAR-mediated signaling in homocysteine-induced neurotoxicity.

Glutaminergic tonic action potentiate MPP+-induced hydroxyl radical production in rat striatum.

The effect of glycine on 1-methyl-4-phenylpyridinium ion (MPP+)-induced hydroxyl radical (OH) formation in the extracellular fluid of rat striatum were investigated. Rats were anesthetized and sodium salicylate in Ringer's solution (0.5 nmol/µl/min) was infused through a microdialysis probe to detect the generation of OH as reflected by the non-enzymatic formation of 2,3-dihydroxybenzoic acid (2,3-DHBA) in rat striatum. MPP+ (5 mmol/L) produced an increase in OH formation. When glycine (1 mmol/L) was infused into the rat striatum through a microdialysis probe after MPP+ treatment, the marked in the level of 2,3-DHBA was observed in the brain dialysate. However, in the presence of MK-801 (100 µmol/L), a non competitive antagonist of N-methyl-D-aspartate (NMDA), glycine failed to increase the 2,3-DHBA formation by MPP+. When corresponding experiments were performed with nitro-L arginine (L-NNA) (1 mmol/L), a nitric oxide synthase (NOS) inhibitor, same result was obtained. These results suggest that MPP+-induced OH generation may modulated by glycine via NMDA receptor in rat striatum. This increase might be explained because of the presence of a glutaminergic tonic action.

Glutamate ionotropic receptor NMDA type subunit 2A (GRIN2A) gene polymorphism (rs4998386) and Parkinson's disease susceptibility: A meta-analysis.

OBJECTIVE: Dopaminergic neuronal degeneration seen in Parkinson's disease (PD) might result from a single nucleotide polymorphism (SNP) in the glutamate ionotropic receptor NMDA type subunit 2A (GRIN2A) gene. We thus performed a meta-analysis exploring the relationship between the rs4998386 SNP of the GRIN2A gene and PD susceptibility. METHODS: We searched PubMed, EMBASE, Web of Science, Google Scholar, and China National Knowledge Infrastructure for studies published between January 2005 and January 2019. The association between the rs4998386 polymorphism and PD susceptibility was evaluated by calculating the pooled odds ratios (ORs) and 95% confidence intervals (CIs). RESULTS: Meta-analysis results did not show a significant association between the rs4998386 polymorphism of the GRIN2A gene and PD susceptibility when assuming an allelic model (OR, 0.90; 95% CI, 0.76-1.07; P = .22; I 2 = 53%), a dominant model (OR, 0.96; 95% CI, 0.82-1.12; P = .62; I 2 = 64%), or a recessive model (OR, 1.14; 95% CI, 0.93-1.38; P = .22; I 2 = 0%). CONCLUSION: Our meta-analysis found that the rs4998386 polymorphism of the GRIN2A gene is not associated with risk of PD in either Europeans or white Americans. However, large sample studies with different ethnicities should be conducted to establish the role of the rs4998386 polymorphism in PD pathophysiology.

Diagnostics of autoimmune encephalitis associated with antibodies against neuronal surface antigens.

This document presents the guidelines for testing antibodies against neuronal surface antigens that have been developed following a consensus process built on questionnaire-based surveys, internet contacts, and discussions at workshops of the sponsoring Italian Association of Neuroimmunology (AINI) congresses. Essential clinical information on autoimmune encephalitis associated with antibodies against neuronal surface antigens, indications and limits of testing for such antibodies, instructions for result interpretation, and an agreed laboratory protocol (Appendix A) are reported for the communicative community of neurologists and clinical pathologists.

Distribution, properties, and inhibitor sensitivity of zebrafish catechol-O-methyl transferases (COMT).

Catechol-O-methyltransferase (COMT; EC 2.1.1.6) is an enzyme with multiple functions in vertebrates. COMT methylates and thus inactivates catecholamine neurotransmitters and metabolizes xenobiotic catechols. Gene polymorphism rs4680 that influences the enzymatic activity of COMT affects cognition and behavior in humans. The zebrafish is widely used as an experimental animal in many areas of biomedical research, but most aspects of COMT function in this species have remained uncharacterized. We hypothesized that both comt genes play essential roles in zebrafish. Both comt-a and comt-b were widely expressed in zebrafish tissues, but their relative abundance varied considerably. Homogenates of zebrafish organs, including the brain, showed enzymatic COMT activity that was the highest in the liver and kidney. Treatment of larval zebrafish with the COMT inhibitor Ro41-0960 shifted the balance of catecholamine metabolic pathways towards increased oxidative metabolism. Whole-body concentrations of dioxyphenylacetic acid (DOPAC), a product of dopamine oxidation, were increased in the inhibitor-treated larvae, although the dopamine levels were unchanged. Thus, COMT is likely to participate in the processing of catecholamine neurotransmitters in the zebrafish, but the inhibition of COMT in larval fish is compensated efficiently and does not have pronounced effects on dopamine levels.

Determination of dendritic spine morphology by the striatin scaffold protein STRN4 through interaction with the phosphatase PP2A.

Dendritic spines are heterogeneous and exist with various morphologies. Altered spine morphology might underlie the cognitive deficits in neurodevelopmental disorders such as autism, but how different subtypes of dendritic spines are selectively maintained along development is still poorly understood. Spine maturation requires spontaneous activity of N-methyl-d-aspartate (NMDA) receptor and local dendritic protein synthesis. STRN4 (also called zinedin) belongs to the striatin family of scaffold proteins, and some of the potential striatin-interacting proteins are encoded by autism risk genes. Although previous studies have demonstrated their localization in dendritic spines, the function of various striatin family members in the neuron remains unknown. Here, we demonstrate that Strn4 mRNA is present in neuronal dendrites, and the local expression of STRN4 protein depends on NMDA receptor activation. Notably, STRN4 is preferentially expressed in mushroom spines, and STRN4 specifically maintains mushroom spines but not thin spines and filopodia through interaction with the phosphatase PP2A. Our findings have therefore unraveled the local expression of STRN4 as a novel mechanism for the control of dendritic spine morphology.

Amyloid-ß oligomers transiently inhibit AMP-activated kinase and cause metabolic defects in hippocampal neurons.

AMP-activated kinase (AMPK) is a key player in energy sensing and metabolic reprogramming under cellular energy restriction. Several studies have linked impaired AMPK function to peripheral metabolic diseases such as diabetes. However, the impact of neurological disorders, such as Alzheimer disease (AD), on AMPK function and downstream effects of altered AMPK activity on neuronal metabolism have been investigated only recently. Here, we report the impact of Aß oligomers (AßOs), synaptotoxins that accumulate in AD brains, on neuronal AMPK activity. Short-term exposure of cultured rat hippocampal neurons or ex vivo human cortical slices to AßOs transiently decreased intracellular ATP levels and AMPK activity, as evaluated by its phosphorylation at threonine residue 172 (AMPK-Thr(P)172). The AßO-dependent reduction in AMPK-Thr(P)172 levels was mediated by glutamate receptors of the N-methyl-d-aspartate (NMDA) subtype and resulted in removal of glucose transporters (GLUTs) from the surfaces of dendritic processes in hippocampal neurons. Importantly, insulin prevented the AßO-induced inhibition of AMPK. Our results establish a novel toxic impact of AßOs on neuronal metabolism and suggest that AßO-induced, NMDA receptor-mediated AMPK inhibition may play a key role in early brain metabolic defects in AD.

Spontaneous Glutamatergic Synaptic Activity Regulates Constitutive COX-2 Expression in Neurons: OPPOSING ROLES FOR THE TRANSCRIPTION FACTORS CREB (cAMP RESPONSE ELEMENT BINDING) PROTEIN AND Sp1 (STIMULATORY PROTEIN-1).

Burgeoning evidence supports a role for cyclooxygenase metabolites in regulating membrane excitability in various forms of synaptic plasticity. Two cyclooxygenases, COX-1 and COX-2, catalyze the initial step in the metabolism of arachidonic acid to prostaglandins. COX-2 is generally considered inducible, but in glutamatergic neurons in some brain regions, including the cerebral cortex, it is constitutively expressed. However, the transcriptional mechanisms by which this occurs have not been elucidated. Here, we used quantitative PCR and also analyzed reporter gene expression in a mouse line carrying a construct consisting of a portion of the proximal promoter region of the mouse COX-2 gene upstream of luciferase cDNA to characterize COX-2 basal transcriptional regulation in cortical neurons. Extracts from the whole brain and from the cerebral cortex, hippocampus, and olfactory bulbs exhibited high luciferase activity. Moreover, constitutive COX-2 expression and luciferase activity were detected in cortical neurons, but not in cortical astrocytes, cultured from wild-type and transgenic mice, respectively. Constitutive COX-2 expression depended on spontaneous but not evoked excitatory synaptic activity and was shown to be N-methyl-d-aspartate receptor-dependent. Constitutive promoter activity was reduced in neurons transfected with a dominant-negative cAMP response element binding protein (CREB) and was eliminated by mutating the CRE-binding site on the COX-2 promoter. However, mutation of the stimulatory protein-1 (Sp1)-binding site resulted in an N-methyl-d-aspartate receptor-dependent enhancement of COX-2 promoter activity. Basal binding of the transcription factors CREB and Sp1 to the native neuronal COX-2 promoter was confirmed. In toto, our data suggest that spontaneous glutamatergic synaptic activity regulates constitutive neuronal COX-2 expression via Sp1 and CREB protein-dependent transcriptional mechanisms.

Functions of Polyamines in Mammals.

The content of spermidine and spermine in mammalian cells has important roles in protein and nucleic acid synthesis and structure, protection from oxidative damage, activity of ion channels, cell proliferation, differentiation, and apoptosis. Spermidine is essential for viability and acts as the precursor of hypusine, a post-translational addition to eIF5A allowing the translation of mRNAs encoding proteins containing polyproline tracts. Studies with Gy mice and human patients with the very rare X-linked genetic condition Snyder-Robinson syndrome that both lack spermine synthase show clearly that the correct spermine:spermidine ratio is critical for normal growth and development.

The CaMKII/GluN2B Protein Interaction Maintains Synaptic Strength.

Learning, memory, and cognition are thought to require normal long-term potentiation (LTP) of synaptic strength, which in turn requires binding of the Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) to the NMDA-type glutamate receptor (NMDAR) subunit GluN2B. For LTP induction, many additional required players are known. Here we tested the hypothesis that CaMKII/GluN2B binding also mediates the more elusive maintenance of synaptic strength. Intriguingly, the CaMKII inhibitor tatCN21 reduces synaptic strength only at high concentrations necessary for CaMKII/NMDAR disruption (20 µm) but not at lower concentrations sufficient for kinase inhibition (5 µm). However, increased concentration also causes unrelated effects. Thus, to distinguish between correlation and causality, we used a pharmacogenetic approach. In a mouse with a mutant NMDAR GluN2B subunit that is CaMKII binding-incompetent, any tatCN21 effects that are specific to the CaMKII/GluN2B interaction should be abolished, and any remaining tatCN21 effects have to be nonspecific (i.e. mediated by other targets). The results showed that the persistent reduction of synaptic strength by transient application of 20 µm tatCN21 had a nonspecific presynaptic component (on fiber volley amplitude) that was unrelated to the CaMKII/GluN2B interaction or CaMKII activity. However, the remaining component of the persistent tatCN21 effect was almost completely abolished in the GluN2B mutant mouse. These results highlight the requirement for stringent pharmacogenetic approaches to separate specific on-target effects from nonspecific off-target effects. Importantly, they also demonstrate that the CaMKII/GluN2B interaction is required not only for normal LTP induction but also for the maintenance of synaptic strength.

Glutamate in peripheral organs: Biology and pharmacology.

Glutamate is a versatile molecule existing in both the central nervous system and peripheral organs. Previous studies have mainly focussed on the biological effect of glutamate in the brain. Recently, abundant evidence has demonstrated that glutamate also participates in the regulation of physiopathological functions in peripheral tissues, including the lung, kidney, liver, heart, stomach and immune system, where the glutamate/glutamate receptor/glutamate transporter system plays an important role in the pathogenesis of certain diseases, such as myocardial ischaemia/reperfusion injury and acute gastric mucosa injury. All these findings provide new insight into the biology and pharmacology of glutamate and suggest a potential therapeutic role of glutamate in non-neurological diseases.