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We previously demonstrated that the blockade of mGluR5 by 2-methyl-6(phenylethynyl)pyridine (MPEP) reduces both cold and warm ischemia/reperfusion injury. Here we evaluated whether MPEP reduces the hepatic preservation injury in rat livers from cardiac-death-donors (DCDs). Livers from DCD rats were isolated after an in situ warm ischemia (30 min) and preserved for 22 h at 4 °C with UW solution. Next, 10 mg/Kg MPEP or vehicle were administered 30 min before the portal clamping and added to the UW solution (3 µM). LDH released during washout was quantified. Liver samples were collected for iNOS, eNOS, NO, TNF-α, ICAM-1, caspase-3 and caspase-9 protein expression and nuclear factor-erythroid-2-related factor-2 (Nrf2) gene analysis. Lower LDH levels were detected in control grafts versus DCD groups. An increase in eNOS and NO content occurred after MPEP treatment; iNOS and TNF-α content was unchanged. ICAM-1 expression was reduced in the MPEP-treated livers as well as the levels of caspase-3 and caspase-9. Nrf2, oxidative stress-sensitive gene, was recovered to control value by MPEP. These results suggest that MPEP can be used to reclaim DCD livers subjected to an additional period of cold ischemia during hypothermic storage. MPEP protects against apoptosis and increased eNOS, whose overexpression has been previously demonstrated to be protective in hepatic ischemia/reperfusion damage.

Objective: To understand the involvement of the mGluR5-mediated JNK signaling pathway in rats with diabetic retinopathy (DR). Methods: This study established rat models of diabetes mellitus (DM), which were divided into Normal, DM, DM + CHPG (mGluR5 agonist CHPG), and DM + MTEP (mGluR5 antagonist MTEP) groups. The blood glucose and weight of rats were recorded. EB staining was used for observation of blood–retinal barrier (BRB) damage. Neural retina function was measured by pattern electroretinogram (ERG). PAS and NG2 immunohistochemistry were conducted to evaluate the retinal vascular morphology. The TUNEL assay and active caspase-3 immunohistochemistry were performed to detect retinal cell apoptosis. Additionally, the expression levels of superoxide dismutase (SOD) and methylenedioxyamphetamine (MDA) were measured. Moreover, expression levels of mGluR5 and JNK pathway-related proteins were detected by western blot. Results: When compared with control rats, rats in the DM group showed decreased amplitude and latency of the peak times in the ERG test; further, DM group rats presented increases in blood glucose, BRB permeability, a retinal capillary area density, retinal cell apoptosis with an increased number of active caspase-3-positive cells, MDA level, mGluR5 levels, and the ratio of p-JNK/JNK, and they showed reductions in body weight and SOD activity, as well as in the number of pericytes and in the pericyte coverage (all P < 0.05). However, rats in DM + CHPG group had stronger negative effects than those in DM group (all P < 0.05). Rats from DM + MTEP group showed an opposite trend compared with the DM rats (all P < 0.05). Conclusion: The level of mGluR5 in DR rats was upregulated, whereas inhibition of mGluR5 alleviated retinal pathological damage and decreased cell apoptosis to improve DR via suppression of the JNK signaling pathway, which provided a scientific theoretical basis for the clinical treatment of DR. [ABSTRACT FROM AUTHOR]

mGluR5-containing synapses have essential roles in synaptic plasticity, circuit physiology, and learning, and dysfunction at these synapses is implicated in specific neurological disorders. As mGluR5-containing synapses are embedded in large and complex distributed circuits containing many neuron and synapse types, it is challenging to elucidate the roles of these synapses and to develop treatments for the associated disorders. Thus, it would be advantageous to deliver different genes into pre- and postsynaptic neurons connected by a mGluR5-containing synapse. Here, we develop this capability: The first gene transfer, into the presynaptic neurons, uses standard techniques to deliver a vector that expresses a synthetic peptide neurotransmitter. This peptide neurotransmitter has three domains: a dense core vesicle sorting domain, a mGluR5-binding domain composed of a single-chain variable fragment anti-mGluR5, and the His tag. Upon release, this peptide neurotransmitter binds to mGluR5, predominately located on the postsynaptic neurons. Selective gene transfer into these neurons uses antibody-mediated, targeted gene transfer and anti-His tag antibodies, as the synthetic peptide neurotransmitter contains the His tag. For the model system, we studied the connection between neurons in two neocortical areas: postrhinal and perirhinal cortices. Targeted gene transfer was over 80% specific for mGluR5-containing synapses, but untargeted gene transfer was only ~ 15% specific for these synapses. This technology may enable studies on the roles of mGluR5-containing neurons and synapses in circuit physiology and learning and support gene therapy treatments for specific disorders that involve dysfunction at these synapses. [ABSTRACT FROM AUTHOR]

Highlights • Repeated alcohol administration produces robust locomotor sensitization in male DBA/2J mice. • Treatment with the mGluR5 antagonist MPEP blunts the induction of sensitization. • Sensitization is associated with changes in ERK1/2 phosphorylation in multiple brain regions. • MPEP administered during induction blocked alcohol-induced changes in pERK1/2 in nucleus accumbens shell and lateral habenula. • mGluR5-ERK1/2 signaling may be required for the induction of alcohol sensitization. Abstract Metabotropic glutamate receptor subtype-5 (mGluR5) activity regulates a variety of behavioral pathologies associated with alcohol addiction. The main goal of this study was to determine if mGluR5 regulates the induction of ethanol-induced locomotor sensitization, which is a model of experience-dependent plasticity following initial exposure to drugs of abuse. The extracellular signal-regulated kinase (ERK 1/2) pathway is downstream of mGluR5 and implicated in alcohol addiction; however, its role in sensitization remains unexplored. We sought to determine if mGluR5-mediated changes in ethanol-induced sensitization are associated with changes in ERK 1/2 phosphorylation (pERK 1/2) in specific brain regions. Adult male DBA/2 J mice were tested for acute locomotor response to ethanol (0 or 2 g/kg, IP) followed by a 9-day induction period in which the mGluR5 antagonist MPEP (0 or 30 mg/kg, IP) was administered prior to ethanol (0 or 2.5 g/kg, IP). One day later, ethanol (2 g/kg) produced a robust within- and between-group increase in locomotor activity, indicating sensitization in mice that received MPEP (0 mg/kg) during induction. MPEP (30 mg/kg) treatment during induction resulted in locomotor response to ethanol (2 g/kg) challenge that was equivalent to an acute response, indicating full blockade of sensitization. Sensitization was associated with increased pERK 1/2 immunoreactivity (IR) in nucleus accumbens shell (AcbSh) and a reduction in lateral habenula (LHb), both of which were blocked by MPEP treatment during induction. Sensitization was also associated with mGluR5-independent increases in pERK 1/2 IR in the nucleus accumbens core and decreases in the dentate gyrus and lateral septum. These data indicate that mGluR5 activity is required for the induction of ethanol locomotor sensitization and associated changes in ERK 1/2 phosphorylation in the AcbSh and LHb, which raises the hypothesis that mGluR5-mediated cell signaling in these brain regions may mediate the induction of sensitization. Elucidating mechanisms of sensitization may increase understanding of how ethanol hijacks behavioral functions during the development of addiction. [ABSTRACT FROM AUTHOR]

• Whole body mGluR5 KO mice displays antidepressant effects. • Proinflammatory cytokines are decreased in bone marrows from mGluR5 KO mice. • Transplanting mGluR5 KO bone marrow to WT mice displays antidepressant effects by decreasing cytokines in serum and CSF. • Inhibited cytokines decrease activation of brain endothelial cells in ERK dependent manner. Antidepressant-like effects of metabotropic glutamate receptor 5 (mGluR5) have been verified by specific antagonists or whole body knock-out (KO) mice. Previous experiments indicate that blocking mGluR5 exerts antidepressant-like effects through neuronal mechanisms, like modulating NMDA receptor activity or 5-HT system. Here we found that transplanting bone marrow from mGluR5 KO mice to WT mice could also show antidepressant-like effects, which were confirmed by sucrose preference test and tail suspension test. Furthermore, mGluR5 deficiency dramatically inhibits cytokines release from bone marrow cells, such as IL-1β, TNF-α and IL-6, alleviating proinflammatory responses in LPS-induced depression model. In addition, inhibited cytokines could decrease the activation of brain endothelial cells in ERK-dependent manner. These data provide the evidence that blocking mGluR5 could improve depression through inhibiting peripheral immune responses, confirming the causal relationship between peripheral immune phenotype and brain behavior. [ABSTRACT FROM AUTHOR]

Targeting mGluR5 has been an attractive strategy to modulate glutamate excitotoxicity for neuroprotection. Although human clinical trials using mGluR5 negative allosteric modulators (NAMs) have included some disappointments, recent investigations have added several more attractive small molecules to this field, providing a promise that the identification of more additional strategies to modulate mGluR5 activity might be potentially beneficial for the advancement of PD treatment. Here, we determined the role of the interacting partner CAL (cystic fibrosis transmembrane conductance regulator-associated ligand) in mGluR5-mediated protection in vitro and in vivo. In astroglial C6 cells, CAL deficiency blocked (S)-3, 5-dihydroxyphenylglycine (DHPG)-elicited p-AKT and p-ERK1/2, subsequently prevented group I mGluRs-mediated anti-apoptotic protection, which was blocked by receptor antagonist 1-aminoindan-1, 5-dicarboxylic acid (AIDA), and PI3K or MEK inhibitor LY294002 or U0126. In rotenone-treated MN9D cells, both CAL and mGluR5 expressions were decreased in a time- and dose-dependent manner, and the correlation between these 2 proteins was confirmed by lentivirus-delivered CAL overexpression and knockdown. Moreover, CAL coupled with mGluR5 upregulated mGluR5 protein expression by inhibition of ubiquitin-proteasome-dependent degradation to suppress mGluR5-mediated p-JNK and to protect against cell apoptosis. Additionally, CAL also inhibited rotenone-induced glutamate release to modulate mGluR5 activity. Furthermore, in the rotenone-induced rat model of PD, AAV-delivered CAL overexpression attenuated behavioral deficits and dopaminergic neuronal death, while CAL deficiency aggravated rotenone toxicity. On the other hand, the protective effect of the mGluR5 antagonist MPEP was weakened by knocking down CAL. In vivo experiments also confirmed that CAL inhibited ubiquitination-proteasome-dependent degradation to modulate mGluR5 expression and JNK phosphorylation. Our findings show that CAL protects against cell apoptosis via modulating mGluR5 activity, and may be a new molecular target for an effective therapeutic strategy for PD. [ABSTRACT FROM AUTHOR]

Hypofunction of NMDA receptors has been considered a possible cause for the pathophysiology of schizophrenia. More recently, indirect ways to regulate NMDA that would be less disruptive have been proposed and metabotropic glutamate receptor subtype 5 (mGluR5) represents one such candidate. To characterize the cell populations involved, we demonstrated here that knock-out (KO) of mGluR5 in cholinergic, but not glutamatergic or parvalbumin (PV)-positive GABAergic, neurons reduced prepulse inhibition of the startle response (PPI) and enhanced sensitivity to MK801-induced locomotor activity. Inhibition of cholinergic neurons in the medial septum by DREADD (designer receptors exclusively activated by designer drugs) resulted in reduced PPI further demonstrating the importance of these neurons in sensorimotor gating. Volume imaging and quantification were used to compare PV and cholinergic cell distribution, density, and total cell counts in the different cell-type-specific KO lines. Electrophysiological studies showed reduced NMDA receptormediated currents in cholinergic neurons of the medial septum in mGluR5 KO mice. These results obtained from male and female mice indicate that cholinergic neurons in the medial septum represent a key cell type involved in sensorimotor gating and are relevant to pathologies associated with disrupted sensorimotor gating such as schizophrenia. [ABSTRACT FROM AUTHOR]

Recent evidence implicates dysregulation of metabotropic glutamatergic receptor 5 (mGluR5) in pathophysiology of PTSD and suicidality. Using positron emission tomography and [18F]FPEB, we quantified mGluR5 availability in vivo in individuals with PTSD (n = 29) and MDD (n = 29) as a function of suicidal ideation (SI) to compare with that of healthy comparison controls (HC; n = 29). Volume of distribution was computed using a venous input function in the five key frontal and limbic brain regions. We observed significantly higher mGluR5 availability in PTSD compared with HC individuals in all regions of interest (P's = 0.001-0.01) and compared with MDD individuals in three regions (P's = 0.007). mGluR5 availability was not significantly different between MDD and HC individuals (P = 0.17). Importantly, we observed an up-regulation in mGluR5 availability in the PTSD-SI group (P's = 0.001-0.007) compared with PTSD individuals without SI. Findings point to the potential role for mGluR5 as a target for intervention and, potentially, suicide risk management in PTSD. [ABSTRACT FROM AUTHOR]

l-DOPA is the main pharmacological therapy for Parkinson's disease. However, long-term exposure to l-DOPA induces involuntary movements termed dyskinesia. Clinical trials show that dyskinesia is attenuated by metabotropic glutamate receptor type 5 (mGluR5) antagonists. Further, the onset of dyskinesia is delayed by nicotine and mGluR5 expression is lower in smokers than in non-smokers. However, the mechanisms by which mGluR5 modulates dyskinesia and how mGluR5 and nicotine interact have not been established. To address these issues, we studied the role of mGluR5 in D1R-containing neurons in dyskinesia and examined whether nicotine reduces dyskinesia via mGluR5. In the aphakia mouse model of Parkinson's disease, we selectively knocked down mGluR5 in D1R-containing neurons (aphakia-mGluR5KD-D1). We found that genetic downregulation of mGluR5 decreased dyskinesia in aphakia mice. Although chronic nicotine increased the therapeutic effect of l-DOPA in both aphakia and aphakia-mGluR5KD-D1 mice, it caused a robust reduction in dyskinesia only in aphakia, and not in aphakia-mGluR5KD-D1 mice. Downregulating mGluR5 or nicotine treatment after l-DOPA decreased ERK and histone 3 activation, and FosB expression. Combining nicotine and mGluR5 knockdown did not have an added antidyskinetic effect, indicating that the effect of nicotine might be mediated by downregulation of mGluR5 expression. Treatment of aphakia-mGluR5KD-D1 mice with a negative allosteric modulator did not further modify dyskinesia, suggesting that mGluR5 in non-D1R-containing neurons does not play a role in its development. In conclusion, this work suggests that mGluR5 antagonists reduce dyskinesia by mainly affecting D1R-containing neurons and that the effect of nicotine on dyskinetic signs in aphakia mice is likely via mGluR5. [ABSTRACT FROM AUTHOR]

Highlights • mGluR1/5 binding is increased in the putamen of Cloninger type 1 and 2 alcoholics. • mGluR1/5 binding is increased in the caudate head of both alcoholic subtypes. • mGluR1/5 binding is increased also in the anterior insula of both alcoholic subtypes. • In the hippocampus, mGluR1/5 binding is increased in Cloninger type 2 alcoholics. Abstract The function of group I metabotropic glutamate receptors mGluR1 and mGluR5 is involved in the hyperglutamatergic state caused by chronic alcohol. Preclinical studies suggest that group I mGluR modulation could serve as a novel treatment of alcoholism. Considering the wide role of glutamatergic neurochemistry in addiction, group I mGluR binding was studied in brain areas involved in decision-making, learning and memory. Post-mortem whole hemisphere autoradiography was used to study the binding density of [³H]quisqualic acid, a potent group I mGluR agonist, in 9 Cloninger type 1 alcoholics, 8 Cloninger type 2 alcoholics and 10 controls. Binding was studied in the dorsal striatum, hippocampus and cortex. Alcoholics displayed a trend towards increased [³H]quisqualic acid binding in all brain areas. The most robust findings were in the putamen (p = 0.006) and anterior insula (p = 0.005), where both alcoholic subtypes displayed increased binding compared to the controls. These findings suggest altered group I mGluR function in alcoholic subjects in the dorsal striatum, which is involved in habitual learning, and in the anterior insula, which has a pivotal role in the perception of bodily sensations. Increased [³H]quisqualic acid binding might suggest a beneficial impact of mGluR1/5 modulators in the treatment of alcoholism. [ABSTRACT FROM AUTHOR]

Purpose: The purpose of this study was to assess, in a large sample of healthy young adults, sex differences in the binding potential of [11C]ABP688, a positron emission tomography (PET) tracer selective for the metabotropic glutamate type 5 (mGlu5) receptor. Methods: High resolution [11C]ABP688 PET scans were acquired in 74 healthy volunteers (25 male, 49 female, mean age 20 ± 3.0). Mean binding potential (BPND = fND * (Bavail / KD)) values were calculated in the prefrontal cortex, striatum, and limbic regions using the simplified reference tissue model with cerebellar grey matter as the reference region. Results: [11C]ABP688 BPND was significantly higher in men compared to women in the prefrontal cortex (p < 0.01), striatum (p < 0.001), and hippocampus (p < 0.05). Whole-brain BPND was 17% higher in men. BPND was not related to menstrual phase in women. Conclusions: Binding availability of mGlu5 receptors as measured by PET [11C]ABP688 is higher in healthy men than women. This likely represents a source of variability in [11C]ABP688 studies and could have relevance for sex differences in cognitive-behavioral functions and neuropsychiatric disorders. [ABSTRACT FROM AUTHOR]

Purpose: To determine how the low-affinity (Z)-isomer of the radiotracer [11C]ABP688 affects binding potential values in vivo in humans. Methods: High-resolution [11C]ABP688 PET scans were acquired on 74 healthy volunteers (25 male, 49 female, mean age 20 ± 3.0). The relative contents of (E)- and (Z)-isomers were determined prior to injection using analytical high-performance liquid chromatography [rt(E) = 10 min, rt(Z) = 8.5 min]. Mean binding potential [BPND = fND * (Bavail/KD)] values were calculated in the striatum, limbic regions, and prefrontal cortex using the simplified reference tissue model with cerebellar grey matter as reference. Results: Mean ± SD (E)-isomer content in [11C]ABP688 production was 92 ± 3.8% (range 78–97%). Percent (E)-isomer was positively correlated with BPND in the striatum (ρ = 0.28, p = 0.015) and limbic regions (ρ = 0.25, p = 0.036). In multiple regression analysis, sex (β = 0.39, p = 0.001) and (E)-isomer content (β = 0.23, p = 0.040) were significant predictors of BPND. Conclusions: Even modest levels of (Z)-[11C]ABP688 can reduce estimates of tracer binding in vivo. Future studies should use production methods that enrich levels of (E)-[11C]ABP688, report tracer isomer ratios, and account for this factor in their analyses. [ABSTRACT FROM AUTHOR]

Abstract Previous work has demonstrated that neuroimaging biomarkers which capture functional connectivity of the brain can be used to define a specific and robust endophenotype in Fmr1 -/y mice, a well-established animal model of human Fragile-X Syndrome (FXS). However, it is currently unknown whether this macroscopic measure of brain connectivity is sufficiently sensitive to reliably detect changes caused by pharmacological interventions. Here we inhibited the activity of the metabotropic glutamate receptor-5 (mGluR5) using AFQ056/Mavoglurant, a drug that is assumed to normalize excitatory/inhibitory neural signaling imbalances in FXS. We employed resting-state-fMRI (rs-fMRI) and diffusion-weighted imaging (DWI) to test whether Mavoglurant re-established brain connectivity - at least partly - within some of the affected circuits in Fmr1 -/y mice that are related to social behavior deficits. In line with previous findings, we observed that Fmr1 -/y mice exhibited impaired social interaction, reduced connectivity in three main functional networks and altered network topology. At the group level, Mavoglurant did neither rescue abnormal social behavioral nor white matter abnormalities; however, for some, but not all of these circuits Mavoglurant had a genotype-specific effect of restoring functional connectivity. These results show that rs-fMRI connectivity is sufficiently sensitive to pick up system-level changes after the pharmacological inhibition of mGluR5 activity. However, our results also show that the effects of Mavoglurant are confined to specific networks suggesting that behavioral benefits might be restricted to narrow functional domains. [ABSTRACT FROM AUTHOR]

Many PET tracers enable determination of fluctuations in neurotransmitter release, yet glutamate specifically can not be visualized in a noninvasive manner. Several studies point to the possibility of visualizing fluctuations in glutamate release by changes in affinity of the mGluR5 radioligand [11C]ABP688. These studies use pharmacological challenges to alter glutamate levels, and so probe release, but have not measured chronic alterations in receptor occupancy due to altered neurotransmission relevant to chronic neuropsychiatric disorders or their treatment. In this regard, the GLS1 heterozygous mouse has known reductions in activity of the glutamate-synthetic enzyme glutaminase, brain glutamate levels and release. We imaged this model to elucidate glutamatergic systems. Dynamic [11C]ABP688 microPET scans were performed for mGluR5. Western blot was used as an ex vivo validation. No significant differences were found in BPND between WT and GLS1 Hets. SPM showed voxel-wise increased in BPND in GLS1 Hets compared to WT consistent with lower synaptic glutamate. This was not due to alterations in mGluR5 levels, as western blot results showed lower mGluR5 levels in GLS1 Hets. We conclude that because of the chronic glutaminase deficiency and subsequent decrease in glutamate, the mGluR5 protein levels are lowered. Due to these decreased endogenous glutamate levels, however, there is increased [11C]ABP688 binding to the allosteric site in selected regions. We speculate that lower endogenous glutamate leads to less conformational change to the receptors, and thus higher availability of the binding site. The lower mGluR5 levels, however, lessen [11C]ABP688 binding in GLS1 Hets, in part masking the increase in binding due to diminished endogenous glutamate levels as confirmed with voxel-wise analysis. [ABSTRACT FROM AUTHOR]

There are limited studies focused on the precise mechanism of gonadotropin-releasing hormone (GnRH) secretion dysfunction after overexposure to manganese (Mn). The objective of the present study was to explore the mechanism of Mn disruption of GnRH synthesis via nuclear factor erythroid-2-related factor-2 (Nrf2)/metabotropic glutamate receptor-5 (mGluR5)/cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2) signaling pathway in vitro and in vivo. Primary astrocytes were cultured and treated with different doses of Mn, tert-butylhydroquinonet (tBHQ; Nrf2 agonists), 3-[(2-methyl-4-thaizolyl) ethynyl] pyridine (MTEP; mGluR5 inhibitor), and celecoxib (COX-2 inhibitor) to measure the levels of COX-2, mGluR5, Nrf2, and Nrf2 target genes. Mice were randomly divided into 11 groups, of which included the control group, 12.5-, 25-, and 50-mg/kg MnCl2 group, dimethyl sulfoxide (DMSO) group, tBHQ control group, tBHQ pretreatment group, MTEP control group, MTEP pretreatment group, celecoxib control group, and celecoxib pretreatment group. The injection was administered every day for 2 weeks. Then, levels of GnRH, PGE2, COX-2, mGluR5, Nrf2, Nrf2 target genes, and morphological changes in the hypothalamus of mice were measured. Mn reduced protein levels of Nrf2 and mRNA expression of Nrf2 target genes and increased mGluR5, COX-2, PGE2, and GnRH levels. Meanwhile, injury-related histomorphology changes in the hypothalamus of mice were significantly present. In conclusion, excessive exposure to Mn disrupts GnRH secretion through Nrf2/mGluR5/COX-2/PGE2 signaling pathway. [ABSTRACT FROM AUTHOR]

Objective To discuss the influence of Shenzhifin (Ginseng, Cassia Twig and Indian Bread) Oral Solution (SZL) on expressions of hippocampal α7-nAChRs and mGluR5 (neuronal synapse signaling related proteins) in APP/PS1 double transgenic mice. Methods Male APP/PS1 double transgenic mice (3 months old) were randomly divided into 5 groups; model group, donepezil group .0.92 mg/(kg · d)], high dose SZL group .50 g/< kg · d)], mid-dose SZL group [25 g/(kg · d)] and low dose SZL group 12.5 g/

Highlights • MPP+ selectively increased the expression of RTN1-C in SN4741 cells. • Downregulation of RTN1-C alleviates MPP+-induced neuronal injury. • Downregulation of RTN1-C reduces mGluR5 expression. • Downregulation of RTN1-C preserves intracellular Ca2+ homeostasis. Abstract Reticulons (RTNs) are a group of membrane-bound proteins that are dominantly localized to the endoplasmic reticulum (ER). RTN1-C, one isoform of RTNs highly expressed in the brain, has been shown to mediate neuronal injury in cerebral ischemia models. The aim of this study was to investigate the role of RTN1-C in an in vitro model of Parkinson's disease (PD) mimicked by 1-methyl-4-phenylpyridinium (MPP+) treatment in SN4741 cells. We found that MPP+ significantly increased the expression of RTN1-C, with no effect on RTN1-A and RTN1-B. Downregulation of RTN1-C using siRNA (Si-RTN1-C) markedly increased cell viability and inhibited apoptosis induced by MPP+ treatment. The results of western blot showed that downregulation of RTN1-C inhibited the surface expression of metabotropic glutamate receptor 5 (mGluR5) but had no effect on mGluR1. The protective effects of Si-RTN1-C were partially prevented by activating mGluR5, not mGluR1. In addition, the results of Ca2+ imaging showed that downregulation of RTN1-C attenuated intracellular Ca2+ release induced by MPP+, which could be nullified by activation of mGluR5 pathway. In conclusion, our data suggest that downregulation of RTN1-C protects SN4741 cells against MPP+ through mGluR5-mediated preservation of Ca2+ homeostasis. Therefore, RTN1-C might represent a therapeutic target for the treatment of neuronal injury in experimental PD models. [ABSTRACT FROM AUTHOR]

Although several studies have found that metabotropic glutamate 5 receptor (mGluR5) may play an important role in autism spectrum disorders (ASD), the mechanisms remain unclear. Here, we used a Shank3 gene complete knockout mouse model (Shank3B−/−) to explore the change in mGluR5 in the brain. To assess whether deletion of Shank3 in mice results in ASD-like behavior, we conducted a battery of behavioral experiments to characterize Shank3B−/− mice, including repetitive grooming behavior tests, three-chamber tests and resident-intruder tests. Wild-type C57/BL6 and Shank3B−/− mice underwent PET scans with [18F]FPEB, which was highly specific to mGluR5. Mouse brains were extracted post-scan, and mGluR5 protein levels were assessed by immunoblotting. The binding potential (BPnd) of mGluR5 was rich in the hippocampus, thalamus, striatum, and amygdala. More importantly, Shank3B−/− mice showed significantly increased BPnd compared to the control mice in these brain regions. Immunoblotting revealed elevated mGluR5 levels in the hippocampus, thalamus, and amygdala but not in the striatum compared with control mice. These findings indicated that [18F]FPEB could visualize mGluR5 in the mouse brain. The deficiency of Shank3 can impair mGluR5 expression in multiple brain regions. Future work is also needed to understand the reasons for different results between in vivo PET and ex vivo immunoblotting. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Morphine tolerance leads to increased mGluR5 levels and phosphorylation of NR2B. • Loss of mGluR5 prevents the development of tolerance and thermal hyperalgesia. • Disruption of NMDA receptor and mGluR5 connection by mutation of shank3 doesn't prevent the development of morphine-induced tolerance. Abstract Background and purpose The aim of this study was to investigate the interactions between mGluR5 and NMDAR during the development of tolerance to the analgesic effect of chronic morphine following disruption of the link between mGluR5 and NMDAR. Methods For these experiments, mGluR5 knockout (KO) mice, Shank3 ΔC mutant mice and their littermate controls were used. Repeated morphine treatment through intrathecal injections (i.t. 10 μg/10 μl twice daily for 5 days) was used to induce antinociceptive tolerance and thermal hyperalgesia, which was determined by tail-flick latency and calculated as a percentage of the maximum possible effect (%MPE). Expression of NMDAR subunits (NR1, NR2A and NR2B) and phosphorylation of NR2B in cortex were evaluated by Western blot. Results Chronic morphine treatment increased the total expression of mGluR5 and NR2A but not NR1 and NR2B in the cortex and enhanced the phosphorylation of NR2B in wild-type mice after tolerance developed. mGluR5 KOs had attenuated morphine-induced tolerance and thermal hyperalgesia and reduced chronic morphine-induced phosphorylation of NR2B but not total NR2A expression. Shank3 ΔC mice were used to imitate the loss of the physiological connection between mGluR5 and NMDAR, and the mice showed a similar analgesic effect to morphine as their littermate controls, and there were no differential effects on NMDAR subunits compared with controls during the development of morphine-induced tolerance. Conclusions It may be mGluR5-mediated PKC signalling that plays the most significant role in the mechanisms underlying morphine-induced antinociceptive tolerance rather than the physiological connection between mGluR5 and NMDAR. [ABSTRACT FROM AUTHOR]

Highlights • Inhibition of mGluR5 under hypoxia promoted cell death. • Inhibition of mGluR5 induced expression of genes of mitochondrial oxidative function. • Akt activation reversed mGluR5 inhibition-induced facilitation of glioma cell death. Abstract Glioma is a primary brain tumor with high frequency and dismal prognosis. As there is no permanent cure available, identifying new therapy or mediator to augment the effectiveness of existing therapy is urgently needed. In the current study we tested the effect of group I metabotropic glutamate receptors (mGluRs): mGluR1 and mGluR5 on the viability of glioma cell lines. We analyzed cell viability using lactate dehydrogenase (LDH) release assay and evaluated apoptosis by propidium iodide (PI) staining. We used qPCR to evaluate change in mitochondrial gene expression and Western blot to evaluate the phosphorylation of Akt and ERK. Inhibition of mGluR5 by a selective antagonist MPEP under hypoxia promoted cell death, and induced expression of mitochondrial oxidative function related genes, with concurrent lowering of AKT phosphorylation level in glioma cell lines. Akt activation reversed mGluR5 inhibition on hypoxia-induced glioma cell death. These results suggest mGluR5 as a potential therapeutic target for hypoxic tumors such as malignant glioma. [ABSTRACT FROM AUTHOR]

Abstract Glutamate, a major neurotransmitter in the central nervous system of human, plays a crucial role in various neurological pathways by activating the ligand-gated ion channels such as mGluR and iGluR. Dysfunction of mGluR 5 can cause Alzheimer's disease, Parkinson's disease, epilepsy, depression, anxiety, etc. In the current study, we have developed the energetically optimized pharmacophore model to screen the eMolecules database having more than 6 million compounds with the help of reported cocrystal structure with 3-chloro-5-[6-(5-fluoropyridin-2 yl)pyrimidin-4-yl]benzonitrile (PDB ID: 5CGD). The obtained hits were docked into the allosteric site of the target and further validation of E-pharmacophore was done by enrichment calculations followed by the molecular dynamics simulations to analyze the specific amino acid interactions with the compound present in the allosteric site of the receptor. [ABSTRACT FROM AUTHOR]

Astrocyte expression of metabotropic glutamate receptor 5 (mGluR5) is consistently observed in resected tissue from patients with epilepsy and is equally prevalent in animal models of epilepsy. However, little is known about the functional signaling properties or downstream consequences of astrocyte mGluR5 activation during epilepsy development. In the rodent brain, astrocyte mGluR5 expression is developmentally regulated and confined in expression/function to the first weeks of life, with similar observations made in human control tissue. Herein, we demonstrate that mGluR5 expression and function dramatically increase in a mouse model of temporal lobe epilepsy. Interestingly, in both male and female mice, mGluR5 function persists in the astrocyte throughout the process of epileptogenesis following status epilepticus. However, mGluR5 expression and function are transient in animals that do not develop epilepsy over an equivalent time period, suggesting that patterns of mGluR5 expression may signify continuing epilepsy development or its resolution. We demonstrate that, during epileptogenesis, astrocytes reacquire mGluR5-dependent calcium transients following agonist application or synaptic glutamate release, a feature of astrocyte-neuron communication absent since early development. Finally, we find that the selective and conditional knock-out of mGluR5 signaling from astrocytes during epilepsy development slows the rate of glutamate clearance through astrocyte glutamate transporters under high-frequency stimulation conditions, a feature that suggests astrocyte mGluR5 expression during epileptogenesis may recapitulate earlier developmental roles in regulating glutamate transporter function. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is the most common cause of dementia and is characterized by the progressive loss of memory and cognition in the aging population. However, the etiology of and therapies for AD remain far from understood. Astrocytes, the most abundant neuroglia in the brain, have recently aroused substantial concern due to their involvement in synaptotoxicity, amyloidosis, neuroinflammation, and oxidative stress. In this review, we summarize the candidate molecules of astrocytes, especially receptors and transporters, that may be involved in AD pathogenesis. These molecules include excitatory amino acid transporters (EAATs), metabotropic glutamate receptor 5 (mGluR5), the adenosine 2A receptor (A2AR), the α7-nicotinic acetylcholine receptor (α7-nAChR), the calcium-sensing receptor (CaSR), S100β, and cannabinoid receptors. We describe the characteristics of these molecules and the neurological and pharmacological underpinnings of these molecules in AD. Among these molecules, EAATs, A2AR, and mGluR5 are strongly related to glutamate-mediated synaptotoxicity and are involved in glutamate transmission or the clearance of extrasynaptic glutamate in the AD brain. The α7-nAChR, CaSR, and mGluR5 are receptors of Aβ and can induce a plethora of toxic effects, such as the production of excess Aβ, synaptotoxicity, and NO production triggered by changes in intracellular calcium signaling. Antagonists or positive allosteric modulators of these receptors can repair cognitive ability and modify neurobiological changes. Moreover, blocking S100β or activating cannabinoid receptors reduces neuroinflammation, oxidative stress, and reactive astrogliosis. Thus, targeting these molecules might provide alternative approaches for treating AD. [ABSTRACT FROM AUTHOR]

Background and Purpose: Dopamine and glutamate reciprocally regulate each other in some of the neurocircuits affected by Parkinson's disease (PD). The objective of this pilot study was to explore relationships between these neurotransmitter systems with positron emission tomography.Methods: The sample consisted of nine patients with PD and eight healthy volunteers (HVs). Dynamic images of the brain were acquired after the IV administration of ∼370 MBq (10 mCi) of [11 C]PE2i, a dopamine transporter (DaT) imaging agent, and ∼185 MBq (∼5 mCi) of [18 F]FPEB, a selective metabotropic glutamate receptor 5 (mGluR5) antagonist. Multiple volumes of interest were semiautomatically placed on contemporaneously acquired MRI scans. Nondisplaceable binding potentials (BPND ) were calculated with the Logan reference tissue model using cerebellar white matter as the reference region.Results: The findings showed that average [18 F]FPEB BPND values were slightly more than 20% higher in PD than HVs in several mesocortical regions, including the bilateral putamen (P = .01), hippocampus (P = .02), and amygdala (P = .05). Average [11 C]PE2i BPND was significantly reduced by about half or more in patients with PD in the bilateral caudate (P < .001) and putamen (P < .001).Conclusions: mGluR5 seems upregulated in strategic dopaminergic brain regions adversely affected by PD. The findings seem to confirm that DaT tracers are better discriminatory biomarkers for diagnosing PD; however, mGluR5 tracers might deserve further exploration as potential biomarkers of response in clinical trials. [ABSTRACT FROM AUTHOR]

The fragile X mental retardation 1 knockout (Fmr1 KO) mouse replicates behavioral deficits associated with autism, fragile X syndrome, and schizophrenia. Less is known whether protein expression changes are consistent with findings in subjects with schizophrenia. In the current study, we used liquid chromatography tandem mass spectrometry (LC‐MS/MS) proteomics to determine the protein expression of four subcellular fractions in the forebrains of Fmr1 KO mice vs. C57BL/6 J mice and the effect of a negative allosteric modulator of mGluR5—2‐Methyl‐6‐(phenylethynyl)pyridine (MPEP)—on protein expression. Strain‐ and treatment‐specific differential expression of proteins was observed, many of which have previously been observed in the brains of subjects with schizophrenia. Western blotting verified the direction and magnitude of change for several proteins in different subcellular fractions as follows: neurofilament light protein (NEFL) and 2′,3′‐cyclic‐nucleotide 3′‐phosphodiesterase (CNP) in the total homogenate; heterogeneous nuclear ribonucleoproteins C1/C2 (HNRNPC) and heterogeneous nuclear ribonucleoprotein D0 (HNRNPD) in the nuclear fraction; excitatory amino acid transporter 2 (EAAT2) and ras‐related protein rab 3a (RAB3A) in the synaptic fraction; and ras‐related protein rab 35 (RAB35) and neuromodulin (GAP43) in the rough endoplasmic reticulum fraction. Individuals with FXS do not display symptoms of schizophrenia. However, the biomarkers that have been identified suggest that the Fmr1 KO model could potentially be useful in the study of schizophrenia. Concordant changes in protein expression in subcellular compartments [total homogenate, synapse, nucleus, and rough endoplasmic reticulum (rER)] via proteomics and western blotting in forebrain neurons of fragile X mental retardation knockout (Fmr1 KO) mice. [ABSTRACT FROM AUTHOR]

Abstract Repeated, recreational ketamine (KE) or methamphetamine (MA) administration seldom produce neurotoxicity, while combining MA and KE administration have been thought to render changes in neural plasticity and motivational behavior. In this study, we sought to assess whether pre-exposure to multiple MA injections and withdrawal may affect low-dose KE-produced rewarding effects, social interaction behavior and its neurochemical underpinnings. A 10-day MA injections (2 mg/kg/day) and 10-day withdrawal regimen was found to cause reliable behavioral sensitization. While KE (1 mg/kg) induced weak conditioned place preference (CPP), pre-exposure to this MA-withdrawal regimen enhanced such KE CPP magnitude. This MA-withdrawal regimen also caused impairments in the social interaction behavior in the sociability, social novelty test. Compared with the mice undergoing the 10-day saline-withdrawal or MA regimen, mice receiving the 10-day MA-withdrawal regimen exhibited lower dopamine-releasing probability in the nucleus accumbens, inferring the MA-withdrawal regimen-primed preference for KE rewarding effects. Likewise, mice receiving the MA-withdrawal regimen had high expression in mGluR5 protein but unaltered EAAT3, Homer2 expression in hippocampal tissues. Pretreatment with MPEP, an mGluR5 antagonist, prevented the MA-withdrawal regimen-induced increment in the KE CPP magnitude and impairments in social interaction behavior. We, thus, conclude that repeated MA administration and abstinence may enhance KE rewarding effects and produce eminent deficits in social recognition and interest. And these effects correlate with the mGluR5 over-expression and modulation of the KE-stimulating effect on dopamine release. Highlights • Ketamine (KE, 1 mg/kg) induces weak conditioned place preference (CPP). • A methamphetamine (MA)-withdrawal (WD) regimen enhances KE (1 mg/kg) CPP magnitude. • The same regimen deteriorates mouse sociability and social novelty preference. • The MA-WD regimen prime for the plasticity in ketamine-stimulating dopamine release • MPEP Pretreatment prevents the regimen-induced changes in KE CPP and social behavior. [ABSTRACT FROM AUTHOR]

Gonadal hormones play a vital role in driving motivated behavior. They not only modulate responses to naturally rewarding stimuli, but also influence responses to drugs of abuse. A commonality between gonadal hormones and drugs of abuse is that they both impact the neurocircuitry of reward, including the regulation of structural plasticity in the nucleus accumbens (NAc). Previous hormonal studies have focused on the mechanisms and behavioral correlates of estradiol-induced dendritic spine changes in the female NAc. Here we sought to determine the effects of androgens on medium spiny neuron (MSN) spine plasticity in the male NAc. Following treatment with the androgen receptor agonist dihydrotestosterone (DHT), MSNs in castrated male rats exhibited a significant decrease in dendritic spine density. This effect was isolated to the shell subregion of the NAc. The effect of DHT was dependent on mGluR5 activity, and local mGluR5 activation and subsequent endocannabinoid signaling produce an analogous NAc shell spine decrease. Somewhat surprisingly, DHT-induced conditioned place preference remained intact following systemic inhibition of mGluR5. These findings indicate that androgens can utilize mGluR signaling, similar to estrogens, to mediate changes in NAc dendritic structure. In addition, there are notable differences in the direction of spine changes, and site specificity of estrogen and androgen action, suggesting sex differences in the hormonal regulation of motivated behaviors. [ABSTRACT FROM AUTHOR]

Abstract Metabotropic glutamate receptor 5 (mGluR 5) is a drug target for central nervous system disorders such as fragile X syndrome that involve excessive glutamate-induced excitation. We tested the efficacy of a novel negative allosteric modulator of mGluR 5 developed by Merz Pharmaceuticals, MRZ-8456, in comparison to MPEP and AFQ-056 (Novartis, a.k.a. mavoglurant) in both in vivo and in vitro assays in a mouse model of fragile X syndrome, Fmr1 KO mice. The in vivo assays included susceptibility to audiogenic-induced seizures and pharmacokinetic measurements of drug availability. The in vitro assays included dose response assessments of biomarker expression and dendritic spine length and density in cultured primary neurons. Both MRZ-8456 and AFQ-056 attenuated wild running and audiogenic-induced seizures in Fmr1 KO mice with similar pharmacokinetic profiles. Both drugs significantly reduced dendritic expression of amyloid-beta protein precursor (APP) and rescued the ratio of mature to immature dendritic spines. These findings demonstrate that MRZ-8456, a drug being developed for the treatment of motor complications of L-DOPA in Parkinson’s disease and which completed a phase I clinical trial, is effective in attenuating both well-established (seizures and dendritic spine maturity) and exploratory biomarker (APP expression) phenotypes in a mouse model of fragile X syndrome. Highlights • Metabotropic glutamate receptor 5 antagonist MRZ-8456 attenuates seizures. • MRZ-8456 reduces dendritic expression of amyloid-beta protein precursor. • MRZ-8456 rescues the ratio of mature to immature dendritic spines in Fmr1 KO neurons. [ABSTRACT FROM AUTHOR]

It is well-appreciated that phosphorylation is an essential post-translational mechanism of regulation for several proteins, including group 1 metabotropic glutamate receptors (mGluRI), mGluR1, and mGluR5 subtypes. While contributions of various serine/threonine protein kinases on mGluRI modulation have been recognized, the functional role of tyrosine kinases (TKs) is less acknowledged. Here, while describing current evidence supporting that mGluRI are targets of TKs, we mainly focus on the modulatory roles of the ErbB tyrosine kinases receptors—activated by the neurotrophic factors neuregulins (NRGs)—on mGluRI function. Available evidence suggests that mGluRI activity is tightly dependent on ErbB signaling, and that ErbB’s modulation profoundly influences mGluRI-dependent effects on neurotransmission, neuronal excitability, synaptic plasticity, and learning and memory processes.

[18F]FPEB is a positron emission tomography (PET) radiopharmaceutical used for imaging the abundance and distribution of mGluR5 in the central nervous system (CNS). Efficient radiolabeling of the aromatic ring of [18F]FPEB has been an ongoing challenge. Herein, five metal-free precursors for the radiofluorination of [18F]FPEB were compared, namely, a chloro-, nitro-, sulfonium salt, and two spirocyclic iodonium ylide (SCIDY) precursors bearing a cyclopentyl (SPI5) and a new adamantyl (SPIAd) auxiliary. The chloro- and nitro-precursors resulted in a low radiochemical yield (18F]FPEB in the highest RCYs of 25% and 36%, respectively. Preliminary PET/CT imaging studies with [18F]FPEB were conducted in a transgenic model of Alzheimer’s Disease (AD) using B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J (APP/PS1) mice, and data were compared with age-matched wild-type (WT) B6C3F1/J control mice. In APP/PS1 mice, whole brain distribution at 5 min post-injection showed a slightly higher uptake (SUV = 4.8 ± 0.4) than in age-matched controls (SUV = 4.0 ± 0.2). Further studies to explore mGluR5 as an early biomarker for AD are underway.

Glutamate plays a crucial role in the treatment of depression by interacting with the metabotropic glutamate receptor subtype 5 (mGluR5), whose negative allosteric modulators (NAMs) are thus promising antidepressants. At present, to explore the structural features of 106 newly synthesized aryl benzamide series molecules as mGluR5 NAMs, a set of ligand-based three-dimensional quantitative structure-activity relationship (3D-QSAR) analyses were firstly carried out applying comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) methods. In addition, receptor-based analysis, namely molecular docking and molecular dynamics (MD) simulations, were performed to further elucidate the binding modes of mGluR5 NAMs. As a result, the optimal CoMSIA model obtained shows that cross-validated correlation coefficient Q2 = 0.70, non-cross-validated correlation coefficient R2ncv = 0.89, predicted correlation coefficient R2pre = 0.87. Moreover, we found that aryl benzamide series molecules bind as mGluR5 NAMs at Site 1, which consists of amino acids Pro655, Tyr659, Ile625, Ile651, Ile944, Ser658, Ser654, Ser969, Ser965, Ala970, Ala973, Trp945, Phe948, Pro903, Asn907, Val966, Leu904, and Met962. This site is the same as that of other types of NAMs; mGluR5 NAMs are stabilized in the “linear” and “arc” configurations mainly through the H-bonds interactions, π−π stacking interaction with Trp945, and hydrophobic contacts. We hope that the models and information obtained will help understand the interaction mechanism of NAMs and design and optimize NAMs as new types of antidepressants.

Genetic defects in SHANK genes are associated with autism. Deletions and truncating mutations suggest haploinsufficiency for Shank3 as a major cause of disease which may be analyzed in appropriate Shank deficient mouse models. Here we will focus on the functional analysis of missense mutations found in SHANK genes. The relevance of most of these mutations for Shank function, and their role in autism pathogenesis is unclear. This is partly due to the fact that mutations spare the most well studied functional domains of Shank3, such as the PDZ and SAM domains, or the short proline-rich motifs which are required for interactions with postsynaptic partners Homer, Cortactin, dynamin, IRSp53 and Abi-1. One set of mutations affects the N-terminal part, including the highly conserved SPN domain and ankyrin repeats. Functional analysis from several groups has indicated that these mutations (e.g., R12C; L68P; R300C, and Q321R) interfere with the critical role of Shank3 for synapse formation. More recently the structural analysis of the SPN-ARR module has begun to shed light on the molecular consequences of mutations in the SPN of Shank3. The SPN was identified as a Ras association domain, with high affinities for GTP-bound, active forms of Ras and Rap. The two autism related mutations in this part of the protein, R12C and L68P, both abolish Ras binding. Further work is directed at identifying the consequences of Ras binding to Shank proteins at postsynaptic sites.

Impairment of the metabotropic glutamate receptor 5 (mGluR5) has been implicated with various neurologic disorders. Although mGluR5 density can be quantified with the PET radiotracer [11C]ABP688, the methods for reproducible quantification of [11C]ABP688 PET imaging in mice have not been thoroughly investigated yet. Thus, this study aimed to assess and validate cerebellum as reference region for simplified reference tissue model (SRTM), investigate the feasibility of a noninvasive cardiac image-derived input function (IDIF) for relative quantification, to validate the use of a PET template instead of an MRI template for spatial normalization, and to determine the reproducibility and within-subject variability of [11C]ABP688 PET imaging in mice. Blocking with the mGluR5 antagonist MPEP resulted in a reduction of [11C]ABP688 binding of 41% in striatum (p < 0.0001), while no significant effect could be found in cerebellum (−4.8%, p > 0.99) indicating cerebellum as suitable reference region for mice. DVR-1 calculated using a noninvasive IDIF and an arteriovenous input function correlated significantly when considering the cerebellum as the reference region (striatum: DVR-1, r = 0.978, p < 0.0001). Additionally, strong correlations between binding potential calculated from SRTM (BPND) with DVR-1 based on IDIF (striatum: r = 0.980, p < 0.0001) and AV shunt (striatum: r = 0.987, p < 0.0001). BPND displayed higher discrimination power than VT values in determining differences between wild-types and heterozygous Q175 mice, an animal model of Huntington's disease. Furthermore, we showed high agreement between PET- and MRI-based spatial normalization approaches (striatum: r = 0.989, p < 0.0001). Finally, both spatial normalization approaches did not reveal any significant bias between test-retest scans, with a relative difference below 5%. This study indicates that noninvasive quantification of [11C]ABP688 PET imaging is reproducible and cerebellum can be used as reference region in mice.

The present study was aimed to further characterize the pharmacological profile of N-[4-(trifluoromethyl) benzyl]-4-methoxybutyramide (GET73), a putative negative allosteric modulator (NAM) of metabotropic glutamate subtype 5 receptor (mGluR5) under development as a novel medication for the treatment of alcohol dependence. This aim has been accomplished by means of a series of in vitro functional assays. These assays include the measure of several down-stream signaling [intracellular Ca++ levels, inositol phosphate (IP) formation and CREB phosphorylation (pCREB)] which are generally affected by mGluR5 ligands. In particular, GET73 (0.1 nM–10 μM) was explored for its ability to displace the concentration-response curve of some mGluR5 agonists/probes (glutamate, L-quisqualate, CHPG) in different native preparations. GET73 produced a rightward shift of concentration-response curves of glutamate- and CHPG-induced intracellular Ca++ levels in primary cultures of rat cortical astrocytes. The compound also induced a rightward shift of concentration response curve of glutamate- and L-quisqualate-induced increase in IP turnover in rat hippocampus slices, along with a reduction of CHPG (10 mM)-induced increase in IP formation. Moreover, GET73 produced a rightward shift of concentration-response curve of glutamate-, CHPG- and L-quisqualate-induced pCREB levels in rat cerebral cortex neurons. Although the engagement of other targets cannot be definitively ruled out, these data support the view that GET73 acts as an mGluR5 NAM and support the significance of further investigating the possible mechanism of action of the compound.

We studied group-I metabotropic glutamate (mGlu) receptors in Pahenu2 (ENU2) mice, which mimic the genetics and neurobiology of human phenylketonuria (PKU), a metabolic disorder characterized, if untreated, by autism, and intellectual disability (ID). Male ENU2 mice showed increased mGlu5 receptor protein levels in the hippocampus and corpus striatum (but not in the prefrontal cortex) whereas the transcript of the mGlu5 receptor was unchanged. No changes in mGlu1 receptor mRNA and protein levels were found in any of the three brain regions of ENU2 mice. We extended the analysis to Homer proteins, which act as scaffolds by linking mGlu1 and mGlu5 receptors to effector proteins. Expression of the long isoforms of Homer was significantly reduced in the hippocampus of ENU2 mice, whereas levels of the short Homer isoform (Homer 1a) were unchanged. mGlu5 receptors were less associated to immunoprecipitated Homer in the hippocampus of ENU2 mice. The lack of mGlu5 receptor-mediated long-term depression (LTD) in wild-type mice (of BTBR strain) precluded the analysis of hippocampal synaptic plasticity in ENU2 mice. We therefore performed a behavioral analysis to examine whether pharmacological blockade of mGlu5 receptors could correct behavioral abnormalities in ENU2 mice. Using the same apparatus we sequentially assessed locomotor activity, object exploration, and spatial object recognition (spatial novelty test) after displacing some of the objects from their original position in the arena. Systemic treatment with the mGlu5 receptor antagonist, MPEP (20 mg/kg, i.p.), had a striking effect in the spatial novelty test by substantially increasing the time spent in exploring the displaced objects in ENU2 mice (but not in wild-type mice). These suggest a role for mGlu5 receptors in the pathophysiology of ID in PKU and suggest that, also in adult untreated animals, cognitive dysfunction may be improved by targeting these receptors with an appropriate therapy.

Fragile X syndrome (FXS) is the most common monogenetic cause of intellectual disability and autism. The disorder is characterized by altered synaptic plasticity in the brain. Synaptic plasticity is tightly regulated by a complex balance of different synaptic pathways. In FXS, various synaptic pathways are disrupted, including the excitatory metabotropic glutamate receptor 5 (mGluR5) and the inhibitory γ-aminobutyric acid (GABA) pathways. Targeting each of these pathways individually, has demonstrated beneficial effects in animal models, but not in patients with FXS. This lack of translation might be due to oversimplification of the disease mechanisms when targeting only one affected pathway, in spite of the complexity of the many pathways implicated in FXS. In this report we outline the hypothesis that targeting more than one pathway simultaneously, a combination therapy, might improve treatment effects in FXS. In addition, we present a glance of the first results of chronic combination therapy on social behavior in Fmr1 KO mice. In contrast to what we expected, targeting both the mGluR5 and the GABAergic pathways simultaneously did not result in a synergistic effect, but in a slight worsening of the social behavior phenotype. This does implicate that both pathways are interconnected and important for social behavior. Our results underline the tremendous fine-tuning that is needed to reach the excitatory-inhibitory balance in the synapse in relation to social behavior. We believe that alternative strategies focused on combination therapy should be further explored, including targeting pathways in different cellular compartments or cell-types.

To date, histone H2B monoubiquitination (H2Bub), a mark associated with transcriptional elongation and ongoing transcription, has not been linked to the development or maintenance of neuropathic pain states. Here, using male Sprague Dawley rats, we demonstrated spinal nerve ligation (SNL) induced behavioral allodynia and provoked ring finger protein 20 (RNF20)-dependent H2Bub in dorsal horn. Moreover, SNL provoked RNF20-mediated H2Bub phosphorylated RNA polymerase II (RNAPII) in the promoter fragments of mGluR5, thereby enhancing mGluR5 transcription/expression in the dorsal horn. Conversely, focal knockdown of spinal RNF20 expression reversed not only SNL-induced allodynia but also RNF20/H2Bub/RNAPII phosphorylation-associated spinal mGluRS transcription/expression. Notably, TNF-a injection into naive rats and specific neutralizing antibody injection into SNL-induced allodynia rats revealed that TNF-a-associated allodynia involves the RNF20/H2Bub/RNAPII transcriptional axis to upregulate mGluR5 expression in the dorsal horn. Collectively, our findings indicated TNF-a induces RNF20-drived H2B monoubiquitination, which facilitates phosphorylated RNAPII-dependent mGluR5 transcription in the dorsal horn for the development of neuropathic allodynia. [ABSTRACT FROM AUTHOR]

Genetic defects in SHANK genes are associated with autism. Deletions and truncating mutations suggest haploinsufficiency for Shank3 as a major cause of disease which may be analyzed in appropriate Shank deficient mouse models. Here we will focus on the functional analysis of missense mutations found in SHANK genes. The relevance of most of these mutations for Shank function, and their role in autism pathogenesis is unclear. This is partly due to the fact that mutations spare the most well studied functional domains of Shank3, such as the PDZ and SAM domains, or the short proline-rich motifs which are required for interactions with postsynaptic partners Homer, Cortactin, dynamin, IRSp53 and Abi-1. One set of mutations affects the N-terminal part, including the highly conserved SPN domain and ankyrin repeats. Functional analysis from several groups has indicated that these mutations (e.g., R12C; L68P; R300C, and Q321R) interfere with the critical role of Shank3 for synapse formation. More recently the structural analysis of the SPN-ARR module has begun to shed light on the molecular consequences of mutations in the SPN of Shank3. The SPN was identified as a Ras association domain, with high affinities for GTP-bound, active forms of Ras and Rap. The two autism related mutations in this part of the protein, R12C and L68P, both abolish Ras binding. Further work is directed at identifying the consequences of Ras binding to Shank proteins at postsynaptic sites. [ABSTRACT FROM AUTHOR]

The metabotropic glutamatergic receptor subtype 5 (mGluR5) may represent a promising therapeutic target for stress-related psychiatric disorders. Here, we describe mGluR5 findings in stress disorders, particularly major depressive disorder (MDD), highlighting insights from positron emission tomography studies. Positron emission tomography studies report either no differences or lower mGluR5 in MDD, potentially reflecting MDD heterogeneity. Unlike the rapidly acting glutamatergic agent ketamine, mGluR5-specific modulation has not yet shown antidepressant efficacy in MDD and bipolar disorder. Although we recently showed that ketamine may work, in part, through significant mGluR5 modulation, the specific role of mGluR5 downregulation in ketamine’s antidepressant response is unclear. In contrast to MDD, there has been much less investigation of mGluR5 in bipolar disorder, yet initial studies indicate that mGluR5-specific treatments may aid in both depressed and manic mood states. The direction of modulation needed may be state dependent, however, limiting clinical feasibility. There has been relatively little study of posttraumatic stress disorder or obsessive-compulsive disorder to date, although there is evidence for the upregulation of mGluR5 in these disorders. However, while antagonism of mGluR5 may reduce fear conditioning, it may also reduce fear extinction. Therefore, studies are needed to determine the role mGluR5 modulation might play in the treatment of these conditions. Further challenges in modulating this prevalent neurotransmitter system include potential induction of significant side effects. As such, more research is needed to identify level and type (positive/negative allosteric modulation or full antagonism) of mGluR5 modulation required to translate existing knowledge into improved therapies. [ABSTRACT FROM AUTHOR]

Long-term levodopa (l-dopa) treatment in patients with Parkinson´s disease (PD) is associated with the development of motor complications (ie, motor fluctuations and dyskinesias). The principal etiopathogenic factors are the degree of nigro-striatal dopaminergic loss and the duration and dose of l-dopa treatment. In this review article we concentrate on analysis of the mechanisms underlying l-dopa-induced dyskinesias, a phenomenon that causes disability in a proportion of patients and that has not benefited from major therapeutic advances. Thus, we discuss the main neurotransmitters, receptors, and pathways that have been thought to play a role in l-dopa-induced dyskinesias from the perspective of basic neuroscience studies. Some important advances in deciphering the molecular pathways involved in these abnormal movements have occurred in recent years to reveal potential targets that could be used for therapeutic purposes. However, it has not been an easy road because there have been a plethora of components involved in the generation of these undesired movements, even bypassing the traditional and well-accepted dopamine receptor activation, as recently revealed by optogenetics. Here, we attempt to unify the available data with the hope of guiding and fostering future research in the field of striatal activation and abnormal movement generation. © 2017 International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Impairment of the metabotropic glutamate receptor 5 (mGluR5) has been implicated with various neurologic disorders. Although mGluR5 density can be quantified with the PET radiotracer [11C]ABP688, the methods for reproducible quantification of [11C]ABP688 PET imaging in mice have not been thoroughly investigated yet. Thus, this study aimed to assess and validate cerebellum as reference region for simplified reference tissue model (SRTM), investigate the feasibility of a noninvasive cardiac image-derived input function (IDIF) for relative quantification, to validate the use of a PET template instead of an MRI template for spatial normalization, and to determine the reproducibility and within-subject variability of [11C]ABP688 PET imaging in mice. Blocking with the mGluR5 antagonist MPEP resulted in a reduction of [11C]ABP688 binding of 41% in striatum (p < 0.0001), while no significant effect could be found in cerebellum (-4.8%, p > 0.99) indicating cerebellum as suitable reference region for mice. DVR-1 calculated using a noninvasive IDIF and an arteriovenous input function correlated significantly when considering the cerebellum as the reference region (striatum: DVR-1, r = 0.978, p < 0.0001). Additionally, strong correlations between binding potential calculated from SRTM (BPND) with DVR-1 based on IDIF (striatum: r = 0.980, p < 0.0001) and AV shunt (striatum: r = 0.987, p < 0.0001). BPND displayed higher discrimination power than VT values in determining differences between wild-types and heterozygous Q175 mice, an animal model of Huntington's disease. Furthermore, we showed high agreement between PET- and MRI-based spatial normalization approaches (striatum: r = 0.989, p < 0.0001). Finally, both spatial normalization approaches did not reveal any significant bias between test-retest scans, with a relative difference below 5%. This study indicates that noninvasive quantification of [11C]ABP688 PET imaging is reproducible and cerebellum can be used as reference region in mice. [ABSTRACT FROM AUTHOR]

Atom-based three dimensional-quantitative structure-activity relationship (3D-QSAR) model was developed on the basis of 5-point pharmacophore hypothesis (AARRR) with two hydrogen bond acceptors (A) and three aromatic rings for the derivatives of thieno[2,3-b]pyridine, which modulates the activity to inhibit the mGluR5 receptor. Generation of a highly predictive 3D-QSAR model was performed using the alignment of predicted pharmacophore hypothesis for the training set (R2 = 0.84, SD = 0.26, F = 45.8, N = 29) and test set (Q2 = 0.74, RMSE = 0.235, Pearson-R = 0.94, N = 9). The best pharmacophore hypothesis AARRR was selected, and developed three dimensional-quantitative structure activity relationship (3D-QSAR) model also supported the outcome of this study by means of favorable and unfavorable electron withdrawing group and hydrophobic regions of most active compound 42d and least active compound 18b. Following, induced fit docking and binding free energy calculations reveals the reliable binding orientation of the compounds. Finally, molecular dynamics simulations for 100 ns were performed to depict the protein-ligand stability. We anticipate that the resulted outcome could be supportive to discover potent negative allosteric modulators for metabotropic glutamate receptor 5 (mGluR5). [ABSTRACT FROM AUTHOR]

The Arc gene is robustly transcribed in specific neural ensembles in response to experience-driven activity. Upon induction, Arc mRNA is transported to dendrites, where it can be rapidly and locally translated by activation of metabotropic glutamate receptors (mGluR1/5). mGluR-induced dendritic synthesis of Arc is implicated in weakening or elimination of excitatory synapses by triggering endocytosis of postsynaptic AMPARs in both hippocampal CA1 and cerebellar Purkinje neurons. Importantly, CA1 neurons with experience-induced Arc mRNA are susceptible, or primed for mGluR-induced long-term synaptic depression (mGluR-LTD). Here we review mechanisms and function of Arc in mGluR-LTD and synapse elimination and propose roles for these forms of plasticity in Arc-dependent formation of sparse neural representations of learned experience. We also discuss accumulating evidence linking dysregulation of Arc and mGluR-LTD in human cognitive disorders such as intellectual disability, autism and Alzheimer’s disease. [ABSTRACT FROM AUTHOR]

The aim of the present study was to evaluate functional changes of mGluR5 expression in advanced Alzheimer's disease (AD) using positron emission tomography (PET) with an mGluR5 specific radiotracer ([ 18 F]FPEB) in 5xFAD AD model. Subsequently, in the same animal, mGluR5 expression was quantified by immunoassay techniques. The non-displaceable binding potential values for mGluR5 was estimated by the Logan's graphical analysis. Brain PET imaging revealed that radioactivities in the hippocampus and the striatum were significantly lower in 5xFAD mice compared to control animals. Binding values were also significantly lowered in 5xFAD mice. This decline was validated by immunoblotting of protein isolates from brain tissues, as the mean band density for 5xFAD mice had a lower mGluR5 intensity than for wild type mice. These results indicated that mGluR5 levels in 5xFAD mice were down regulated in the limbic system. [ABSTRACT FROM AUTHOR]

Abstract: Dysregulation of prefrontal cortical glutamatergic signalling via NMDA receptor hypofunction has been implicated in cognitive dysfunction and impaired inhibitory control in such neuropsychiatric disorders as schizophrenia, attention‐deficit hyperactivity disorder and drug addiction. Although NMDA receptors functionally interact with metabotropic glutamate receptor 5 (mGluR5), the consequence of this interaction for glutamate release in the prefrontal cortex (PFC) remains unknown. We therefore investigated the effects of positive and negative allosteric mGluR5 modulation on changes in extracellular glutamate efflux in the medial PFC (mPFC) induced by systemic administration of the non‐competitive NMDA receptor antagonist dizocilpine (or MK801) in rats. Extracellular glutamate efflux was measured following systemic administration of the positive allosteric mGluR5 modulator [S‐(4‐Fluoro‐phenyl)‐{3‐[3‐(4‐fluoro‐phenyl)‐[1,2,4]‐oxadiazol‐5‐yl]‐piperidin‐1‐yl}‐methanone] (ADX47273; 100 mg/kg, p.o.) and negative allosteric mGluR5 modulator [2‐chloro‐4‐{[1‐(4‐fluorophenyl)‐2,5‐dimethyl‐1H‐imidazol‐4‐yl]ethynyl}pyridine] (RO4917523; 0.3 mg/kg, p.o.), using a wireless glutamate biosensor in awake, freely moving rats. The effect of MK801 (0.03–0.06 mg/kg, s.c.) on mPFC glutamate efflux was also investigated in addition to the effects of MK801 (0.03 mg/kg, s.c.) following ADX47273 (100 mg/kg, p.o.) pre‐treatment. ADX47273 produced a sustained increase in glutamate efflux and increased the effect of NMDA receptor antagonism on glutamate efflux in the mPFC. In contrast, negative allosteric mGluR5 modulation with RO4917523 decreased glutamate efflux in the mPFC. These findings indicate that positive and negative allosteric mGluR5 modulators produce long lasting and opposing actions on extracellular glutamate efflux in the mPFC. Positive and negative allosteric modulators of mGluR5 may therefore be viable therapeutic agents to correct abnormalities in glutamatergic signalling present in a range of neuropsychiatric disorders. [ABSTRACT FROM AUTHOR]

The present study was aimed to further characterize the pharmacological profile of N-[4-(trifluoromethyl) benzyl]-4-methoxybutyramide (GET73), a putative negative allosteric modulator (NAM) of metabotropic glutamate subtype 5 receptor (mGluR5) under development as a novel medication for the treatment of alcohol dependence. This aim has been accomplished by means of a series of in vitro functional assays. These assays include the measure of several down-stream signaling [intracellular Ca++ levels, inositol phosphate (IP) formation and CREB phosphorylation (pCREB)] which are generally affected by mGluR5 ligands. In particular, GET73 (0.1 nM-10 mM) was explored for its ability to displace the concentration-response curve of some mGluR5 agonists/probes (glutamate, L-quisqualate, CHPG) in different native preparations. GET73 produced a rightward shift of concentration-response curves of glutamate- and CHPG-induced intracellular Ca++ levels in primary cultures of rat cortical astrocytes. The compound also induced a rightward shift of concentration response curve of glutamate- and L-quisqualate-induced increase in IP turnover in rat hippocampus slices, along with a reduction of CHPG (10 mM)-induced increase in IP formation. Moreover, GET73 produced a rightward shift of concentration-response curve of glutamate-, CHPGand L-quisqualate-induced pCREB levels in rat cerebral cortex neurons. Although the engagement of other targets cannot be definitively ruled out, these data support the view that GET73 acts as an mGluR5 NAM and support the significance of further investigating the possible mechanism of action of the compound. [ABSTRACT FROM AUTHOR]

Cell–cell communication plays a central role in the guidance of migrating neuronal precursor cells during the development of the cerebral cortex. Endocannabinoids (eCBs) have previously been shown to be one of the central factors regulating neuronal migration. In this study the effects of eCBs on different parameters, expected to affect embryonic cortical neuronal motility have been analyzed in neurosphere-derived neuroblasts using time-lapse microscopy. Increased endogenous production of the endocannabinoid 2-arachidonyl glycerol (2-AG) causes bursts of neuroblast motility. The neuroblasts move longer distances and show a low frequency of turning, and the number of neuron–neuron contacts are reduced. Similar changes occur interfering with the function of the metabotropic glutamate receptor 5 (mGluR5) or its transducer canonical transient receptor potential channel 3 (TRPC3) or the neuregulin receptor ErbB4. Blocking of 2-AG production reverses these effects. The data suggest that eCB-regulated neuronal motility is controlled by mGluR5/TRPC3 activity possibly via NRG/ErbB4 signaling. [ABSTRACT FROM AUTHOR]

GRM5 (coding for metabotropic glutamate receptor 5, mGluR5) is a promising target for the treatment of cognitive deficits in schizophrenia, but there has been little investigation of its association with cognitive and brain phenotypes within this disorder. We examined the effects of common genetic variation in GRM5 with cognitive function, hippocampal volume, and hippocampal mGluR5 protein levels in schizophrenia patients relative to healthy controls. Two independent GRM5 variants rs60954128 [C>T] and rs3824927 [G>T] were genotyped in a schizophrenia case/control cohort (n=249/261). High-resolution anatomical brain scans were available for a subset of the cohort (n=103 schizophrenia /78 control). All participants completed a standard set of neuropsychological tests. In a separate postmortem cohort (n=19 schizophrenia/20 controls), hippocampal mGluR5 protein levels were examined among individuals of different GRM5 genotypes. Schizophrenia minor allele carriers of rs60954128 had reduced right hippocampal volume relative to healthy controls of the same genotype (-12.3%); this effect was exaggerated in males with schizophrenia (-15.6%). For rs3824927, compared to major allele homozygotes, minor allele carriers with schizophrenia had lower Intelligence Quotients (IQ). Examination in hippocampal postmortem tissue showed no difference in mGluR5 protein expression according to genotype for either rs60954128 or rs3824927. While these genetic variants in GRM5 were associated with cognitive impairments and right hippocampal volume reduction in schizophrenia, they did not affect protein expression. Further study of these mechanisms may help to delineate new targets for the treatment of cognitive deficits in schizophrenia, and may be relevant to other disorders. [ABSTRACT FROM AUTHOR]

2-Methyl-6-(phenylethynyl)pyridine (MPEP), a negative allosteric modulator of the metabotropic glutamate receptor (mGluR) 5, protects hepatocytes from ischemic injury. In astrocytes and microglia, MPEP depletes ATP. These findings seem to be self-contradictory, since ATP depletion is a fundamental stressor in ischemia. This study attempted to reconstruct the mechanism of MPEP-mediated ATP depletion and the consequences of ATP depletion on protection against ischemic injury. We compared the effects of MPEP and other mGluR5 negative modulators on ATP concentration when measured in rat hepatocytes and acellular solutions. We also evaluated the effects of mGluR5 blockade on viability in rat hepatocytes exposed to hypoxia. Furthermore, we studied the effects of MPEP treatment on mouse livers subjected to cold ischemia and warm ischemia reperfusion. We found that MPEP and 3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pyridine (MTEP) deplete ATP in hepatocytes and acellular solutions, unlike fenobam. This finding suggests that mGluR5s may not be involved, contrary to previous reports. MPEP, as well as MTEP and fenobam, improved hypoxic hepatocyte viability, suggesting that protection against ischemic injury is independent of ATP depletion. Significantly, MPEP protected mouse livers in two different ex vivo models of ischemia reperfusion injury, suggesting its possible protective deployment in the treatment of hepatic inflammatory conditions. [ABSTRACT FROM AUTHOR]

The D2 receptor partial agonist, aripiprazole, has shown increased therapeutic efficacy for schizophrenia, autism and Tourette's syndrome compared to traditional antipsychotics such as the D2 receptor antagonist, haloperidol. Recent evidence suggests this superior profile may be associated with downstream effects on glutamatergic synapses. Group 1 metabotropic glutamate receptors (mGluRs) and their endogenous modulators, Norbin and Homer1, are regulated by D2 receptor activity, particularly within the nucleus accumbens (NAc), a target region of aripiprazole and haloperidol. This study sought to evaluate the effects of aripiprazole on Group 1 mGluRs, Norbin and Homer1 in the NAc, in comparison to haloperidol. Sprague-Dawley rats were orally administered daily doses of aripiprazole (2.25 mg/kg), haloperidol (0.3 mg/kg) or vehicle for 1 or 10-weeks. Immunoblot analyses revealed Group 1 mGluR protein levels were not altered following 1-week and 10-week aripiprazole or haloperidol treatment, compared to vehicle treated rodents. However, 1-week aripiprazole and haloperidol treatment significantly elevated Homer1a and Norbin protein expression, respectively. After 10 weeks of treatment, aripiprazole, but not haloperidol, significantly increased Norbin expression. These findings indicate the antipsychotics, aripiprazole and haloperidol, exert differential temporal effects on Norbin and Homer1 expression that may have consequences on synaptic glutamatergic transmission underlying their therapeutic profile. [ABSTRACT FROM AUTHOR]