Glu Receptors Research Articles

Balancing Neuroprotection with Functional Recovery: The Potential Role of the PNN in Preventing Excitotoxicity after SCI

In spinal cord injury (SCI), initial mechanical trauma causes debilitating primary damage to neural cells and blood vessels. Following this, secondary cascades of downstream events occur, including inflammation, ischemia, and excitotoxicity — an increase in intracellular Ca2+ concentration from overactive glutamate (Glu) receptors leading to cell death. Additionally, there is an upregulation of the perineuronal net (PNN), a lattice‐like structure of the extracellular matrix (ECM) which modulates neural communication and homeostasis. The PNN is partially composed of negatively charged chondroitin sulfate proteoglycans (CSPGs). While CSPGs can stabilize plasticity and neuronal growth during development, these molecules become inhibitory to regeneration, sprouting and plasticity after injury, as well as contribute greatly to the glial scar. However, administration of the bacterial enzyme chondroitinase ABC (ChABC) can digest the PNN and CSPGs, ultimately promoting functional recovery. What remains unknown are the other impacts of removing the PNN at very acute stages of injury. We hypothesize that the PNN and its negatively charged CSPGs are upregulated after SCI as a neuroprotective response that attenuates excitotoxicity by acting as a buffer for Ca2+. To test our hypothesis, we induced excitotoxicity by injecting rats with a threshold dose of NMDA with or without ChABC utilizing the well defined spinal respiratory motor system. 59% of SCI occurs at the cervical level, and mechanical ventilation is the leading cause of death and restriction of independence in these cases. Therefore, we administered the dose instraspinally at the C4 level and paired treatment with intrapleural injection of cholera toxin‐B to retrogradely label the phrenic motor neuron pool, which innervates the diaphragm. Preliminary findings suggest that animals lacking the PNN have exacerbated cell death versus control. This implies that, following SCI, the body's main focus is to survive and not necessarily to preserve function. PNN upregulation after injury could be a conserved mechanism which promotes survival and CNS tissue preservation at the expense of plasticity and functional regeneration. The next steps are to detail the spread of ChABC and how long it takes for the PNN to regenerate. Future directions will investigate the optimal timing of ChABC administration to balance preserving cell survival with promoting functional recovery.Support or Funding InformationNINDS R01NS101105 (WJA)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Attenuation of Novelty-Induced Hyperactivity of Gria1-/- Mice by Cannabidiol and Hippocampal Inhibitory Chemogenetics.

Gene-targeted mice with deficient AMPA receptor GluA1 subunits (Gria1-/- mice) show robust hyperlocomotion in a novel environment, suggesting them to constitute a model for hyperactivity disorders such as mania, schizophrenia and attention deficit hyperactivity disorder. This behavioral alteration has been associated with increased neuronal activation in the hippocampus, and it can be attenuated by chronic treatment with antimanic drugs, such as lithium, valproic acid, and lamotrigine. Now we found that systemic cannabidiol strongly blunted the hyperactivity and the hippocampal c-Fos expression of the Gria1-/- mice, while not affecting the wild-type littermate controls. Acute bilateral intra-dorsal hippocampal infusion of cannabidiol partially blocked the hyperactivity of the Gria1-/- mice, but had no effect on wild-types. The activation of the inhibitory DREADD receptor hM4Gi in the dorsal hippocampus by clozapine-N-oxide robustly inhibited the hyperactivity of the Gria1-/- mice, but had no effect on the locomotion of wild-type mice. Our results show that enhanced neuronal excitability in the hippocampus is associated with pronounced novelty-induced hyperactivity of GluA1 subunit-deficient mice. When this enhanced response of hippocampal neurons to novel stimuli is specifically reduced in the hippocampus by pharmacological treatment or by chemogenetic inhibition, Gria1-/- mice recover from behavioral hyperactivity, suggesting a hippocampal dysfunction in hyperactive behaviors that can be treated with cannabidiol.

Role of Cdk5 in Kalirin7-Mediated Formation of Dendritic Spines.

A majority of excitatory synapses in the brain are localized on the dendritic spines. Alterations of spine density and morphology are associated with many neurological diseases. Understanding the molecular mechanisms underlying spine formation is important for understanding these diseases. Kalirin7 (Kal-7) is localized to the postsynaptic side of excitatory synapses in the neurons. Overexpression of Kal-7 causes an increase in spine density whereas knockdown expression of endogenous Kal-7 results in a decrease in spine density in primary cultured cortical neurons. However, the mechanisms underlying Kal-7-mediated spine formation are not entirely clear. Cyclin-dependent kinase 5 (Cdk5) plays a vital role in the formation of spines and synaptic plasticity. Kal-7 is phosphorylated by CDK5 at Thr1590, the unique Cdk5 phosphorylation site in the Kal-7 protein. This study was to explore the role of CDK5-mediated phosphorylation of Kal-7 in spine formation and the underlying mechanisms. Our results showed expression of Kal-7T/D (mimicked phosphorylation), Kal-7T/A mutants (blocked phosphorylation) or wild-type (Wt) Kal-7 caused in a similar increase in spine density, while spine size of Wt Kal-7-expressing cortical neurons was bigger than that in Kal-7 T\A-expressing neurons, but smaller than that in Kal-7T/D-expressing neurons. The fluorescence intensity of NMDA receptor subunit NR2B (GluN2B) staining was stronger along the MAP2 positive dendrites of Kal-7T/D-expressing neurons than that in Kal-7T/A- or Wt Kal-7-expressing neurons. The fluorescence intensity of AMPA receptor subunit GluR1 (GluA1) staining showed the same trend as GluN2B staining. These findings suggest that Cdk5 affects the function of Kal-7 on spine morphology and function via GluN2B and GluA1 receptors during dendritic spine formation.

Reconsolidation of sucrose instrumental memory in rats: The role of retrieval context

Memory reconsolidation enables the update of a previously consolidated memory trace after its reactivation. Although Pavlovian memory reconsolidation has been widely demonstrated, instrumental memory reconsolidation is still debated. The most critical issue on instrumental memory reconsolidation findings have mainly been linked to the presence of specific boundary conditions for reactivation, for instance contextual parameters. In this study, we investigated the role of the spatial context on molecular markers of sucrose instrumental memory reactivation.Following withdrawal, rats previously conditioned to sucrose self-administration underwent either instrumental memory retrieval or no-retrieval in the conditioned context (Context A, AA condition) or in a modified version of the conditioned context (Context B, AB condition). Two hours later, the level of GluA1 and GluN2B receptors, Zif268 and phosphorylated-rpS6 (rpS6P) was measured in key brain areas for memory reactivation.Retrieval in Context A significantly increased GluA1Rs and GluN2BRs in amygdala compared to no-retrieval, indicating that memory successfully reactivated and destabilized. Moreover, Zif268 level was significantly increased after retrieval in Context A in the nucleus accumbens shell, central and basolateral amygdala but not in the hippocampus, while retrieval in Context B significantly increased Zif268 level in all brain areas. On the other hand, rpS6P level was increased in the nucleus accumbens shell and central amygdala, but decreased in the hippocampus, after retrieval in Context A, while retrieval in Context B did not change rpS6P level in brain areas, except for a small but significant decrease in hippocampus.While the increase of Zif268 level indicated that memory reactivation has been triggered in both the conditions, the lack of change in rpS6P levels after retrieval in Context B – in particular in the central amygdala – suggests that the reconsolidation process could not occur after memory reactivation in a context different from the conditioned one.

NMDA Receptors in Astrocytes: In Search for Roles in Neurotransmission and Astrocytic Homeostasis.

Studies of the last two decades have demonstrated the presence in astrocytic cell membranes of N-methyl-d-aspartate (NMDA) receptors (NMDARs), albeit their apparently low abundance makes demonstration of their presence and function more difficult than of other glutamate (Glu) receptor classes residing in astrocytes. Activation of astrocytic NMDARs directly in brain slices and in acutely isolated or cultured astrocytes evokes intracellular calcium increase, by mutually unexclusive ionotropic and metabotropic mechanisms. However, other than one report on the contribution of astrocyte-located NMDARs to astrocyte-dependent modulation of presynaptic strength in the hippocampus, there is no sound evidence for the significant role of astrocytic NMDARs in astrocytic-neuronal interaction in neurotransmission, as yet. Durable exposure of astrocytic and neuronal co-cultures to NMDA has been reported to upregulate astrocytic synthesis of glutathione, and in this way to increase the antioxidative capacity of neurons. On the other hand, overexposure to NMDA decreases, by an as yet unknown mechanism, the ability of cultured astrocytes to express glutamine synthetase (GS), aquaporin-4 (AQP4), and the inward rectifying potassium channel Kir4.1, the three astroglia-specific proteins critical for homeostatic function of astrocytes. The beneficial or detrimental effects of astrocytic NMDAR stimulation revealed in the in vitro studies remain to be proven in the in vivo setting.

Acute administration of ketamine attenuates the impairment of social behaviors induced by social defeat stress exposure as juveniles via activation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors

The impairment of social behaviors induced by social defeat stress exposure as juveniles is resistant to some antidepressants and an antipsychotic, although the underlying mechanisms and/or therapeutic target are not yet clear. In this study, we investigated the involvement of the glutamatergic neuronal system in the impairment of social behaviors in this model, as this system is known to be involved in many central pathologies. Acute administration of ketamine, a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist and subsequent stimulation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors, attenuated the expression of impairment of social behaviors. Lack of the NMDA receptor GluN2A subunit or acute administration of ifenprodil, an NMDA receptor GluN2B subunit antagonist, did not cause an effect. There were no significant changes in NMDA function, as determined by the ratios of phosphorylated NMDA receptor subunits in the prefrontal cortex and hippocampus. 2,3-Dihydroxy-6-nitro-7-sulfamoyl-benzo[f]quinoxaline-2,3-dione, a selective AMPA receptor antagonist, prevented the effect of ketamine on the expression of impairment of social behaviors. On the contrary, the ratio of phosphorylated AMPA receptor GluA1 subunit in the hippocampus was significantly increased in the non-tested, defeated group. Ketamine increased the level of total protein, but not the ratio of phosphorylated GluA1 in the hippocampus of the non-tested, defeated group. In conclusion, exposure to social defeat stress as juveniles may induce the expression of impairment of social behaviors in adolescents via functional changes in GluA1. Activators of AMPA receptor signaling, such as ketamine, may constitute a novel treatment strategy for stress-related psychiatric disorders in adolescents with adverse juvenile experiences.

Sustained antidepressant action of the N-methyl-D-aspartate receptor antagonist MK-801 in a chronic unpredictable mild stress model.

(5S,10R)-5-methyl-10,11-dihydro-5H-dibenzo(A,D)cyclohepten-5,10-imine hydrogen maleate (MK-801) is an N-methyl-D-aspartate non-competitive antagonist that possesses useful biological properties, including anticonvulsant and anesthetic activities. Studies have indicated the rapid antidepressant effects of MK-801 in animal models. However, there are no reports concerning a sustained antidepressant effect in the chronic unpredictable mild stress (CUMS) model. Furthermore, the antidepressant mechanism remains unclear. The aim of the present study was to examine the effects of MK-801 (0.1 mg/kg) and rapastinel (10 mg/kg) on depression-like behavior in CUMS mice and measure the protein expression of brain-derived neurotrophic factor (BDNF), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (GluA1) and phosphorylated mammalian target of rapamycin (p-mTOR). In the tail suspension and forced swim tests, MK-801 significantly attenuated the increased immobility time in CUMS mice compared with the vehicle group. In the sucrose preference test, a single-dose injection of MK-801 significantly ameliorated the decreased sucrose preference in CUMS mice compared with the vehicle group. Western blot analyses indicated that MK-801 significantly attenuated the decreased BDNF, GluA1 and p-mTOR protein levels in the medial prefrontal cortex (mPFC), dentate gyrus (DG) and CA3 of the hippocampi of CUMS mice. Conversely, this compound had no effect on increased BDNF, GluA1 and p-mTOR protein levels in the nucleus accumbens of CUMS mice. Therefore, the present study revealed the sustained antidepressant effects of MK-801 in the CUMS model. Furthermore, synaptogenesis and neuronal regeneration in the prelimbic regions of mPFC, DG and CA3 of the hippocampus may be implicated as mechanisms that promote a sustained antidepressant response.

Mechanisms underlying the rapid-acting antidepressant-like effects of neuropeptide VGF (non-acronymic) C-terminal peptide TLQP-62

Previous studies have revealed that neuropeptide VGF (non-acronymic) C-terminal peptide TLQP-62 rapidly activates brain-derived neurotrophic factor (BDNF)/tropomyosin receptor kinase B (TrkB)/α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor/mammalian target of rapamycin (mTOR) signaling and produces antidepressant-like actions in rodents. In addition, acute TLQP-62 infusion also markedly changes the AMPA receptor GluA1 subunit phosphorylation at Ser 845 (pGluA1 Ser845) in the PFC of mice, indicating that the GluA1 may contributes to the rapid antidepressant-like effects of TLQP-62. However, how to regulate the TrkB-mediated signaling and GluA1 changes in the prefrontal cortex (PFC) by TLQP-62 remains unclear. Herein, acute administration of TLQP-62 into PFC produced rapid-acting antidepressant-like effects in mice. Additionally, we confirmed that TLQP-62 ameliorated the depression-like behaviors induced by chronic social defeat stress (CSDS) in mice. Further investigation demonstrated that this effect of TLQP-62 was mediated by activation of TrkB and mTOR, which proceeded to decrease bicaudal C homolog 1 gene (BICC1) and increase synaptic protein expression, including GluA1 subunit and pGluA1 Ser845. Notably, we further found that beneficial effects of TLQP-62 on depression-like behaviors and TrkB/mTOR/BICC1 signaling, GluA1 phosphorylation and GluA1 activation in the PFC of mice were significantly abolished by TrkB antagonist ANA-12. In conclusion, our findings indicate that TrkB/mTOR/BICC1 signaling, GluA1 phosphorylation and GluA1 activation in the PFC may involve in the rapid-acting antidepressant-like actions of TLQP-62 in mice.

Essential roles of neuropeptide VGF regulated TrkB/mTOR/BICC1 signaling and phosphorylation of AMPA receptor subunit GluA1 in the rapid antidepressant-like actions of ketamine in mice

Previous studies have suggested that rapid reductions in depression-like behaviors are observed in response to sub-anesthetic-doses of ketamine, an N-methyl-d-aspartate receptor (NMDAR) antagonist. Neuropeptide VGF (non-acronymic) is a critical effector of depression-like behaviors and is thought to be involved in the antidepressant actions of ketamine that have been demonstrated. However, the mechanism underlying the involvement of VGF in the anti-depressant action of ketamine remains unclear. We found that single dose ketamine treatment reversed CSDS-induced depression-like behaviors and decrease of VGF in the PFC of mice. To investigate the involvement of VGF in the antidepressant-like effects of ketamine, a lentivirus vector for VGF was constructed to knockdown the expression of VGF in the prefrontal cortex (PFC) of mice. The biochemical and behavioral effects of this VGF knockdown were examined, using the open field, forced swim, and sucrose preference tests. Our results show that knockdown of VGF increased the immobility time and decreased the sucrose preference in mice. These effects were not improved by ketamine administration. In addition, we found that knockdown of VGF significantly decreased the expression of phosphorylation of tropomyosin receptor kinase B (TrkB), mammalian target of rapamycin (mTOR), and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluA1 Ser845 and increased the expression of bicaudal C homolog 1 (BICC1) in the mouse PFC, and blocked the regulation of TrkB/mTOR/BICC1 signaling and GluA1 phosphorylation by ketamine. Our results indicate that the rapid onset antidepressant-like actions of ketamine require VGF to regulate TrkB/mTOR/BICC1 signaling and AMPA receptor GluA1 phosphorylation.

Electroacupuncture ameliorates cognitive impairment through inhibition of Ca2+-mediated neurotoxicity in a rat model of cerebral ischaemia–reperfusion injury

BackgroundThe hippocampus is vulnerable to severe damage after cerebral ischaemia–reperfusion (I/R) injury. This study aimed to explore the effect of electroacupuncture (EA) on cognitive impairment and its relationship with Ca2+neurotoxicity...

Hippocampal amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid GluA1 (AMPA GluA1) receptor subunit involves in learning and memory improvement following treatment with Centella asiatica extract in adolescent rats.

Introduction Centella asiatica is an herbal plant that contains phytochemicals that are widely believed to have positive effects on cognitive function. The adolescent stage is a critical development period for the maturation of brain processes that encompass changes in physical and psychological systems. However, the effect of C. asiatica has not been extensively studied in adolescents. The aim of this study was therefore to investigate the effects of a C. asiatica extract on the enhancement of learning and memory in adolescent rats.MethodsThe locomotor activity, learning, and memory were assessed by using open field test and water T‐maze test. This study also examined changes in neuronal cell morphology using cresyl violet and apoptosis staining. We also performed immunohistochemical study to analyse the expression of the glutamate AMPA receptor (α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid) GluA1 subunit and the GABA receptor (γ‐Aminobutyric Acid) subtype GABAA α1 subunit in the hippocampus of the same animals.ResultsWe found no significant changes in locomotor activity (p > 0.05). The water T‐maze data showed that 30 mg/kg dose significantly (p < 0.05) improved learning, memory, and the memory consolidation phase but had no effect on reversal learning (p > 0.05). Histological data revealed no neuronal morphological changes. Immunohistochemical analysis revealed increased expression of the AMPA GluA1 receptor subunit but there was no effect on GABAA receptor α1 subunit expression in the CA1 and CA2 subregions of the hippocampus.ConclusionsThe C. asiatica extract therefore improved hippocampus‐dependent spatial learning and memory in a dose‐dependent manner in rats through the GluA1‐containing AMPA receptor in the CA1 and CA2 sub regions of the hippocampus.

Activation of Protein Kinase G After Repeated Cocaine Administration Is Necessary for the Phosphorylation of α-Amino-3-Hydroxy-5-Methyl-4-Isoxazolepropionic Acid Receptor GluA1 at Serine 831 in the Rat Nucleus Accumbens.

Phosphorylation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors in the striatum plays a crucial role in regulating the receptor-coupled signaling cascades leading to behavioral changes associated with psychostimulant exposure. The present study determined if activation of protein kinase G (PKG) contributes to the phosphorylation of AMPA receptor GluA1 subunit at the position of serine 831 (GluA1-S831) in the rat nucleus accumbens (NAc) after repeated cocaine administration. The results demonstrated that repeated intraperitoneal (i.p.) injections of cocaine (20 mg/kg) once a day for seven consecutive days significantly increased the level of phosphorylated (p)GluA1-S831. This increase was decreased by the intra-NAc infusion of either the cyclic guanosine monophosphate (cGMP) analog, Rp-8-Br-PET-cGMPS (5 nmol/1 μL), or the PKG inhibitor, KT5823 (2 nmol/1 μL). Repeated cocaine administration increased PKG binding activity to GluA1. This increase in GluA1-S831 phosphorylation after repeated cocaine administration was decreased by the intra-NAc infusion of the synthetic peptide (Tat-tagged interfering peptide (Tat-GluA1-i)), that interferes with the binding of PKG to GluA1. Intra-NAc infusion of the interfering peptide also reduced the repeated cocaine-induced increase in locomotor activity. These findings suggest that activated PKG, after repeated exposure to cocaine, binds to AMPA receptor GluA1 and is required for the phosphorylation of S831, contributing to behavioral changes.

Developmental Roles and Evolutionary Significance of AMPA-Type Glutamate Receptors.

Organogenesis and metamorphosis require the intricate orchestration of multiple types of cellular interactions and signaling pathways. Glutamate (Glu) is an excitatory extracellular signaling molecule in the nervous system, while Ca2+ is a major intracellular signaling molecule. The first Glu receptors to be cloned are Ca2+ -permeable receptors in mammalian brains. Although recent studies have focused on Glu signaling in synaptic mechanisms of the mammalian central nervous system, it is unclear how this signaling functions in development. Our recent article demonstrated that Ca2+ -permeable AMPA-type Glu receptors (GluAs) are essential for formation of a photosensitive organ, development of some neurons, and metamorphosis, including tail absorption and body axis rotation, in ascidian embryos. Based on findings in these embryos and mammalian brains, we formed several hypotheses regarding the evolution of GluAs, the non-synaptic function of Glu, the origin of GluA-positive neurons, and the neuronal network that controls metamorphosis in ascidians.

Effect of Pb Exposure on Synaptic Scaling Through Regulation of AMPA Receptor Surface Trafficking.

Homeostatic synaptic plasticity (HSP) helps to stabilize the neuronal network activity, which is essential for optimal information coding. Synaptic scaling is a form of homeostatic plasticity that stabilizes neuronal firing in response to activity blockade. Lead (Pb) is a ubiquitous environmental neuro-toxicant and can impair the input-specific Hebbian type synaptic plasticity, but whether Pb exerts effects in HSP remains unknown. We previously reported that blocking L-type calcium channel induces synaptic scaling, which stimulates the synthesis of all-trans retinoic acid (RA) and the expression of GluA2-lacking α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor. Given Pb is a potent blocker of calcium channel, we hypothesized Pb may participate in synaptic scaling accompanied by RA synthesis and AMPA receptor trafficking. In this study, cultured hippocampal neurons were treated with Pb (1 μM 5 min, 15 min, 4 h, 24 h, and 10 μM 24 h) alone or in combination with tetrodotoxin (TTX, 1 μM, 24 h). The results showed that Pb alone, either at 1 μM or 10 μM, cannot induce synaptic scaling. But Pb participated in synaptic scaling when concurrent with TTX (10 μM Pb + 1 μM TTX, 24 h). Further results showed that surface heteromeric GluA1 and GluA2 AMPA receptors were increased in TTX+ Pb-induced synaptic scaling. In addition, RA was proved not to participate in TTX+ Pb-mediated synaptic scaling. Taken together, our work supported that TTX+ Pb could induce synaptic scaling and enhance synaptic accumulation of AMPAR GluA1 and GluA2 during synaptic up scaling. Our study would help for elucidation of the Pb-induced neuronal network instability mechanism.

Feasibility study for a glutamate driven subretinal prosthesis: local subretinal application of glutamate on blind retina evoke network-mediated responses in different types of ganglion cells

Objective. A feasibility study for a transmitter based subretinal prosthesis, generating visual responses in blind mouse retina is presented. Approach. Degenerated rd1 mouse retina were stimulated in subretinal configuration by local glutamate (Glu) or NMDA application via micropipettes (~1.5 μm) and thereby the outer retinal activity was recorded by calcium-imaging or the ganglion cell (GC) activity was recorded by the multi-electrode array system. The network mediated activation of GC via bipolar cells was approved by the administration of Glu receptor blockers. Main results. Data of the degenerated and blind rd1 mouse retina reveals that the outer retina is Glu sensitive and that the subretinal Glu stimulation promotes network mediated GC responses. Analysis of the spatial activity-spread indicates that the Glu induced cell activation radius in the outer retina (~12.5 μm) and postsynaptically activated GC (~40 μm) is focal to the stimulation pipette tip. Moreover, the application of NMDA in subretinal space also evoked network mediated GC responses. The Glu-activated GC were identified as ON–OFF, OFF and two ON cells types. Significance. This study evaluates the prerequisite for the function of a transmitter based implant, that after the loss of the photoreceptors, the remnant blind retinal network is Glu sensitive and functional, positively. The differential activation of ON (hyperpolarisation) and OFF (depolarisation) bipolar cells by transmitter Glu is a unique feature and of high interest for retinal implants. Therefore, the respective bipolar cell types could only be driven by glutamatergic stimulation accurately and not by electrical stimulation. The preserved functionality of the blind retina at the onset of complete blindness is motivating to continue research on a transmitter-based prosthesis. Since the artificial Glu stimulation mimics the natural retinal input, early implantation of a Glu-prosthesis might delay the devastating retinal remodelling positively, due to the neuronal-plasticity.

Ultrastructural Changes in the Frog Brain in the Presence of High Concentrations of Glutamate and an NO-Generating Compound

This review summarizes published data and original findings on the morphology of neurons and glial cells in normal conditions and model conditions that simulate those in the human brain during stroke. Ultrastructural changes that occur in the presence of high concentrations of glutamate (Glu, 0.1–5.0 M) and the NO-generating compound NaNO2 (0.1–5.0 mM) were studied in the cerebella (frog cerebellum), which is one of the simplest circuitries structurally. Such studies and data analyses are important because hyperstimulation of Glu receptors is a leading pathogenetic factor of neuronal damage during a stroke. High Glu concentrations exert a toxic effect and damage cerebellar neurons and glial cells. Mitochondria are de-energized, ionic homeostasis is distorted, the intracellular Ca2+ concentration increases, and constitutive NO synthases are activated in the process. The changes result in an increase in the contents of NO and its transformation products, which are involved in a negative feedback mechanism from postsynaptic neurons to presynaptic cells. Biochemical processes are consequently affected, and morphological changes are induced in neurons and glial cells, leading to their swelling. At the same time, ultrastructural compensatory adaptive mechanisms develop to reduce the damaging effect of high concentrations of Glu and NO-generating compounds.

F196. DIFFERENTIAL EFFECTS OF MGLU5 RECEPTOR BLOCKADE ON BEHAVIOR, SCHIZOPHRENIA-RELEVANT GENE EXPRESSION AND NEURONAL ACTIVATION PATTERNS FROM DEVELOPMENT TO AGING MICE

BackgroundThe glutamate system is implicated both in schizophrenia and mood disorders. Mice lacking metabotropic mGlu5 receptors (mGluR5 KO) display schizophrenia-like abnormalities. Additionally, mGluR5 antagonists represent promising alternative anxiolytics/antidepressants. However, the underlying age-specific molecular/cellular mechanisms are only partially understood. We aimed at identifying molecular alterations associated with a genetically induced mGluR5 deletion, which results in a schizophrenia-like phenotype. Additionally, we investigated age-specific effects of mGluR5 antagonists on emotional behaviour and c-fos activation.MethodsFor analysis of mRNA and protein levels we performed Real-time RT-PCR and Western blot investigations in the hippocampus and prefrontal/frontal cortex (PFC/FC) of mice with a genetic deletion of the metabotropic glutamate receptor 5 (mGlu5), addressing key components of the GABAergic and glutamatergic systems. Additionally, we used classical behavioral tests for determining anxiety- and depression-like changes triggered by the mGluR5 antagonist 2-Methyl-6-(phenylethynyl)pyridine (MPEP). Finally, we used profiling of c-Fos expression, as marker of neuronal activity, induced by MPEP from postnatal day 16 (P16) to adulthood (P90).ResultsmGlu5 knockout (KO) mice showed a significant reduction of reelin, GAD65, GAD67 and parvalbumin mRNA levels, which is specific for the PFC/FC, and that is paralleled by a significant reduction of protein levels in male KO mice. We also analysed the main NMDA and AMPA receptor subunits, namely GluN1, GluN2A, GluN2B and GluA1, and observed that mGlu5 deletion determined a significant reduction of their mRNA levels, also within the hippocampus, with differences between the two genders. We measured age-specific alterations in emotional behaviour of mGluR5 KO mice, with marked increase of anxiety during aging. There was a remarkably conserved activation of the paraventricular nucleus of the hypothalamus, implicated in stress regulation, by MPEP at all investigated ages, whereas the extended amygdala was specifically activated in adulthood only.DiscussionOur data suggest that neurochemical abnormalities impinging the glutamatergic and GABAergic systems may be responsible for the behavioral phenotype associated with mGlu5 KO animals and point to the close interaction of these molecular players for the development of neuropsychiatric disorders such as schizophrenia. These data could contribute to a better understanding of the involvement of mGlu5 alterations in the molecular imbalance between excitation and inhibition underlying the emergence of a schizophrenic-like phenotype and to understand the potential of mGlu5 modulators in reversing the deficits characterizing the schizophrenic pathology.

Glial loss of the metallo β-lactamase domain containing protein, SWIP-10, induces age- and glutamate-signaling dependent, dopamine neuron degeneration

Across phylogeny, glutamate (Glu) signaling plays a critical role in regulating neural excitability, thus supporting many complex behaviors. Perturbed synaptic and extrasynaptic Glu homeostasis in the human brain has been implicated in multiple neuropsychiatric and neurodegenerative disorders including Parkinson’s disease, where theories suggest that excitotoxic insults may accelerate a naturally occurring process of dopamine (DA) neuron degeneration. In C. elegans, mutation of the glial expressed gene, swip-10, results in Glu-dependent DA neuron hyperexcitation that leads to elevated DA release, triggering DA signaling-dependent motor paralysis. Here, we demonstrate that swip-10 mutations induce premature and progressive DA neuron degeneration, with light and electron microscopy studies demonstrating the presence of dystrophic dendritic processes, as well as shrunken and/or missing cell soma. As with paralysis, DA neuron degeneration in swip-10 mutants is rescued by glial-specific, but not DA neuron-specific expression of wildtype swip-10, consistent with a cell non-autonomous mechanism. Genetic studies implicate the vesicular Glu transporter VGLU-3 and the cystine/Glu exchanger homolog AAT-1 as potential sources of Glu signaling supporting DA neuron degeneration. Degeneration can be significantly suppressed by mutations in the Ca2+ permeable Glu receptors, nmr-2 and glr-1, in genes that support intracellular Ca2+ signaling and Ca2+-dependent proteolysis, as well as genes involved in apoptotic cell death. Our studies suggest that Glu stimulation of nematode DA neurons in early larval stages, without the protective actions of SWIP-10, contributes to insults that ultimately drive DA neuron degeneration. The swip-10 model may provide an efficient platform for the identification of molecular mechanisms that enhance risk for Parkinson’s disease and/or the identification of agents that can limit neurodegenerative disease progression.

M1 muscarinic receptor facilitates cognitive function by interplay with AMPA receptor GluA1 subunit.

M1 muscarinic acetylcholine receptors (M1 mAChRs) are the most abundant muscarinic receptors in the hippocampus and have been shown to have procognitive effects. AMPA receptors (AMPARs), an important subtype of ionotropic glutamate receptors, are key components in neurocognitive networks. However, the role of AMPARs in procognitive effects of M1 mAChRs and how M1 mAChRs affect the function of AMPARs remain poorly understood. Here, we found that basal expression of GluA1, a subunit of AMPARs, and its phosphorylation at Ser845 were maintained by M1 mAChR activity. Activation of M1 mAChRs promoted membrane insertion of GluA1, especially to postsynaptic densities. Impairment of hippocampus-dependent learning and memory by antagonism of M1 mAChRs paralleled the reduction of GluA1 expression, and improvement of learning and memory by activation of M1 mAChRs was accompanied by the synaptic insertion of GluA1 and its increased phosphorylation at Ser845. Furthermore, abrogation of phosphorylation of Ser845 residue of GluA1 ablated M1 mAChR-mediated improvement of learning and memory. Taken together, these results show a functional correlation of M1 mAChRs and GluA1 and the essential role of GluA1 in M1 mAChR-mediated cognitive improvement.-Zhao, L.-X., Ge, Y.-H., Xiong, C.-H., Tang, L., Yan, Y.-H., Law, P.-Y., Qiu, Y., Chen, H.-Z. M1 muscarinic receptor facilitates cognitive function by interplay with AMPA receptor GluA1 subunit.

AMPA receptor GluA1 Ser831 phosphorylation is critical for nitroglycerin-induced migraine-like pain

Migraine is the third most common disease worldwide; however, the mechanisms underlying migraine headache are still not fully understood. Previous studies have demonstrated that α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor phosphorylation plays an important role in central sensitization of pain transmission. In the present study, we observed that AMPA receptor GluA1 Ser831 phosphorylation was enhanced in the spinal trigeminal nucleus caudalis (Sp5C) after intraperitoneal injection of nitroglycerin (NTG). The NTG injection induced acute migraine-like pain including photophobia and mechanical hypersensitivity as reported previously. Interestingly, targeted mutation of GluA1 Ser831 site to prevent phosphorylation significantly inhibited NTG-induced migraine-like pain. Moreover, NTG incubation caused a robust Ca2+ influx in cultured brainstem neurons, which was dramatically inhibited by GluA1 S831A (serine at the 831 site of GluA1 is mutated to alanine) phospho-deficient mutation, and treatment with 1-naphthyl acetyl spermine (NASPM), a selective Ca2+-permeable AMPA receptor channel blocker, dose-dependently blocked the NTG-evoked increase of Ca2+ influx in the cultured neurons. We further found that intra-Sp5C injection of NASPM significantly inhibited NTG-produced mechanical hypersensitivity. These results suggest that AMPA receptor phosphorylation at the Ser831 site in the Sp5C is critical for NTG-induced migraine-like pain.