ro-25-6981 and 5-(alpha-methyl-4-bromobenzylamino)phosphonomethyl-1-4-dihydroquinoxaline-2-3-dione

Long-term potentiation (LTP) is a neurobiological mechanism of cognitive function, and the N-methyl-D-aspartate (NMDA) receptors is fundamental for LTP. Previous studies showed that over activation of NMDA receptors may be a crucial cause of LTP and cognitive impairment induced by stress or corticosterone. However, other studies showed that the function of NMDA receptors is insufficient since the NMDA receptors co-agonist D-serine could improve stress-induced cognitive impairment. The purpose of this study is to clarify whether over activation of NMDA receptors or hypofunction of NMDA receptors is involved in hippocampal impairment of LTP by corticosterone and the underlying mechanisms. Results showed that hippocampal LTP and object location recognition memory were impaired in corticosterone-treated mice. Corticosterone increased the glutamate level in hippocampal tissues, neither NMDA receptors antagonist nor its subtype antagonists alleviated impairment of LTP, while enhancing the function of NMDA receptors by D-serine did alleviate impairment of LTP by corticosterone, suggesting that hypofunction of NMDA receptors might be one of the main reasons for impairment of LTP by corticosterone. Further results showed that the level of D-serine and its precursor L-serine did not change. D-serine release-related protein Na

Topics: Animals; Corticosterone; Dentate Gyrus; Excitatory Amino Acid Antagonists; Long-Term Potentiation; Male; Mice, Inbred BALB C; Mice, Inbred C57BL; Open Field Test; Perforant Pathway; Phenols; Piperidines; Quinolones; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Serine

24S-hydroxycholesterol (24HC) is the major metabolic breakdown product of cholesterol in the brain. Among its other effects on neurons, 24HC modulates N-methyl-d-aspartate (NMDA or GluN) receptors, but our understanding of this mechanism is poor. We used whole-cell patch clamp recordings and various pharmacological approaches in mouse brain slices to record isolated NMDAR-mediated (I

Topics: Animals; Cholesterol 24-Hydroxylase; Dentate Gyrus; Evoked Potentials; Excitatory Postsynaptic Potentials; Hydroxycholesterols; Interneurons; Male; Mice; Mice, Knockout; Neurons; Phenols; Piperidines; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Synaptic Potentials

Cannabis affects cognitive performance through the activation of the endocannabinoid system, and the molecular mechanisms involved in this process are poorly understood. Using the novel object-recognition memory test in mice, we found that the main psychoactive component of cannabis, delta9-tetrahydrocannabinol (THC), alters short-term object-recognition memory specifically involving protein kinase C (PKC)-dependent signaling. Indeed, the systemic or intra-hippocampal pre-treatment with the PKC inhibitors prevented the short-term, but not the long-term, memory impairment induced by THC. In contrast, systemic pre-treatment with mammalian target of rapamycin complex 1 inhibitors, known to block the amnesic-like effects of THC on long-term memory, did not modify such a short-term cognitive deficit. Immunoblot analysis revealed a transient increase in PKC signaling activity in the hippocampus after THC treatment. Thus, THC administration induced the phosphorylation of a specific Ser residue in the hydrophobic-motif at the C-terminal tail of several PKC isoforms. This significant immunoreactive band that paralleled cognitive performance did not match in size with the major PKC isoforms expressed in the hippocampus except for PKCθ. Moreover, THC transiently enhanced the phosphorylation of the postsynaptic calmodulin-binding protein neurogranin in a PKC dependent manner. These data demonstrate that THC alters short-term object-recognition memory through hippocampal PKC/neurogranin signaling.

Topics: Animals; Anisomycin; Dizocilpine Maleate; Dose-Response Relationship, Drug; Dronabinol; Drug Interactions; Hippocampus; Isoenzymes; Male; Memory, Short-Term; Mice; Microinjections; Neurogranin; Phenols; Phosphorylation; Piperidines; Protein Kinase C; Quinoxalines; Rimonabant; Signal Transduction; Sirolimus

While it has been shown that the blockade of N-methyl-d-aspartate type glutamate receptors (NMDARs) impairs memory acquisition, recent studies have reported that the post-acquisition administration of NMDAR antagonists suppresses spatial memory decay. These findings suggest that NMDARs are important not only for the acquisition of new memories but also for the decay of previously acquired memories. The present study investigated the contributions of specific NMDAR subunits to spatial memory decay using NVP-AAM077 (NVP), an NMDAR antagonist that preferentially binds to GluN2A subunits, and the selective GluN2B blocker Ro 25-6981 (Ro). Following Morris water maze training (four trials/day for four days), NVP and/or Ro were subchronically infused into the rat hippocampus for five days. Seven days after training, NVP-treated rats and NVP/Ro-treated rats explored the target area significantly more than the control and Ro-treated rats. These results demonstrate that post-acquisition treatment with NVP, but not Ro, suppresses the forgetting of previously acquired spatial memories. The NVP-treated rats more persistently explored the target area in the second test, which was conducted one day after the first, while the NVP/Ro-treated rats did not, which suggest that Ro treatment downregulates memory retention. In conclusion, the present results indicate that the NMDAR GluN2A and GluN2B subunits contribute to spatial memory deterioration and maintenance, respectively.

Topics: Animals; Behavior, Animal; Excitatory Amino Acid Antagonists; Hippocampus; Maze Learning; Phenols; Piperidines; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Retention, Psychology; Spatial Memory

Brain-derived neurotrophic factor (BDNF) has been investigated for its positive role in regulation of fear acquisition and memory. The precursor of BDNF, proBDNF, has been identified as different protein from its mature form. The prelimbic (PL) and infralimbic (IL) sub-regions of the medial prefrontal cortex (mPFC) are functionally distinct in fear behavior. However, the role of PL and IL proBDNF in fear memory is unclear. Here, through the infusion of cleavage-resistant proBDNF and its antibody, we identified the dissociable roles of PL and IL proBDNF in fear expression and extinction memory as well as explored proBDNF's potential mechanism of action. The results suggest that the infusion of proBDNF in the IL facilitates induction of fear extinction, while infusion in the PL depresses fear expression. Blocking proBDNF by using its antibody disrupted the acquisition of fear extinction in the IL, but not the PL. Furthermore, proBDNF-induced extinction was sufficient for extinguishing new and older memories, and required NR2B, but not NR2A, -containing NMDA receptors. We also observed extinction-related proBDNF expression increased in the PL and IL during successful fear expression and extinction, respectively. Importantly, enhanced proBDNF was required for maintaining an extinguished behavior. The extinction effects of proBDNF did not involve degrading the original fear memory. Therefore, proBDNF in the IL and PL differentially contribute to the inhibitory control of fear extinction behavior. Our findings provide a strong link between proBDNF activity and deficits in fear extinction, a hallmark of several psychiatric disorders.

Topics: Analysis of Variance; Animals; Antibodies; Brain-Derived Neurotrophic Factor; Conditioning, Psychological; Excitatory Amino Acid Antagonists; Extinction, Psychological; Fear; Male; Phenols; Piperidines; Prefrontal Cortex; Quinoxalines; Rats; Rats, Sprague-Dawley

In the CA1 area of the hippocampus N-methyl-d-aspartate receptors (NMDARs) mediate the induction of long-term depression (LTD), short-term potentiation (STP) and long-term potentiation (LTP). All of these forms of synaptic plasticity can be readily studied in juvenile hippocampal slices but the involvement of particular NMDAR subunits in the induction of these different forms of synaptic plasticity is currently unclear. Here, using NVP-AAM077, Ro 25-6981 and UBP145 to target GluN2A-, 2B- and 2D-containing NMDARs respectively, we show that GluN2B-containing NMDARs (GluN2B) are involved in the induction of LTD, STP and LTP in slices prepared from P14 rat hippocampus. A concentration of Ro (1 μM) that selectively blocks GluN2B-containing diheteromers is able to block LTD. It also inhibits a component of STP without affecting LTP. A higher concentration of Ro (10 μM), that also inhibits GluN2A/B triheteromers, blocks LTP. UBP145 selectively inhibits the Ro-sensitive component of STP whereas NVP inhibits LTP. These data are consistent with a role of GluN2B diheretomers in LTD, a role of both GluN2B- and GluN2D- containing NMDARs in STP and a role of GluN2A/B triheteromers in LTP. This article is part of the Special Issue entitled 'Ionotropic glutamate receptors'.

Topics: Animals; Hippocampus; Long-Term Potentiation; Long-Term Synaptic Depression; Phenols; Piperidines; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

Ischemic postconditioning (Post C), which involves administration of a brief ischemia after the initial ischemic event, has been demonstrated to be strongly neuroprotective against global cerebral ischemia (GCI) and to improve cognitive outcome. To enhance understanding of the underlying mechanisms, the current study examined the role of NMDA receptors in mediating the beneficial effects of Post C (3 min ischemia) administered 2 days after GCI in adult male rats. The results revealed that Post C was strongly neuroprotective against GCI, and that this effect was blocked by administration of the NMDA receptor antagonist MK-801. Further work revealed that the NR2A-type NMDA receptors mediate the Post C beneficial effects as administration of a NR2A-preferring antagonist (NVP-A) blocked Post C neuroprotection and cognitive enhancement, while administration of a NR2B-preferring antagonist (Ro25) was without effect. Post C significantly up-regulated NR2A levels and phosphorylation of NR2A in the hippocampal CA1 region after Post C. Post C also increased Ca(2+) influx and activation/phosphorylation of CamKIIα at Thr(286), effects that were NR2A mediated as they were blocked by NVP-A. Phosphorylation of ERK and CREB was also increased by Post C, as were two downstream CREB-dependent prosurvival factors, brain derived neurotropic factor (BDNF) and Bcl2, effects that were blocked by the NR2A antagonist, NVP-A. Taken as a whole, the current study provides evidence that NR2A-activation and downstream prosurvival signaling is a critical mediator of Post C-induced neuroprotection and cognitive enhancement following GCI.

Topics: Analysis of Variance; Animals; Brain Ischemia; Calcium; CREB-Binding Protein; Dizocilpine Maleate; Hippocampus; Immunoprecipitation; Male; Maze Learning; Neuroprotective Agents; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Time Factors

Neonatal hypoxia-ischemia brain damage is an important cause of death by affecting prognosis of neural diseases. It is difficult to find effective methods of prevention and treatment due to the complexity of its pathogenesis. N-methyl-D-aspartate (NMDA), as an excitotoxicity amino acids, has proven to play an important role in hypoxic-ischemic. However, the exact effects of the NMDA subunits, NR2A and NR2B, during hypoxic-ischemic have not been investigated in detail. Therefore, we sought to study whether the NMDA receptor antagonist could confer neuroprotective effects in a neonatal rat hypoxia-ischemia model. The effects of intraperitoneal injections of different drugs, namely MK-801 (0.5 mg/kg), NVP-AAM077 (5 mg/kg), and Ro25-6981 (5 mg/kg), on the expressions of anti-apoptotic protein Bcl-2 and apoptosis protein Bax in the subventricular zone were analyzed by immunohistochemical staining to explore the roles of NMDA subunits (NR2A and NR2B) in hypoxic-ischemic. We found that the NR2B antagonist (Ro25-6981) could inhibit hypoxic-ischemic with the increasing Bcl-2 expression. NR2A antagonists (NVP-AAM077) can increase cerebral hypoxia-ischemia in neonatal rats, promoting the expression of apoptotic protein Bax.

Topics: Animals; bcl-2-Associated X Protein; Disease Models, Animal; Dizocilpine Maleate; Hypoxia-Ischemia, Brain; Immunohistochemistry; Lateral Ventricles; Neuroprotective Agents; Phenols; Piperidines; Protein Subunits; Proto-Oncogene Proteins c-bcl-2; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate

Paradoxically, N-methyl-D-aspartate (NMDA) receptor antagonists are used to model certain aspects of schizophrenia as well as to treat refractory depression. However, the role of different subunits of the NMDA receptor in both conditions is poorly understood. Here we used biochemical and behavioral readouts to examine the in vivo prefrontal efflux of serotonin and glutamate as well as the stereotypical behavior and the antidepressant-like activity in the forced swim test elicited by antagonists selective for the GluN2A (NVP-AAM077) and GluN2B (Ro 25-6981) subunits. The effects of the non-subunit selective antagonist, MK-801; were also studied for comparison. The administration of MK-801 dose dependently increased the prefrontal efflux of serotonin and glutamate and markedly increased the stereotypy scores. NVP-AAM077 also increased the efflux of serotonin and glutamate, but without the induction of stereotypies. In contrast, Ro 25-6981 did not change any of the biochemical and behavioral parameters tested. Interestingly, the administration of NVP-AAM077 and Ro 25-6981 alone elicited antidepressant-like activity in the forced swim test, in contrast to the combination of both compounds that evoked marked stereotypies. Our interpretation of the results is that both GluN2A and GluN2B subunits are needed to induce stereotypies, which might be suggestive of potential psychotomimetic effects in humans, but the antagonism of only one of these subunits is sufficient to evoke an antidepressant response. We also propose that GluN2A receptor antagonists could have potential antidepressant activity in the absence of potential psychotomimetic effects.

Topics: Animals; Antidepressive Agents; Depressive Disorder, Treatment-Resistant; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Excitatory Amino Acid Antagonists; Glutamic Acid; Male; Phenols; Piperidines; Prefrontal Cortex; Quinoxalines; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Schizophrenia; Serotonin; Stereotyped Behavior

Dual probe microdialysis was used to investigate whether GluN2A and GluN2B NMDA receptor subunits regulate striatal output pathways under dyskinetic conditions. The preferential GluN2A antagonist NVP-AAM077 perfused in the dopamine-depleted striatum of 6-hydroxydopamine hemilesioned dyskinetic rats reduced GABA and glutamate levels in globus pallidus whereas the selective GluN2B antagonist Ro 25-6981 elevated glutamate without affecting pallidal GABA. Moreover, intrastriatal NVP-AAM077 did not affect GABA but elevated glutamate levels in substantia nigra reticulata whereas Ro 25-6981 elevated GABA and reduced nigral glutamate. To investigate whether GluN2A and GluN2B NMDA receptor subunits are involved in motor pathways underlying dyskinesia expression, systemic NVP-AAM077 and Ro 25-6981 were tested for their ability to attenuate levodopa-induced abnormal involuntary movements. NVP-AAM077 failed to prevent dyskinesia while Ro 25-6981 mildly attenuated it. We conclude that in the dyskinetic striatum, striatal GluN2A subunits tonically stimulate the striato-pallidal pathway whereas striatal GluN2B subunits tonically inhibit striato-nigral projections. Moreover, GluN2A subunits are not involved in dyskinesia expression whereas GluN2B subunits minimally contribute to it.

Topics: Animals; Corpus Striatum; Dopamine; Dopamine Agents; Dyskinesia, Drug-Induced; gamma-Aminobutyric Acid; Globus Pallidus; Glutamic Acid; Levodopa; Male; Microdialysis; Neostriatum; Oxidopamine; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Substantia Nigra

Chronic global N-methyl-d-aspartate receptor (NMDAR) blockade leads to changes in glutamatergic transmission. The impact of more subunit-selective NMDAR inhibition on glutamatergic circuits remains incomplete. To this end, organotypic hippocampal slice cultures were treated for 17-21 days with the high-affinity competitive antagonist d-aminophosphonovaleric acid (d-APV), the allosteric GluN2B-selective antagonist Ro25-6981, or the newer competitive GluN2A-preferring antagonist NVP-AAM077. Electrophysiological recordings from dentate granule cells revealed that chronic d-APV treatment increased, whereas chronic Ro25-6981 reduced, epileptiform event-associated large-amplitude spontaneous excitatory postsynaptic currents (sEPSC) compared with all other treatment groups, consistent with opposite effects on glutamatergic networks. Presynaptically, chronic d-APV or Ro25-6981 increased small-amplitude sEPSCs and AMPA/kainate receptor-mediated miniature EPSCs (mEPSCAMPAR) frequency. Chronic d-APV or NVP-AAM077, but not Ro25-6981, increased putative vGlut1-positive glutamatergic synapses. Postsynaptically, chronic d-APV dramatically increased mEPSCAMPAR and profoundly decreased NMDAR-mediated mEPSC (mEPSCNMDAR) measures, suggesting increased AMPAR/NMDAR ratio. Ro25-6981 decreased mEPSCAMPAR charge transfer and modestly decreased mEPSCNMDAR frequency and decay, suggesting downward scaling of AMPAR and NMDAR function without dramatically altering AMPAR/NMDAR ratio. Extrasynaptically, threo-β-benzyloxyaspartate-enhanced "tonic" NMDAR current amplitude and activated channel number estimates were significantly increased only by chronic Ro25-6981. For intrinsic excitability, action potential threshold was slightly more negative following chronic d-APV or NVP-AAM077. The predominant pro-excitatory effects of chronic d-APV are consistent with increased glutamatergic transmission and network excitability. The minor effects of chronic NVP-AAM077 on action potential threshold and synapse number are consistent with minimal effects on circuit function. The chronic Ro25-6981-induced downward scaling of synaptic AMPAR and NMDAR function is consistent with decreased postsynaptic glutamate receptors and reduced network excitability.

Topics: 2-Amino-5-phosphonovalerate; Animals; Dentate Gyrus; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Miniature Postsynaptic Potentials; Neuronal Plasticity; Neurons; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Vesicular Glutamate Transport Protein 1

GluN receptors are heteromers of obligatory GluN1 subunits and GluN2(A-D) subunits. In the present study, we addressed the question whether GluN2A and GluN2B subunits play distinct roles in the formation of filopodia and dendrites during the early development of hippocampal neurons. In hippocampal neurons brought into culture at embryonic day 17, we studied 2-3 days later the effects of N-methyl-D-aspartic acid (NMDA) on the numbers of filopodia, growth cones, and primary as well as secondary dendrites. Antagonists specific for GluN2A and GluN2B subunits were applied together with NMDA. NMDA only induced the formation of filopodia and secondary dendrites. Whereas filopodia were generated within 15 min by NMDA alone, secondary dendrites were only induced by the joint application of NMDA and the Rho kinase inhibitor Y-27632 for 24 h. The GluN2B antagonists ifenprodil and Ro 25-6981 prevented the NMDA-induced formation of filopodia, whereas the GluN2A antagonists NVP-AAM007 and EAA-090 prevented the formation of secondary dendrites. We conclude that both GluN2 subunits have differential roles in dendritic arborization.

Topics: Animals; Azabicyclo Compounds; Cells, Cultured; Dendrites; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; HeLa Cells; Hippocampus; Humans; N-Methylaspartate; Neurons; Organophosphonates; Phenols; Piperidines; Protein Subunits; Pseudopodia; Quinoxalines; Rats; Receptors, N-Methyl-D-Aspartate

The role of N-methyl-D-aspartate receptors (NMDARs) in the generation and maintenance of epileptic seizures has been widely investigated, however, little is known of possible separate roles played by NMDARs that contain different NR2 subunits. A better comprehension of how distinct NMDARs subtypes participate in seizure generation and/or diffusion may lead to the development of more targeted pharmacologic strategies to treat epilepsy. Therefore, we have performed an electrophysiologic investigation using a multielectrode array device, on slices comprising entorhinal cortex (EC) and hippocampus, continuously perfused in a Mg(2+) -free medium, with added 4-aminopiridine (4AP; 10-15 μm). Two separate rhythmic patterns of interictal-like activity were generated in EC and hippocampus, with EC seizures entrained to those in CA3, so that a significant degree of cross-correlation occurred. Perfusion with the NR2A-containing NMDAR antagonist [(R)-[(S)-1-(4-bromo-phenyl)-ethylamino]-(2,3-dioxo-1,2,3,4-tetrahydroquinoxalin-5-yl)-methyl]-phosphonic acid (NVP-AAM077; 50 nm) or Zn(2+) (200 nm), did not affect the rate of interictal-like events in EC and hippocampus; however, it significantly reduced their cross-correlation, causing a substantial decoupling of the two rhythm generators. The same effect was observed with (αR,βS)-α-(4-hydroxyphenyl)-β-methyl-4-(phenylmethyl)-1-piperidinepropanol maleate (Ro25-6981; 1 μm), when coapplied with a subthreshold dose of NVP-AAM077. Our results suggest that NR2 subunits may be crucial in entraining cortical networks, leading to recruitment of wider range oscillations during epilepsy. Therefore, a pharmacologic strategy directed onto NR2 subunits may help to limit seizure diffusion and recruitment of potentially entrained oscillatory networks.

Topics: 2-Amino-5-phosphonovalerate; Action Potentials; Animals; Dose-Response Relationship, Drug; Drug Interactions; Entorhinal Cortex; Excitatory Amino Acid Antagonists; Hippocampus; In Vitro Techniques; Neural Pathways; Patch-Clamp Techniques; Phenols; Piperidines; Potassium Channel Blockers; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate

Converging evidence shows that monoamine oxidase A (MAO A), the key enzyme catalyzing serotonin (5-hydroxytryptamine; 5-HT) and norepinephrine (NE) degradation, is a primary factor in the pathophysiology of antisocial and aggressive behavior. Accordingly, male MAO A-deficient humans and mice exhibit an extreme predisposition to aggressive outbursts in response to stress. As NMDARs regulate the emotional reactivity to social and environmental stimuli, we hypothesized their involvement in the modulation of aggression mediated by MAO A. In comparison with WT male mice, MAO A KO counterparts exhibited increases in 5-HT and NE levels across all brain regions, but no difference in glutamate concentrations and NMDAR binding. Notably, the prefrontal cortex (PFC) of MAO A KO mice exhibited higher expression of NR2A and NR2B, as well as lower levels of glycosylated NR1 subunits. In line with these changes, the current amplitude and decay time of NMDARs in PFC was significantly reduced. Furthermore, the currents of these receptors were hypersensitive to the action of the antagonists of the NMDAR complex (dizocilpine), as well as NR2A (PEAQX) and NR2B (Ro 25-6981) subunits. Notably, systemic administration of these agents selectively countered the enhanced aggression in MAO A KO mice, at doses that did not inherently affect motor activity. Our findings suggest that the role of MAO A in pathological aggression may be mediated by changes in NMDAR subunit composition in the PFC, and point to a critical function of this receptor in the molecular bases of antisocial personality.

Topics: Aggression; Animals; Antisocial Personality Disorder; Autoradiography; Binding Sites; Blotting, Western; Corpus Striatum; Dizocilpine Maleate; Electrophysiological Phenomena; Excitatory Amino Acid Antagonists; Glutamic Acid; Hippocampus; Male; Mice; Mice, Knockout; Monoamine Oxidase; Motor Activity; Norepinephrine; Patch-Clamp Techniques; Phenols; Piperidines; Prosencephalon; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Serotonin

Glutamate-induced delayed calcium dysregulation (DCD) is causally linked to excitotoxic neuronal death. The mechanisms of DCD are not completely understood, but it has been proposed that the excessive influx of external Ca(2+) is essential for DCD. The NMDA-subtype of glutamate receptor (NMDAR) and the plasmalemmal Na(+)/Ca(2+) exchanger operating in the reverse mode (NCX(rev)) have been implicated in DCD. In experiments with "younger" neurons, 6-8 days in vitro (6-8 DIV), in which the NR2A-containing NMDAR expression is low, ifenprodil, an inhibitor of NR2B-containing NMDAR, completely prevented DCD whereas PEAQX, another NMDAR antagonist that preferentially interacts with NR2A-NMDAR, was without effect. With "older" neurons (13-16 DIV), in which NR2A- and NR2B-NMDARs are expressed to a greater extent, both ifenprodil and PEAQX applied separately failed to prevent DCD. However, combined application of ifenprodil and PEAQX completely averted DCD. Ifenprodil and ifenprodil-like NR2B-NMDAR antagonists Ro 25-6981 and Co 101244 but not PEAQX or AP-5 inhibited gramicidin- and Na(+)/NMDG-replacement-induced increases in cytosolic Ca(2+) mediated predominantly by NCX(rev). This suggests that ifenprodil, Ro 25-6981, and Co 101244 inhibit NCX(rev). The ability of ifenprodil to inhibit NCX(rev) correlates with its efficacy in preventing DCD and emphasizes an important role of NCX(rev) in DCD. Overall our data suggest that both NR2A- and NR2B-NMDARs are involved in DCD in "older" neurons, and it is necessary to inhibit both NMDARs and NCX(rev) to prevent glutamate-induced DCD.

Topics: Animals; Blotting, Western; Calcium Signaling; Cell Survival; Cells, Cultured; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Hippocampus; Microscopy, Fluorescence; N-Methylaspartate; Neurons; Patch-Clamp Techniques; Phenols; Piperidines; Quinoxalines; Rats; Receptors, N-Methyl-D-Aspartate; Sodium-Calcium Exchanger

Natural rewards and addictive drugs are believed to exert their reinforcing actions by influencing synaptic plasticity in reward-related brain regions such as the nucleus accumbens (NAc). Long-lasting changes in the efficacy of excitatory synaptic transmission in the NAc are critically dependent on efficient interactions between the dopaminergic and the glutamatergic neurotransmitter systems. Potential targets to the actions of dopamine and of addictive drugs include the GluN2 subunits that compose the N-Methyl-D-Aspartate (NMDA) type of glutamate receptors. However, the ability of dopamine to induce synaptic plasticity by modulating specific subunits of the NMDA receptor has not been examined. The present study shows that in the mouse NAc, dopamine produces a long-lasting depression of NMDA responses which occludes long-term depression (LTD) induced by high frequency stimulation (HFS) of glutamatergic fibers. LTD induced by dopamine or by HFS does not involve a change in the subunit composition of NMDA receptors. Although GluN2B contributes to synaptic responses in the NAc and is affected by dopamine, this subunit might not be a direct target to the actions of dopamine. The results, however, identify a critical role for GluN2A in dopamine-induced and HFS-induced synaptic plasticity. This study suggests a possible mechanism of action for dopamine in the regulation of reward-related behaviors.

Topics: Animals; Dopamine; Electric Stimulation; Excitatory Postsynaptic Potentials; In Vitro Techniques; Long-Term Synaptic Depression; Male; Mice; Mice, Inbred C57BL; Nucleus Accumbens; Patch-Clamp Techniques; Phenols; Piperidines; Quinoxalines; Receptors, N-Methyl-D-Aspartate; Synaptic Transmission

Although NMDA receptor (NMDAR)-dependent long-term potentiation (LTP) and long-term depression (LTD) of glutamatergic transmission are candidate mechanisms for long-term spatial memory, the precise contributions of LTP and LTD remain poorly understood. Here, we report that LTP and LTD in the hippocampal CA1 region of freely moving adult rats were prevented by NMDAR 2A (GluN2A) and 2B subunit (GluN2B) preferential antagonists, respectively. These results strongly suggest that NMDAR subtype preferential antagonists are appropriate tools to probe the roles of LTP and LTD in spatial memory. Using a Morris water maze task, the LTP-blocking GluN2A antagonist had no significant effect on any aspect of performance, whereas the LTD-blocking GluN2B antagonist impaired spatial memory consolidation. Moreover, similar spatial memory deficits were induced by inhibiting the expression of LTD with intrahippocampal infusion of a short peptide that specifically interferes with AMPA receptor endocytosis. Taken together, our findings support a functional requirement of hippocampal CA1 LTD in the consolidation of long-term spatial memory.

Topics: Animals; CA1 Region, Hippocampal; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Hippocampus; Long-Term Potentiation; Long-Term Synaptic Depression; Male; Maze Learning; Memory; Phenols; Piperazines; Piperidines; Protein Subunits; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spatial Behavior

NMDA receptors bidirectionally modulate extracellular signal-regulated kinase (ERK) through the coupling of synaptic NMDA receptors to an ERK activation pathway that is opposed by a dominant ERK shutoff pathway thought to be coupled to extrasynaptic NMDA receptors. In the present study, synaptic NMDA receptor activation of ERK in rat cortical cultures was partially inhibited by the highly selective NR2B antagonist Ro25-6981 (Ro) and the less selective NR2A antagonist NVP-AAM077 (NVP). When Ro and NVP were added together, inhibition appeared additive and equal to that observed with the NMDA open-channel blocker MK-801. Consistent with a selective coupling of extrasynaptic NMDA receptors to the dominant ERK shutoff pathway, pre-block of synaptic NMDA receptors with MK-801 did not alter the inhibitory effect of bath-applied NMDA on ERK activity. Lastly, in contrast to a complete block of synaptic NMDA receptor activation of ERK by extrasynaptic NMDA receptors, activation of extrasynaptic NMDA receptors had no effect upon ERK activation by brain-derived neurotrophic factor. These results suggest that the synaptic NMDA receptor ERK activation pathway is coupled to both NR2A and NR2B containing receptors, and that the extrasynaptic NMDA receptor ERK inhibitory pathway is not a non-selective global ERK shutoff.

Topics: Animals; Blotting, Western; Brain-Derived Neurotrophic Factor; Cells, Cultured; Cerebral Cortex; Cyclic AMP Response Element-Binding Protein; Dizocilpine Maleate; Enzyme Activation; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Extracellular Signal-Regulated MAP Kinases; Extracellular Space; N-Methylaspartate; Neurons; Patch-Clamp Techniques; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction

We previously demonstrated that NMDA receptors containing the NR2A or NR2B subunits differentially regulate striatal output pathways. We now investigate whether such a differential control is altered under parkinsonian conditions and whether subunit selective antagonists have different abilities to attenuate parkinsonian-like motor deficits. Three microdialysis probes were simultaneously implanted in the dopamine-depleted striatum, globus pallidus and substantia nigra reticulata of 6-hydroxydopamine hemilesioned rats. The NR2A antagonist NVP-AAM077 perfused in the striatum reduced pallidal GABA, but not glutamate, levels whereas the NR2B antagonist Ro 25-6981 was ineffective. Neither antagonist affected striatal or nigral amino acid levels. To investigate whether these neurochemical responses were predictive of different antiparkinsonian activities, antagonists were administered systemically and motor activity evaluated in different motor tasks. Neither antagonist attenuated akinesia/bradykinesia in the bar and drag test. However, NVP-AAM077 dually modulated rotarod performance (low doses being facilitatory and higher ones inhibitory) while Ro 25-6981 monotonically improved it. Microdialysis revealed that motor facilitating doses reduced pallidal GABA levels while motor inhibiting doses increased them. We conclude that, under parkinsonian conditions, the striato-pallidal pathway is driven by striatal NR2A subunits. Motor improvement induced by NVP-AAM077 and Ro 25-6981 is accomplished by blockade of striatal NR2A and extrastriatal NR2B subunits, respectively.

Topics: Adrenergic Agents; Animals; Behavior, Animal; Corpus Striatum; Disease Models, Animal; Excitatory Amino Acid Antagonists; Globus Pallidus; Male; Microdialysis; Motor Activity; Neurons; Oxidopamine; Parkinson Disease; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley

The role of NMDA receptors in the induction of long-term potentiation (LTP) and long-term depression (LTD) is well established but which particular NR2 subunits are involved in these plasticity processes is still a matter of controversy. We have studied the effects of subtype selective NMDA receptor antagonists on LTP induced by high frequency stimulation (100 Hz for 1s) and LTD induced by low frequency stimulation (1 Hz for 15 min) in the CA1 region of hippocampal slices from 14 day old Wistar rats. Against recombinant receptors in HEK293 cells NVP-AAM077 (NVP) was approximately 14-fold selective for NR2A vs NR2B receptors, whilst Ro 25-6981 (Ro) was highly selective for NR2B receptors. On NMDA receptor-mediated EPSCs from Schaffer collaterals in CA1 neurones, NVP and Ro both reduced the amplitude but differentially affected the time constant of decay. The data are compatible with the selective effect of NVP (0.1 microM) and Ro (4 microM) on native NR2A and NBR2B receptors, respectively. NVP reduced both LTP and LTD whereas Ro reduced only LTP. Thus, LTP was reduced by 63% at 0.1 microM NVP and almost completely at 0.4 microM whereas 5 microM Ro reduced LTP by 45%. These data are consistent with a role for both NR2A and NR2B in the induction of LTP, under our experimental conditions. In comparison, LTD was unaffected by Ro (5 microM) even in the presence of a glutamate uptake inhibitor threo-beta-benzylaspartic acid (TBOA) to increase the concentration of glutamate at NR2B containing receptors. NVP (0.2-0.4 microM), however, produced a concentration dependent inhibition of LTD which was complete at 0.4 microM. The lack of effect of 0.1 microM NVP on LTD contrasts with its marked effect on LTP and raises the possibility that different NVP-sensitive NR2 subunit-containing NMDA receptors are required for LTP and LTD in this preparation.

Topics: Animals; Animals, Newborn; Cell Line, Transformed; Dose-Response Relationship, Drug; Dose-Response Relationship, Radiation; Drug Interactions; Electric Stimulation; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Hippocampus; Humans; In Vitro Techniques; Long-Term Potentiation; Long-Term Synaptic Depression; N-Methylaspartate; Patch-Clamp Techniques; Phenols; Piperidines; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Transfection

We have shown previously that when postsynaptic NMDA receptors are blocked, the frequency, but not amplitude, of spontaneous EPSCs (sEPSCs) at synapses in the entorhinal cortex is reduced by NMDA receptor antagonists, demonstrating that glutamate release is tonically facilitated by presynaptic NMDA autoreceptors. In the present study, we recorded sEPSCs using whole-cell voltage clamp in neurons in layer V in slices of the rat entorhinal cortex. Using specific antagonists for NR2A [(R)-[(S)-1-(4-bromo-phenyl)-ethylamino]-(2,3-dioxo-1,2,3,4-tetrahydroquinoxalin-5-yl)-methyl]-phosphonic acid] and NR2B [(alphaR, betaS)-alpha-(4-hydroxyphenyl)-beta-methyl-4-(phenylmethyl)-1-piperidinepropanol hydrochloride (Ro 25-6981)] subunit-containing receptors, we confirmed that in slices from juvenile rats (4-6 weeks of age), the autoreceptor is predominantly of the NR1-NR2B subtype. In older (4-6 months of age) control animals, the effect of the NR2B antagonist was less marked, suggesting a decline in autoreceptor function with development. In slices from rats (aged 4-6 months) exhibiting spontaneous recurrent seizures induced with a lithium-pilocarpine protocol, Ro 25-6981 again robustly reduced sEPSC frequency. The effect was equal to or greater than that seen in the juvenile slices and much more pronounced than that seen in the age-matched control animals. In all three groups, the NR2A antagonist was without effect on sEPSCs. These results suggest that there is a developmental decrease in NMDA autoreceptor function, which is reversed in a chronic epileptic condition. The enhanced autoreceptor function may contribute to seizure susceptibility and epileptogenesis in temporal lobe structures.

Topics: Age Factors; Animals; Autoreceptors; Chronic Disease; Entorhinal Cortex; Epilepsy, Generalized; Excitatory Amino Acid Antagonists; Glutamic Acid; Male; Membrane Potentials; Neurons; Patch-Clamp Techniques; Phenols; Pilocarpine; Piperidines; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

Excitotoxicity is generally studied in dissociated neurons, cultured hippocampal slices, or intact animals. However, the requirements of dissociated neurons or cultured slices to use prenatal or juvenile rats seriously limit the advantages of these systems, whereas the complexity of intact animals prevents detailed molecular investigations. In the present experiments, we studied developmental changes in NMDA neurotoxicity in acute hippocampal slices with lactate dehydrogenase (LDH) release in medium, propidium iodide (PI) uptake, and Nissl staining as markers of cell damage. Calpain-mediated spectrin degradation was used to test calpain involvement in NMDA neurotoxicity. NMDA treatment produced increased LDH release, PI uptake, and spectrin degradation in slices from juvenile rats but not adult rats. NMDA-induced changes in slices from young rats were blocked completely by the NMDA receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (MK-801) and by the antagonists of NR2B receptor ifenprodil and R-(R, S)-alpha-(4-hydroxyphenyl)-beta-methyl-4-(phenylmethyl)-1-piperidine propranol and were partly blocked by calpain inhibitor III but were not affected by the NR2A-specific antagonist [(R)-[(S)-1-(4-bromo-phenyl)-ethylamino]-(2,3-dioxo-1,2,3,4-tetrahydroquinoxalin-5-yl)-methyl]-phosphonic acid. NMDA-induced changes in Nissl staining were also different in slices from young and adult rats and blocked by NR2B but not NR2A antagonists. In contrast to NMDA treatment, oxygen/glucose deprivation (OGD) induced neurotoxicity in slices from both young and adult rats, although OGD-induced toxicity was attenuated by MK-801 only in slices from young rats. Our results are consistent with the idea that NMDA-mediated toxicity is caused by activation of NR2B- but not NR2A-containing NMDA receptors leading to calpain activation and that developmental changes in NMDA toxicity reflect developmental changes in NMDA receptor subunit composition.

Topics: Age Factors; Animals; Biomarkers; Calpain; Dipeptides; Disks Large Homolog 4 Protein; Dizocilpine Maleate; Enzyme Activation; Excitatory Amino Acid Antagonists; Fluorescent Dyes; Gene Expression Regulation, Developmental; Glucose; Hippocampus; In Vitro Techniques; Intracellular Signaling Peptides and Proteins; L-Lactate Dehydrogenase; Membrane Proteins; N-Methylaspartate; Neurons; Oxygen; Phenols; Piperidines; Propidium; Protein Subunits; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spectrin; Valine

It has recently been proposed that activation of the NR2A subunit results in Long-term potentiation (LTP) induction, whereas activation of the NR2B subunit results in long-term depression (LTD) induction. The present study undertakes to replicate these findings in vivo to determine if a role for specific subunits in synaptic plasticity can be shown in the intact brain. Field recordings were made from the CA1 subfield of the hippocampus using Schaffer collateral stimulation in anesthetized male Sprague-Dawley rats. Antagonists of the N-methyl-D-aspartate receptors NR2A and NR2B subunits were administered by either intraperitoneal (i.p.) or intrahippocampal (i.h.) injections to assess their involvement in LTP (100 Hz stimuli) and LTD (200 Paired-burst stimuli). i.h. injection of Ro25-6981 (100 microM) significantly attenuated hippocampal LTP expression and completely blocked LTD expression. When administered i.p., Ro25-6981 (6 mg/kg) again blocked LTD, but did not significantly diminish the expression of LTP. When NVP-AAM077 was administered i.h. (80 microM) both LTP and LTD were completely abolished. The administration of this compound i.p. (1.2 mg/kg) also significantly attenuated LTP, but did not affect LTD. These data suggest that both NR2A and NR2B subunits can play roles in LTP and LTD in the hippocampus in vivo.

Topics: Analysis of Variance; Animals; Dose-Response Relationship, Drug; Drug Administration Routes; Electric Stimulation; Excitatory Amino Acid Antagonists; Hippocampus; Male; Neuronal Plasticity; Neurons; Phenols; Piperidines; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Synapses

Glutamate receptors responding to N-methyl-D: -aspartate (NMDA) are involved in neural development, excitotoxicity and neuronal plasticity. Each receptor includes at least two NR2 subunits. Here, we have examined the effects of selective antagonists of NR2A and NR2B subunits (NVP-AAM07 and Ro25-6981 respectively) on the effects of NMDA in the CA1 field of rat hippocampal slices. We have observed that Ro25-6981 potentiates, rather than blocks, the effects of NMD on field EPSPs and paired-pulse interactions (indicators of presynaptic effects) and on postsynaptic depolarisation in hippocampal slices. The NR2A subunit antagonist NVP-AAM077 blocks the effects of NMDA alone, or after potentiation by Ro25-6981. The potentiation of NMDA by Ro25-6981 was not prevented by staurosporine (protein kinase inhibitor), okadaic acid (an inhibitor of serine/threonine protein phosphatases) or anisomycin (protein synthesis inhibitor), but was prevented by cyclosporin A, which inhibits Ca2+/calmodulin-dependent phosphatase 2B [calcineurin]. NMDA-dependent long-term potentiation (LTP) induced by electrical stimulation was not prevented by Ro25-6981 but was prevented by selective blockade of the NR2A subunit. The results suggest that, at both presynaptic and postsynaptic sites in the rat hippocampus, NR2B-subunit-containing receptors limit NMDA receptor function by inhibitory restraint over NR2A-subunit-containing receptors, via calcineurin activation, and that LTP induction critically involves primarily receptors containing the NR2A subunit. Endogenous factors or drugs that modify this NR2B/NR2A interaction could have a major influence on synaptic transmission and plasticity in the brain.

Topics: Animals; Calcineurin; Calcineurin Inhibitors; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Hippocampus; Long-Term Potentiation; Male; Neural Inhibition; Organ Culture Techniques; Phenols; Piperidines; Protein Synthesis Inhibitors; Quinoxalines; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Synapses; Synaptic Transmission

Cortical plasticity is thought to be important for the establishment, consolidation, and retrieval of permanent memory. Hippocampal long-term potentiation (LTP), a cellular mechanism of learning and memory, requires the activation of glutamate N-methyl-D-aspartate (NMDA) receptors. In particular, it has been suggested that NR2A-containing NMDA receptors are involved in LTP induction, whereas NR2B-containing receptors are involved in LTD induction in the hippocampus. However, LTP in the prefrontal cortex is less well characterized than in the hippocampus. Here we report that the activation of the NR2B and NR2A subunits of the NMDA receptor is critical for the induction of cingulate LTP, regardless of the induction protocol. Furthermore, pharmacological or genetic blockade of the NR2B subunit in the cingulate cortex impaired the formation of early contextual fear memory. Our results demonstrate that the NR2B subunit of the NMDA receptor in the prefrontal cortex is critically involved in both LTP and contextual memory.

Topics: Animals; Behavior, Animal; Blotting, Western; Dose-Response Relationship, Drug; Drug Interactions; Egtazic Acid; Electric Stimulation; Electroporation; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Fear; Gene Expression Regulation; Hippocampus; In Vitro Techniques; Long-Term Potentiation; Male; Memory; Mice; Mice, Inbred C57BL; Patch-Clamp Techniques; Phenols; Piperidines; Prefrontal Cortex; Protein Subunits; Pyramidal Cells; Quinoxalines; Receptors, N-Methyl-D-Aspartate; RNA, Small Interfering

It is widely believed that long-term depression (LTD) and its counterpart, long-term potentiation (LTP), involve mechanisms that are crucial for learning and memory. However, LTD is difficult to induce in adult cortex for reasons that are not known. Here we show that LTD can be readily induced in adult cortex by the activation of NMDA receptors (NMDARs), after inhibition of glutamate uptake. Interestingly there is no need to activate synaptic NMDARs to induce this LTD, suggesting that LTD is triggered primarily by extrasynaptic NMDA receptors. We also find that de novo LTD requires the activation of NR2B-containing NMDAR, whereas LTP requires activation of NR2A-containing NMDARs. Surprisingly another form of LTD, depotentiation, requires activation of NR2A-containing NMDARs. Therefore, NMDARs with different synaptic locations and subunit compositions are involved in various forms of synaptic plasticity in adult cortex.

Topics: 2-Amino-5-phosphonovalerate; Animals; Aspartic Acid; Cerebral Cortex; Dicarboxylic Acids; Dizocilpine Maleate; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Glutamic Acid; Long-Term Potentiation; Long-Term Synaptic Depression; N-Methylaspartate; Neurons; Neurotransmitter Uptake Inhibitors; Phenols; Picrotoxin; Piperidines; Protein Subunits; Pyrrolidines; Quinoxalines; Rats; Receptors, Metabotropic Glutamate; Receptors, N-Methyl-D-Aspartate