Non-competitive Antagonist MK-801 Research Articles

Impaired Enzymatic Antioxidant Defense in Erythrocytes of Rats with Ammonia-Induced Encephalopathy: Role of NMDA Receptors.

Hepatic encephalopathy (HE), a neuropsychiatric disorder developing in patients with severe hepatic dysfunction, has been known for more than a century. However, pathogenetic mechanisms of cerebral dysfunction associated with liver disease are still poorly understood. There is a consensus that the primary cause of HE is accumulation of ammonia in the brain as a result of impaired liver detoxification capacity or the portosystemic shunt. Current evidence suggests that ammonia toxicity is mediated by hyperactivation of glutamate receptors, mainly N-methyl-D-aspartate receptors (NMDARs), and affects brain aerobic metabolism, which provides energy for multiple specific functions and neuronal viability. Recent reports on the presence of functional NMDARs in erythrocytes and the data on the deviations of blood parameters from their normal ranges indicate impaired hemodynamics and reduced oxygen-carrying capacity of erythrocytes in most patients with HE, thus suggesting a relationship between erythrocyte damage and cerebral dysfunction. In order to understand how hyperammonemia (HA)-induced disturbances in the energy metabolism in the brain (which needs a constant supply of large amounts of oxygen in the blood) lead to encephalopathy, it is necessary to reveal ammonia-induced impairments in the energy metabolism and antioxidant defense system of erythrocytes and to explore a potential role of ammonia in reduced brain oxygenation. To identify the said missing link, the activities of antioxidant enzymes and concentrations of reduced glutathione (GSH), oxidized glutathione (GSSG), and H2O2 were measured in the erythrocytes of rats with HA that were injected with the noncompetitive NMDAR antagonist MK-801. We found that in rats with HA, ammonia was accumulated in erythrocytes (cells lacking ammonia removal enzymes), which made them more susceptible to the prooxidant environment created during oxidative stress. This effect was completely or partially inhibited by MK-801. The data obtained might help to identify the risk factors in cognitive disorders and facilitate prediction of unfavorable outcomes of hypoperfusion in patients with a blood elevated ammonia concentration.

A human relevant mixture of persistent organic pollutants (POPs) and perfluorooctane sulfonic acid (PFOS) differentially affect glutamate induced excitotoxic responses in chicken cerebellum granule neurons (CGNs) in vitro

Primary cultures of cerebellar granule neurons (CGNs) derived from chicken embryos were used to explore the effects on developmental neurotoxicity by a complex defined mixture of persistent organic pollutants (POPs). Its chemical composition and concentrations were based on blood levels in the Norwegian/Scandinavian population. Perfluorooctane sulfonic acid (PFOS) alone, its most abundant compound was also evaluated. Different stages of CGNs maturation, between day in vitro (DIV) 1, 3, and 5 were exposed to the POP mixture, or PFOS alone. Their combination with glutamate, an excitatory endogenous neurotransmitter important in neurodevelopment, also known to cause excitotoxicity was evaluated. Outcomes with the mixture at 500x blood levels were compared to PFOS at its corresponding concentration of 20 μM. The POP mixture reduced tetrazolium salt (MTT) conversion at earlier stages of maturation, compared to PFOS alone. Glutamate-induced excitotoxicity was enhanced above the level of that induced by glutamate alone, especially in mature CGNs at DIV5. Glutathione (GSH) concentrations seemed to set the level of sensitivity for the toxic insults from exposures to the pollutants. The role of N-methyl-D-aspartate receptor (NMDA-R) mediated calcium influx in pollutant exposures was investigated using the non-competitive and competitive receptor antagonists MK-801 and CGP 39551. Observations indicate a calcium-independent, but still NMDA-R dependent mechanism in the absence of glutamate, and a calcium- and NMDA-R dependent one in the presence of glutamate. The outcomes for the POP mixture cannot be explained by PFOS alone, indicating that other chemicals in the mixture contribute its overall effect.

Clozapine Improves Behavioral and Biochemical Outcomes in a MK-801-Induced Mouse Model of Schizophrenia.

Glutamatergic N-methyl-D-aspartate (NMDA) receptors have critical roles in several neurological and psychiatric diseases. Dizocilpine (MK-801) is a ligand at phencyclidine recognition sites that is associated with NMDA receptor-coupled cation channels, where it acts as a potent noncompetitive antagonist of central glutamate receptors. In this study, we investigate the effect of clozapine on MK-801-induced neurochemical and neurobehavioral alterations in the prefrontal cortex of mice. Acute administration of NMDA noncompetitive antagonist MK-801 impairs motor coordination, grip strength, and locomotor activity. Clozapine is the only medication that is indicated for treating refractory schizophrenia, due to its superior efficacy among all antipsychotic agents; however, its mechanism is not well understood. To understand its mechanism, we investigated the effects of clozapine on motor coordination, locomotor activity, and grip strength in mice against the NMDA receptor antagonist MK-801. MK-801 induced elevations in acetylcholinesterase (AChE) activity, monoamine oxidase (MAO) activity, and c-fos expression. The administration of clozapine inhibited the effects caused by MK-801 (0.2 mg/kg body weight). Motor coordination and grip strength paradigms that had been altered by MK-801 were restored by clozapine. Moreover, clozapine also ameliorated MK-801-induced elevation in AChE and MAO activity. Our immunostaining results demonstrated that clozapine treatment reduced overexpression of the neuronal activity marker c-fos in cortices of the brain. Results of the current study determine that clozapine ameliorated cognition in MK-801-treated mice via cholinergic and neural mechanisms. These findings show that clozapine possesses the potential to augment cognition in diseases such as schizophrenia.

T-lymphocytes response persists following Plasmodium berghei strain Anka infection resolution and may contribute to later experimental cerebral malaria outcomes

Several studies have proposed cerebral malaria (CM) as a CD4+ and CD8+ T lymphocyte-mediated disease. However, there are no data regarding the recruitment and/or persistence of these cells in the CNS following the phase of infection resolution. Glutamate-mediate excitotoxicity has also been implicated in CM. Blockade of glutamate NMDA receptors by its noncompetitive antagonist MK801 modulates cytokine and neurotrophic factors expression preventing cognitive and depressive-like behavior in experimental CM. Herein, we aim to investigate the role of T lymphocytes in later outcomes in CM, and whether the protective role of MK801 is associated with T lymphocytes response.

NMDA Receptor Antagonist MK801 Protects Against 1-Bromopropane-Induced Cognitive Dysfunction.

Occupational exposure to 1-bromopropane (1-BP) induces learning and memory deficits. However, no therapeutic strategies are currently available. Accumulating evidence has suggested that N-methyl-D-aspartate receptors (NMDARs) and neuroinflammation are involved in the cognitive impairments in neurodegenerative diseases. In this study we aimed to investigate whether the noncompetitive NMDAR antagonist MK801 protects against 1-BP-induced cognitive dysfunction. Male Wistar rats were administered with MK801 (0.1mg/kg) prior to 1-BP intoxication (800mg/kg). Their cognitive performance was evaluated by the Morris water maze test. The brains of rats were dissected for biochemical, neuropathological, and immunological analyses. We found that the spatial learning and memory were significantly impaired in the 1-BP group, and this was associated with neurodegeneration in both the hippocampus (especially CA1 and CA3) and cortex. Besides, the protein levels of phosphorylated NMDARs were increased after 1-BP exposure. MK801 ameliorated the 1-BP-induced cognitive impairments and degeneration of neurons in the hippocampus and cortex. Mechanistically, MK801 abrogated the 1-BP-induced disruption of excitatory and inhibitory amino-acid balance and NMDAR abnormalities. Subsequently, MK801 inhibited the microglial activation and release of pro-inflammatory cytokines in 1-BP-treated rats. Our findings, for the first time, revealed that MK801 protected against 1-BP-induced cognitive dysfunction by ameliorating NMDAR function and blocking microglial activation, which might provide a potential target for the treatment of 1-BP poisoning.

Elucidating the mechanisms of fear extinction in developing animals: a special case of NMDA receptor-independent extinction in adolescent rats.

NMDA receptors (NMDARs) are considered critical for the consolidation of extinction but recent work challenges this assumption. Namely, NMDARs are not required for extinction retention in infant rats as well as when extinction training occurs for a second time (i.e., reextinction) in adult rats. In this study, a possible third instance of NMDAR-independent extinction was tested. Although adolescents typically exhibit impaired extinction retention, rats that are conditioned as juveniles and then given extinction training as adolescents (JuvCond-AdolesExt) have good extinction retention. Unexpectedly, this good extinction retention is not associated with an up-regulation of a synaptic plasticity marker in the medial prefrontal cortex, a region implicated in extinction consolidation. In the current study, rats received either the noncompetitive NMDAR antagonist MK801 (0.1 mg/kg, s.c.) or saline before extinction training. In several experiments, rats conditioned and extinguished as juveniles, adolescents, or adults exhibited impaired extinction retention after MK801 compared to saline, but this effect was not observed in JuvCond-AdolesExt rats. Further experiments ruled out several alternative explanations for why NMDAR antagonism did not affect extinction retention in adolescents extinguishing fear learned as a juvenile. These results illustrate yet another circumstance in which NMDARs are not required for successful extinction retention and highlight the complexity of fear inhibition across development.

Buspirone Counteracts MK-801-Induced Schizophrenia-Like Phenotypes through Dopamine D3 Receptor Blockade.

Background: Several efforts have been made to develop effective antipsychotic drugs. Currently, available antipsychotics are effective on positive symptoms, less on negative symptoms, but not on cognitive impairment, a clinically relevant dimension of schizophrenia. Drug repurposing offers great advantages over the long-lasting, risky and expensive, de novo drug discovery strategy. To our knowledge, the possible antipsychotic properties of buspirone, an azapirone anxiolytic drug marketed in 1986 as serotonin 5-HT1A receptor (5-HT1AR) partial agonist, have not been extensively investigated despite its intriguing pharmacodynamic profile, which includes dopamine D3 (D3R) and D4 receptor (D4R) antagonist activity. Multiple lines of evidence point to D3R as a valid therapeutic target for the treatment of several neuropsychiatric disorders including schizophrenia. In the present study, we tested the hypothesis that buspirone, behaving as dopamine D3R antagonist, may have antipsychotic-like activity.Materials and Methods: Effects of acute administration of buspirone was assessed on a wide-range of schizophrenia-relevant abnormalities induced by a single administration of the non-competitive NMDAR antagonist MK-801, in both wild-type mice (WT) and D3R-null mutant mice (D3R-/-).Results: Buspirone (3 mg⋅kg-1, i.p.) was devoid of cataleptogenic activity in itself, but resulted effective in counteracting disruption of prepulse inhibition (PPI), hyperlocomotion and deficit of temporal order recognition memory (TOR) induced by MK-801 (0.1 mg⋅kg-1, i.p.) in WT mice. Conversely, in D3R-/- mice, buspirone was ineffective in preventing MK-801-induced TOR deficit and it was only partially effective in blocking MK-801-stimulated hyperlocomotion.Conclusion: Taken together, these results indicate, for the first time, that buspirone, might be a potential therapeutic medication for the treatment of schizophrenia. In particular, buspirone, through its D3R antagonist activity, may be a useful tool for improving the treatment of cognitive deficits in schizophrenia that still represents an unmet need of this disease.

In vitro and invivo effects of a novel dimeric inhibitor of PSD-95 on excitotoxicity and functional recovery after experimental traumatic brain injury.

PSD-95 inhibitors have been shown to be neuroprotective in stroke, but have only to a very limited extent been evaluated in the treatment of traumatic brain injury (TBI) that has pathophysiological mechanisms in common with stroke. The aims of the current study were to assess the effects of a novel dimeric inhibitor of PSD-95, UCCB01-147, on histopathology and long-term cognitive outcome after controlled cortical impact (CCI) in rats. As excitotoxic cell death is thought to be a prominent part of the pathophysiology of TBI, we also investigated the neuroprotective effects of UCCB01-147 and related compounds on NMDA-induced cell death in cultured cortical neurons. Anesthetized rats were given a CCI or sham injury, and were randomized to receive an injection of either UCCB01-147 (10mg/kg), the non-competitive NMDAR-receptor antagonist MK-801 (1mg/kg) or saline immediately after injury. At 2 and 4weeks post-trauma, spatial learning and memory were assessed in a water maze, and at 3months, brains were removed for estimation of lesion volumes. Overall, neither treatment with UCCB01-147 nor MK-801 resulted in significant improvements of cognition and histopathology after CCI. Although MK-801 provided robust neuroprotection against NMDA-induced toxicity in cultured cortical neurons, UCCB01-147 failed to reduce cell death and became neurotoxic at high doses. The data suggest potential differential effects of PSD-95 inhibition in stroke and TBI that should be investigated further in future studies taking important experimental factors such as timing of treatment, dosage, and anesthesia into consideration.

Knockout of NMDA-receptors from parvalbumin interneurons sensitizes to schizophrenia-related deficits induced by MK-801.

It has been suggested that a functional deficit in NMDA-receptors (NMDARs) on parvalbumin (PV)-positive interneurons (PV-NMDARs) is central to the pathophysiology of schizophrenia. Supportive evidence come from examination of genetically modified mice where the obligatory NMDAR-subunit GluN1 (also known as NR1) has been deleted from PV interneurons by Cre-mediated knockout of the corresponding gene Grin1 (Grin1ΔPV mice). Notably, such PV-specific GluN1 ablation has been reported to blunt the induction of hyperlocomotion (a surrogate for psychosis) by pharmacological NMDAR blockade with the non-competitive antagonist MK-801. This suggests PV-NMDARs as the site of the psychosis-inducing action of MK-801. In contrast to this hypothesis, we show here that Grin1ΔPV mice are not protected against the effects of MK-801, but are in fact sensitized to many of them. Compared with control animals, Grin1ΔPVmice injected with MK-801 show increased stereotypy and pronounced catalepsy, which confound the locomotor readout. Furthermore, in Grin1ΔPVmice, MK-801 induced medial-prefrontal delta (4 Hz) oscillations, and impaired performance on tests of motor coordination, working memory and sucrose preference, even at lower doses than in wild-type controls. We also found that untreated Grin1ΔPVmice are largely normal across a wide range of cognitive functions, including attention, cognitive flexibility and various forms of short-term memory. Taken together these results argue against PV-specific NMDAR hypofunction as a key starting point of schizophrenia pathophysiology, but support a model where NMDAR hypofunction in multiple cell types contribute to the disease.

1-Deoxysphingolipid-induced neurotoxicity involves N-methyl-d-aspartate receptor signaling

1-Deoxysphingolipids (1-deoxySL) are atypical and neurotoxic sphingolipids formed by alternate substrate usage of the enzyme serine-palmitoyltransferase. Pathologically increased 1-deoxySL formation causes hereditary sensory and autosomal neuropathy type 1 (HSAN1) - a progressive peripheral axonopathy. However, the underlying molecular mechanisms by which 1-deoxySL acts are unknown. Herein we studied the effect of 1-deoxysphinganine (1-deoxySA) and its canonical counterpart sphinganine (SA) in aged cultured neurons comparing their outcome on cell survival and cytoskeleton integrity. 1-deoxySA caused rapid neuronal cytoskeleton disruption and modulated important cytoskeletal regulatory and associated components including Rac1, Ezrin and insulin receptor substrate 53. We show that 1-deoxySA is internalized and metabolized downstream to 1-deoxydihydroceramide since inhibition of ceramide synthase protected neurons from 1-deoxySA-mediated cell death. In addition, 1-deoxySA reduced protein levels of N-methyl-d-aspartate receptor (NMDAR) subunit GluN2B, the postsynaptic density protein 95 and induced cleavage of p35 to p25. Notably, blocking NMDAR activation by MK-801 or memantine significantly prevented 1-deoxySA neurotoxicity. Functional studies of differentiating primary neurons via the patch-clamp technique demonstrated that 1-deoxySA irreversibly depolarizes the neuronal membrane potential in an age-dependent manner. Notably, only neuronal cells that displayed functional NMDAR- and NMDA-induced whole-cell currents responded to 1-deoxySA treatment. Furthermore, pre-exposure to the non-competitive antagonist MK-801 blocked the current response of NMDA and glycine, as well as 1-deoxySA. We conclude that 1-deoxySA-induced neurotoxicity compromises cytoskeletal stability and targets NMDAR signaling in an age-dependent manner. Thus stabilization of cytoskeletal structures and/or inhibition of glutamate receptors could be a potential therapeutic approach to prevent 1-deoxySA-induced neurodegeneration.

NMDA receptors promote neurogenesis in the neonatal rat subventricular zone following hypoxic‑ischemic injury.

Evidence suggests the involvement of N-methyl-D-aspartate receptors (NMDAR) in the regulation of neurogenesis. Functional properties of NMDAR are strongly influenced by the type of NR2 subunits in the receptor complex. NR2A- and NR2B-containing receptors are expressed in neonatal fore-brain regions, such as the subventricular zone (SVZ). The aim of the present study was to examine the effect of the protein expression of hypoxic-ischemic injury NMDAR subunits 2A and 2B in the SVZ of neonatal rats. Expression of these and other proteins of interest was performed using immunohistochemistry. The results showed that NR2A expression was decreased at 6 h after hypoxic-ischemic injury. By contrast, a significant increase in NR2B expression was observed at 24 h after hypoxic-ischemic injury, induced by the clamping of the right common carotid artery. The functional effect of NMDAR subunits on neurogenesis was also examined by quantifying Nestin and doublecortin (DCX), the microtubule-associated protein expressed only in immature neurons. In addition, the effects of selective non-competitive NMDAR antagonist MK-801 (0.5 mg/kg), NR2B antagonist Ro25-6981 (5 mg/kg), and NR2A antagonist NVP-AAM077 (5 mg/kg) administered 30 min prior to the hypoxic-ischemic injury were examined. The number of Nestin- and DCX-positive cells increased significantly 48 h after hypoxic-ischemic injury, which was reverted by the MK-801 and Ro25-6981 antagonists. Notably, NVP-AAM077 had no significant effect on the expression of Nestin and DCX. In conclusion, the results of the present study demonstrate that hypoxia-ischemia inhibited the expression of NR2A, but promoted the expression of NR2B. Furthermore, NMDAR promoted neurogenesis in the SVZ of neonatal brains.

Strain-dependent effects on acquisition and reversal of visual and spatial tasks in a rat touchscreen battery of cognition

AimThe use of touch-screen equipped operant boxes is an increasingly popular method for modeling human cognition in the rodent. A concern of this approach is that the dependence upon vision may limit the strains of rats that can be tested in the apparatus. This is of particular concern because of the increased availability of genetically modified rats that are disproportionately on an albino background and may have compromised vision. Here we test pigmented and albino strains of rats on three touch-screen tasks of learning and memory that may require different levels of visual ability. In tests where albino animals have similar levels of performance as the pigmented rats we also tested common pharmacological models of cognitive impairment to determine the generalizability of these challenges across strains. By doing this work we hope to determine the robustness of common models of pharmacological impairment in albino rats. MethodsWe tested four strains of rats (albino: Wistar and Sprague Dawley, pigmented: Long Evans and Lister Hooded) in three touchscreen-based tasks of cognition with differing visual requirements: visual discrimination (VD) acquisition and reversal learning, and the more spatial and less visually demanding, automated spatial search task (AST). Furthermore, we tested the effects of the muscarinic antagonist scopolamine and the non-competitive NMDAR antagonist MK801 on performance of the four strains in AST. Finally, visual acuity was also assessed via a movement detection test. ResultsThe rate of acquisition (% correct) in albino rats was significantly slower than in pigmented rats. Wistar rats were significantly slower to acquire the task, and showed differences in reversal learning when compared to the pigmented strains. Moreover, SD rats performed so badly during the acquisition phase of the VD that they failed to reach inclusion criteria (80% correct responses over 3-sessions) for the reversal phase. In contrast, no effect of strain was found in AST. Some of these differences can likely be attributed to differences in visual acuity as albino animals appeared to have reduced visual acuity when compared to the pigmented animals as previously reported in the literature. Pharmacological challenge with scopolamine or MK801 induced dissociable effects between compounds, but generally comparable impairments in all four strains. ConclusionsAlbino animals showed a clear impairment on tasks that are dependent upon intact vision, while no impairment was observed in the visually less demanding spatial task. Despite a published report to the contrary, these results demonstrate that albino strains may not be appropriate for use in touchscreen tasks that are dependent upon a visual discrimination. Furthermore, the spatial search task showed distinct impairment profiles as a result of treatment with either MK-801 or scopolamine. While an interaction did exist between strain and treatment, the dissociation between MK-801 and scopolamine was consistent across 3 of 4 strains. These results highlight the importance of selecting the appropriate strain for use in tasks of visual learning and memory and also demonstrate the potential robustness of pharmacological models of cognitive impairment across strains.

Selective activation of M4 muscarinic acetylcholine receptors reverses MK-801-induced behavioral impairments and enhances associative learning in rodents.

Positive allosteric modulators (PAMs) of the M4 muscarinic acetylcholine receptor (mAChR) represent a novel approach for the treatment of psychotic symptoms associated with schizophrenia and other neuropsychiatric disorders. We recently reported that the selective M4 PAM VU0152100 produced an antipsychotic drug-like profile in rodents after amphetamine challenge. Previous studies suggest that enhanced cholinergic activity may also improve cognitive function and reverse deficits observed with reduced signaling through the N-methyl-d-aspartate subtype of the glutamate receptor (NMDAR) in the central nervous system. Prior to this study, the M1 mAChR subtype was viewed as the primary candidate for these actions relative to the other mAChR subtypes. Here we describe the discovery of a novel M4 PAM, VU0467154, with enhanced in vitro potency and improved pharmacokinetic properties relative to other M4 PAMs, enabling a more extensive characterization of M4 actions in rodent models. We used VU0467154 to test the hypothesis that selective potentiation of M4 receptor signaling could ameliorate the behavioral, cognitive, and neurochemical impairments induced by the noncompetitive NMDAR antagonist MK-801. VU0467154 produced a robust dose-dependent reversal of MK-801-induced hyperlocomotion and deficits in preclinical models of associative learning and memory functions, including the touchscreen pairwise visual discrimination task in wild-type mice, but failed to reverse these stimulant-induced deficits in M4 KO mice. VU0467154 also enhanced the acquisition of both contextual and cue-mediated fear conditioning when administered alone in wild-type mice. These novel findings suggest that M4 PAMs may provide a strategy for addressing the more complex affective and cognitive disruptions associated with schizophrenia and other neuropsychiatric disorders.

Pharmacological and parametrical investigation of prepulse inhibition of startle and prepulse elicited reactions in Wistar rats

Prepulse inhibition (PPI) is the inhibition of an acoustic startle response (ASR) that is observed when a weak prepulse is presented shortly before a startling stimulus. Here we studied in Wistar rats the dependence of PPI on variations of the interstimulus interval (ISI; from 25–1020ms) after treatment with various drugs that are known to disrupt PPI. The motor response to the prepulse itself (prepulse elicited reaction, PER) was also studied. The direct dopamine receptor agonist apomorphine, the non-competitive NMDA glutamate receptor antagonist MK-801, and the cannabinoid CB1 receptor agonist WIN 55,212-2 all reduced PPI, depending on the ISI, with different effects on the PER and/or pulse alone. The serotonin 2A receptor agonist DOI tended to reduce PPI. The cannabinoid CB1 receptor antagonist AM 251 did neither affect PPI nor the responses to prepulses or startling noise pulses. Taken together this study supports the current notion of a pharmacologically complex pattern of regulation of PPI at different ISIs and suggests that the PER is a miniature ASR that does, however, not predict the level of PPI.

A peripheral neuroimmune link: glutamate agonists upregulate NMDA NR1 receptor mRNA and protein, vimentin, TNF-alpha, and RANTES in cultured human synoviocytes.

Human primary and clonal synovial cells were incubated with glutamate receptor agonists to assess their modulating influence on glutamate receptors N-methyl-d-aspartate (NMDA) NR1 and NR2 and inflammatory cytokines to determine potential for paracrine or autocrine (neurocrine) upregulation of glutamate receptors, as has been shown for bone and chondrocytes. Clonal SW982 synoviocytes constitutively express vimentin, smooth muscle actin (SMA), and NMDA NR1 and NR2. Coincubation (6 h) with glutamate agonists NMDA (5 microM), and the NMDA NR1 glycine site activator (+/-)1-aminocyclopentane-cis-1,3-dicarboxylic acid (5 muM), significantly increases cellular mRNA and protein levels of glutamate receptors, as well as increasing vimentin, SMA, tumor necrosis factor-alpha, and RANTES (regulated on activation, normal T-cell expressed and secreted), assessed qualitatively and quantitatively with nucleotide amplification, image analysis of immunocytochemical staining, fluorescein-activated cell sorting, Western blotting, and immunoassays. Human primary synovial cells harvested from patients with arthritic conditions also constitutively expressed NMDA NR1 with increases after agonist treatment. Glutamate receptor agonist-induced increases were blocked by the noncompetitive glutamate antagonist MK-801 (8 microg/ml) and NR1 blocking antibody. Coincubation with glutamate agonists and phorbol 12-myristate 13-acetate, a protein kinase C activator, significantly enhanced mean levels of TNF-alpha and RANTES in SW982 cell supernatants compared with incubation with either agent alone. Increases were diminished with protein kinase inhibitor and NR1 blocking antibody. The functional activation of glutamate receptors on human synoviocytes establishes a neurogenic cell signaling link between neurotransmitter glutamate released from nerve terminals and target cells in the joint capsule. The influence of glutamate on subsequent release of cellular proinflammatory mediators in non-neural tissue for activation of downstream immune events supports a peripheral neuroimmune link in arthritis.

Central Glucocorticoid Receptors Modulate the Expression and Function of Spinal NMDA Receptors after Peripheral Nerve Injury

Central glucocorticoid receptors (GRs) and NMDA receptors (NMDARs) have been shown to play a significant role in the mechanisms of neuropathic pain after peripheral nerve injury; however, how central GRs and NMDARs interact in this process remains unknown. Here we show that the expression and function of spinal NMDARs after peripheral nerve injury were modulated by central GRs. Chronic constriction nerve injury (CCI) in rats induced a time-dependent upregulation of NR1 and NR2 subunits of the NMDAR within the spinal cord dorsal horn ipsilateral to CCI. The upregulation of NMDARs was significantly diminished by intrathecal administration (twice daily for postoperative days 1-6) of either the GR antagonist RU38486 or an antisense oligonucleotide against GRs. Moreover, this CCI-induced expression of NMDARs was significantly attenuated in rats receiving intrathecal treatment with an interleukin-6 (IL-6) antiserum and in mice with protein kinase Cgamma (PKCgamma) knock-out. Because IL-6 and PKCgamma mediated the upregulation of central GRs after CCI as demonstrated previously, the results suggest that IL-6 and PKCgamma served as cellular mediators contributing to the GR-mediated expression of NMDARs after CCI. Functionally, nociceptive behaviors induced by NMDAR activation and CCI were reversed by a single intrathecal administration of the GR antagonist RU38486. Conversely, a single intrathecal injection with the noncompetitive NMDAR antagonist MK-801 reversed neuropathic pain behaviors exacerbated by the GR agonist dexamethasone in CCI rats. These data suggest that interactions between central GRs and NMDARs through genomic and nongenomic regulation may be an important mechanism critical to neuropathic pain behaviors in rats.

Role of magnesium ions on the regulation of NMDA receptor--a pharmacopathology of memantine

Magnesium ion blocks the ion channel of the NMDA receptor at a stable condition. The ion channel competes with the binding site of the noncompetitive antagonists phencyclidine (PCP) and MK-801, which prevent a brain impairment due to the ischemia and so on. The binding ability of these antagonists is strong, an exchange with the magnesium ion is not easy, then the side effect of the schizophrenia-like behavior is caused. Recently, memantine can be used as a therapeutic drug of the moderate-to-severe Alzheimer's disease. Memantine is the noncompetitive antagonist, too, then those development details and a difference from MK-801 were explained.

Effects of NMDA receptor antagonists on acute μ-opioid analgesia in the rat

Mixed research findings have led to a debate regarding the effect of N-methyl- d-aspartate (NMDA) receptor antagonists on opiate analgesia. NMDA antagonists have been found in various studies to enhance, to inhibit, or to have no effect on opiate analgesia. The present research used a single protocol to explore the effects of six NMDA receptor antagonists on acute morphine (3.0 mg/kg sc) and fentanyl (0.05 mg/kg sc) analgesia in adult male Sprague–Dawley rats. NMDA receptor antagonists were selected based on their abilities to block various sites on the NMDA receptor complex, including the noncompetitive antagonists MK-801 (0.1 and 0.3 mg/kg ip), dextromethorphan (10.0 and 30.0 mg/kg ip), and memantine (3.0 and 10.0 mg/kg ip), a glycine site antagonist, (+)-HA-966 (10.0 and 30.0 mg/kg ip), a competitive antagonist, LY235959 (1.0 and 3.0 mg/kg ip), and a polyamine site antagonist, ifenprodil (1.0 and 3.0 mg/kg ip). Analgesia was assessed using the tail-flick test. A single dose of each opiate was used. The low doses of the antagonists, which are known to produce significant neural and behavioral actions at NMDA receptors, had no effect on morphine or fentanyl analgesia. At the higher doses, morphine analgesia was significantly enhanced by LY235959 (3.0 mg/kg), and fentanyl analgesia was significantly enhanced by LY235959 (3.0 mg/kg), dextromethorphan (30.0 mg/kg), and (+)-HA-966 (30.0 mg/kg). Enhancement of analgesia occurred without any apparent adverse side effects. None of the NMDA antagonists affected tail-flick responses on their own, except the higher dose of LY235959 (3.0 mg/kg), which produced a mild analgesic effect. Because no consistent effects were observed, the data suggest that NMDA receptors are not involved in acute μ-opioid analgesia. The mechanisms underlying the enhancement of opiate analgesia by selected NMDA antagonists, such as LY235959, dextromethorphan, and (+)-HA-966, remain to be determined.

Regulation of transcription factor phosphorylation by metabotropic glutamate receptor-associated signaling pathways in rat striatal neurons

The group I metabotropic glutamate receptors (mGluRs) are positively coupled to phospholipase C. Through phospholipase C, group I mGluR activation increases intracellular concentrations of diacylglycerol which is known as a strong activator of protein kinase C (PKC). This study investigated the putative role of PKC in the regulation of transcription factor phosphorylation induced by group I mGluR activation in the rat striatum in vivo. We found that the group I agonist 3,5-dihydroxyphenylglycine (DHPG) injected into the dorsal striatum (caudate–putamen) increased phosphorylation of the two transcription factors, cAMP response element-binding protein (CREB) and Elk-1, and extracellular signal-regulated kinase 1/2 (ERK1/2) in the injected striatum. Inhibition of PKC with GF109203X significantly attenuated DHPG-stimulated CREB, Elk-1, and ERK1/2 phosphorylation. Activation of PKC with intracaudate injection of 12- O-tetradecanoylphorbol-13-acetate (TPA) mimicked DHPG actions in facilitating the phosphorylation of CREB, Elk-1, and ERK1/2. Blockade of N-methyl- D-aspartate (NMDA) glutamate receptors with the non-competitive antagonist MK801 or the competitive antagonist AP5 attenuated TPA-induced CREB, Elk-1, and ERK1/2 phosphorylation. Similarly, inhibition of Ca 2+/calmodulin-dependent protein kinases (CaMK) with KN62 also resulted in a significant attenuation of TPA induction of the three phosphoproteins. The data obtained from this study indicate that selective activation of PKC is needed for the group I agonist-induced CREB, Elk-1, and ERK1/2 phosphorylation in striatal neurons. Activated PKC may, at least in part, facilitate the phosphorylation of transcription factors via an NMDA/CaMK-sensitive pathway.

Sexually dimorphic effects of NMDA receptor antagonism on brain–pituitary–gonad axis development in the platyfish

The N-methyl- d-aspartate glutamate receptor (NMDAR) is found in hypothalamic nuclei involved in the regulation of reproduction in several species of mammals and fishes. NMDAR is believed to affect reproductive development and function by regulating gonadotropin releasing hormone (GnRH)-producing cells. These pathways are likely to be sexually dimorphic, as are several other neurotransmitter systems involved in reproductive function. In this report, male and female platyfish received intraperitoneal injections of 0, 5, 10, 20, 40 or 60 μg/g body wt. of the non-competitive NMDAR antagonist MK-801. Injections began at 6 weeks of age and continued thrice weekly until control animals reached puberty, as evidenced by anal fin maturation. The percent of pubescent animals was significantly affected by sex and treatment, with fewer MK-801-injected females in puberty than control females at each dose ( P<0.001), and fewer pubescent females than males at 10, 20 and 40 μg/g ( P<0.05). There were no MK-801-related effects in males. Histological analyses revealed typical immature gonads and pituitary glands in treated females, and typical mature morphology in control females and all males. Immunocytochemical distribution of the R1 subunit of the NMDAR within the brain-pituitary-gonad (BPG) axis was limited to GnRH-containing brain cells in all animals; however, NMDAR1 distribution was in an immature pattern in treated females and a mature pattern in all others. Neural concentrations of GnRH were unaffected by MK-801 treatment in both sexes. These data suggest that in the platyfish, NMDAR influence on reproductive development is sexually dimorphic and occurs at, or above, the level of GnRH-containing cells of the BPG axis.