Glutamate Receptor Antagonist MK-801 Research Articles

Bioenergetic and excitotoxic determinants of cofilactin rod formation.

Cofilactin rods (CARs), which are 1:1 aggregates of cofilin-1 and actin, lead to neurite loss in ischemic stroke and other disorders. The biochemical pathways driving CAR formation are well-established, but how these pathways are engaged under ischemic conditions is less clear. Brain ischemia produces both ATP depletion and glutamate excitotoxicity, both of which have been shown to drive CAR formation in other settings. Here, we show that CARs are formed in cultured neurons exposed to ischemia-like conditions: oxygen-glucose deprivation (OGD), glutamate, or oxidative stress. Of these conditions, only OGD produced significant ATP depletion, showing that ATP depletion is not required for CAR formation. Moreover, the OGD-induced CAR formation was blocked by the glutamate receptor antagonists MK-801 and kynurenic acid; the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitors GSK2795039 and apocynin; as well as an ROS scavenger. The findings identify a biochemical pathway leading from OGD to CAR formation in which the glutamate release induced by energy failure leads to activation of neuronal glutamate receptors, which in turn activates NADPH oxidase to generate oxidative stress and CARs.

Scopolamine and MK-801 impair recognition memory in a new spontaneous object exploration task in monkeys

The spontaneous object recognition (SOR) task is one of the most widely used behavioral protocols to assess visual memory in animals. However, only recently was it shown that nonhuman primates also perform well on this task. Here we further characterized this new monkey recognition memory test by assessing the performance of adult marmosets after an acute systemic administration of two putative amnesic agents: the competitive muscarinic acetylcholine receptor antagonist scopolamine (SCP; 0.05 mg/kg) and the noncompetitive N-methyl-d-aspartate glutamate receptor antagonist MK-801 (0.015 mg/kg). We also determined whether the acetylcholinesterase inhibitor donepezil (DNP; 0.50 mg/kg), a clinically-used cognitive enhancer, reverses memory deficits caused by either drug. The subjects had an initial 10 min sample trial where two identical neutral objects could be explored. After a 6 h retention interval, recognition was based on an exploratory preference for a new rather than familiar object during a 10 min test trial. Both SCP and MK-801 impaired the marmosets' performance on the SOR task, as both objects were explored equivalently. Co-administration of 0.50 mg/kg of DNP reversed the SCP- but not the MK-801-induced memory deficit. These results indicate that cholinergic and glutamatergic pathways mediate object recognition memory in the monkey SOR task.

An examination of the roles of glutamate and sex in latent inhibition: Relevance to the glutamate hypothesis of schizophrenia?

The present study examined the effects of the glutamate receptor antagonist MK-801, the glutamate receptor agonist N-methyl-D-aspartate (NMDA), and sexual dimorphism on latent inhibition to elucidate the glutamate hypothesis of schizophrenia. During the pre-exposure phase, 56 male and 65 female Wistar rats were intracerebroventricularly administered normal saline, MK-801 or NMDA, in the left ventricle and then exposed to a passive avoidance box (or a different context) in three trials over 3 days. Then, all of the rats were placed in the light compartment of the passive avoidance box and were allowed to enter the dark compartment, where they each received a footshock (1mA, 2s) in five trials over 5 days. Injections of the glutamate drugs NMDA and MK-801 did not affect latent inhibition. Sexual dimorphism did not occur in latent inhibition. The present data on the male rats indicated that the glutamate system did not affect latent inhibition, indicating that the glutamate system was not like the dopamine system in terms of mediating the positive symptoms of schizophrenia. The glutamate system might be involved in the negative and cognitive symptoms of schizophrenia. The results may provide information for novel treatments of the negative and cognitive symptoms of schizophrenia.

A Neuroprotective Effect of the Glutamate Receptor Antagonist MK801 on Long-Term Cognitive and Behavioral Outcomes Secondary to Experimental Cerebral Malaria.

Cerebral malaria (CM) is a life-threatening complication of Plasmodium falciparum infection, which can result in long-term cognitive and behavioral deficits despite successful anti-malarial therapy. Due to the substantial social and economic burden of CM, the development of adjuvant therapies is a scientific goal of highest priority. Apart from vascular and immune responses, changes in glutamate system have been reported in CM pathogenesis suggesting a potential therapeutic target. Based on that, we hypothesized that interventions in the glutamatergic system induced by blockage of N-methyl-D-aspartate (NMDA) receptors could attenuate experimental CM long-term cognitive and behavioral outcomes. Before the development of evident CM signs, susceptible mice infected with Plasmodium berghei ANKA (PbA) strain were initiated on treatment with dizocilpine maleate (MK801, 0.5mg/kg), a noncompetitive NMDA receptor antagonist. On day 5 post-infection, mice were treated orally with a 10-day course chloroquine (CQ, 30mg/kg). Control mice also received saline, CQ or MK801 + CQ therapy. After 10days of cessation of CQ treatment, magnetic resonance images (MRI), behavioral and immunological assays were performed. Indeed, MK801 combined with CQ prevented long-term memory impairment and depressive-like behavior following successful PbA infection resolution. In addition, MK801 also modulated the immune system by promoting a balance of TH1/TH2 response and upregulating neurotrophic factors levels in the frontal cortex and hippocampus. Moreover, hippocampus abnormalities observed by MRI were partially prevented by MK801 treatment. Our results indicate that NMDA receptor antagonists can be neuroprotective in CM and could be a valuable adjuvant strategy for the management of the long-term impairment observed in CM.

Regulatory effects of glutamate receptor antagonists on the proliferation and migration of WM451LU malignant melanoma cells and their related mechanisms

Objective To evaluate regulatory effects of glutamate receptor antagonists on the proliferation and migration of WM451LU malignant melanoma cells, and to explore their related mechanisms. Methods WM451LU cells at exponential growth phase were classified into 3 groups to be treated with the glutamate receptor antagonist MK-801 at 100 μmol/L(MK-801 group), the glutamate receptor antagonist CPCCOEt at 10 μmol/L(CPCCOEt group), or culture medium(control group). After 24-hour treatment, methyl thiazolyl tetrazolium(MTT)assay was performed to determine cell proliferation rates, scratch assay to evaluate the migration activity of cells, and Western-blot analysis to measure expression levels of proliferating cell nuclear antigen(PCNA), protein kinase Cα(PKCα)both on cell membrane and in cytoplasm, and phosphorylated mitogen-activated protein kinase(p-MAPK). Results After 24-hour treatment, cell proliferation rates were significantly decreased in the MK-801 group and CPCCOEt group compared with the control group(63% ± 3.1% and 60% ± 2.4% vs. 100% ± 1.1%, both P < 0.05). The scratch assay showed that cell-free zones in the control group gradually narrowed over time, and the scratch wound tended to close. However, the cell-free zones in the MK-801 group and CPCCOEt group narrowed more slowly compared with the control group, and were still wide after 24-hour culture with no obvious closure of the scratch. The MK-801 group and CPCCOEt group both showed significantly decreased expressions of PCNA(77.0% ± 5.4% and 72.0% ± 4.2% respectively), PKCα on the cell membrane(0.12 ± 0.02 and 0.14 ± 0.02 respectively), and p-MAPK(0.48 ± 0.03 and 0.36 ± 0.04 respectively)compared with the control group(PCNA: 100.0% ± 1.3%; PKCα: 0.38 ± 0.01; p-MAPK: 1.00 ± 0.02; all P < 0.05). Conclusion In vitro suppression of glutamate receptors can inhibit the proliferation and migration of WM451LU cells, likely through the mediation of the PKCα-MAPK signaling pathway. Key words: Melanoma, experimental; Receptors, glutamate; Cell proliferation; Protein kinase C-alpha; Mitogen-activated protein kinases

Dynamics of the Development of Amnesia Caused by Disruption of Memory Reconsolidation by Neurotransmitter Receptors Antagonists.

The dynamics of amnesia development under conditions of memory reconsolidation disruption by serotonin receptor antagonist methiothepin or NMDA glutamate receptor antagonist MK-801 was studied in snails trained in conventional food aversion. In 2 days after training, injection of methiothepin or MK-801 before reminder induced amnesia development. During repeated training in 3 days after amnesia induction, the skill was formed more rapidly than during the initial training. During repeated training in 10 days after administration of methiothepin and reminder, the dynamics of habit formation was similar to that during initial learning. At the same time, repeated training in 10 days after MK-801 administration and reminder did not result in long-term memory formation. Disruption of reconsolidation of conditioned food aversion memory by antagonists of serotonin or NMDA glutamate receptors led to the development of different types of amnesia that had similar strengthening gradient at the early stages, but differed by the possibility of memory formation during re-training at the late stage.

State-dependent alterations in sleep/wake architecture elicited by the M4 PAM VU0467154 – Relation to antipsychotic-like drug effects

Accumulating evidence indicates direct relationships between sleep abnormalities and the severity and prevalence of other symptom clusters in schizophrenia. Assessment of potential state-dependent alterations in sleep architecture and arousal relative to antipsychotic-like activity is critical for the development of novel antipsychotic drugs (APDs). Recently, we reported that VU0467154, a selective positive allosteric modulator (PAM) of the M4 muscarinic acetylcholine receptor (mAChR), exhibits robust APD-like and cognitive enhancing activity in rodents. However, the state-dependent effects of VU0467154 on sleep architecture and arousal have not been examined. Using polysomnography and quantitative electroencephalographic recordings from subcranial electrodes in rats, we evaluated the effects of VU0467154, in comparison with the atypical APD clozapine and the M1/M4-preferring mAChR agonist xanomeline. VU0467154 induced state-dependent alterations in sleep architecture and arousal including delayed Rapid Eye Movement (REM) sleep onset, increased cumulative duration of total and Non-Rapid Eye Movement (NREM) sleep, and increased arousal during waking periods. Clozapine decreased arousal during wake, increased cumulative NREM, and decreased REM sleep. In contrast, xanomeline increased time awake and arousal during wake, but reduced slow wave activity during NREM sleep. Additionally, in combination with the N-methyl-d-aspartate subtype of glutamate receptor (NMDAR) antagonist MK-801, modeling NMDAR hypofunction thought to underlie many symptoms in schizophrenia, both VU0467154 and clozapine attenuated MK-801-induced elevations in high frequency gamma power consistent with an APD-like mechanism of action. These findings suggest that selective M4 PAMs may represent a novel mechanism for treating multiple symptoms of schizophrenia, including disruptions in sleep architecture without a sedative profile.

Epilepsy and hippocampal neurodegeneration induced by glutamate decarboxylase inhibitors in awake rats

Glutamic acid decarboxylase (GAD), the enzyme responsible for GABA synthesis, requires pyridoxal phosphate (PLP) as a cofactor. Thiosemicarbazide (TSC) and γ-glutamyl-hydrazone (PLPGH) inhibit the free PLP-dependent isoform (GAD65) activity after systemic administration, leading to epilepsy in mice and in young, but not in adult rats. However, the competitive GAD inhibitor 3-mercaptopropionic acid (MPA) induces convulsions in both immature and adult rats. In the present study we tested comparatively the epileptogenic and neurotoxic effects of PLPGH, TSC and MPA, administered by microdialysis in the hippocampus of adult awake rats. Cortical EEG and motor behavior were analyzed during the next 2h, and aspartate, glutamate and GABA were measured by HPLC in the microdialysis-collected fractions. Twenty-four hours after drug administration rats were fixed for histological analysis of the hippocampus. PLPGH or TSC did not affect the motor behavior, EEG or cellular morphology, although the extracellular concentration of GABA was decreased. In contrast, MPA produced intense wet-dog shakes, EEG epileptiform discharges, a >75% reduction of extracellular GABA levels and remarkable neurodegeneration of the CA1 region, with >80% neuronal loss. The systemic administration of the NMDA glutamate receptor antagonist MK-801 30min before MPA did not prevent the MPA-induced epilepsy but significantly protected against its neurotoxic effect, reducing neuronal loss to <30%. We conclude that in adult awake rats, drugs acting on PLP availability have only a weak effect on GABA neurotransmission, whereas direct GAD inhibition produced by MPA induces hyperexcitation leading to epilepsy and hippocampal neurodegeneration. Because this degeneration was prevented by the blockade of NMDA receptors, we conclude that it is due to glutamate-mediated excitotoxicity consequent to disinhibition of the hippocampal excitatory circuits.

Tumor necrosis factor-mediated downregulation of spinal astrocytic connexin43 leads to increased glutamatergic neurotransmission and neuropathic pain in mice.

Spinal cord astrocytes are critical in the maintenance of neuropathic pain. Connexin 43 (Cx43) expressed on spinal dorsal horn astrocytes modulates synaptic neurotransmission, but its role in nociceptive transduction has yet to be fully elaborated. In mice, Cx43 is mainly expressed in astrocytes, not neurons or microglia, in the spinal dorsal horn. Hind paw mechanical hypersensitivity was observed beginning 3days after partial sciatic nerve ligation (PSNL), but a persistent downregulation of astrocytic Cx43 in ipsilateral lumbar spinal dorsal horn was not observed until 7days post-PSNL, suggesting that Cx43 downregulation mediates the maintenance and not the initiation of nerve injury-induced hypersensitivity. Downregulation of Cx43 expression by intrathecal treatment with Cx43 siRNA also induced mechanical hypersensitivity. Conversely, restoring Cx43 by an adenovirus vector expressing Cx43 (Ad-Cx43) ameliorated PSNL-induced mechanical hypersensitivity. The sensitized state following PSNL is likely maintained by dysfunctional glutamatergic neurotransmission, as Cx43 siRNA-induced mechanical hypersensitivity was attenuated with intrathecal treatment of glutamate receptor antagonists MK801 and CNQX, but not neurokinin-1 receptor antagonist CP96345 or the Ca(2+) channel subunit α2δ1 blocker gabapentin. The source of this dysfunctional glutamatergic neurotransmission is likely decreased clearance of glutamate from the synapse rather than increased glutamate release into the synapse. Astrocytic expression of glutamate transporter GLT-1, but not GLAST, and activity of glutamate transport were markedly decreased in mice intrathecally injected with Cx43-targeting siRNA but not non-targeting siRNA. Glutamate release from spinal synaptosomes prepared from mice treated with either Cx43-targeting siRNA or non-targeting siRNA was unchanged. Intrathecal injection of Ad-Cx43 in PSNL mice restored astrocytic GLT-1 expression. The cytokine tumor necrosis factor (TNF) has been implicated in the induction of central sensitization, particularly through its actions on astrocytes, in the spinal cord following peripheral injury. Intrathecal injection of TNF in naïve mice induced the downregulation of both Cx43 and GLT-1 in spinal dorsal horn, as well as hind paw mechanical hypersensitivity, as observed in PSNL mice. Conversely, intrathecal treatment of PSNL mice with the TNF inhibitor etanercept prevented not only mechanical hypersensitivity but also the downregulation of Cx43 and GLT-1 expression in astrocytes. The current findings indicate that spinal astrocytic Cx43 are essential for the maintenance of neuropathic pain following peripheral nerve injury and suggest modulation of Cx43 as a novel target for developing analgesics for neuropathic pain.

Downregulation of connexin36 in mouse spinal dorsal horn neurons leads to mechanical allodynia.

Connexin36 (Cx36), a component of neuronal gap junctions, is crucial for interneuronal communication and regulation. Gap junction dysfunction underlies neurological disorders, including chronic pain. Following a peripheral nerve injury, Cx36 expression in the ipsilateral spinal dorsal horn was markedly decreased over time, which paralleled the time course of hind paw tactile allodynia. Intrathecal (i.t.) injection of Cx36 siRNA (1 and 5 pg) significantly reduced the expression of Cx36 protein in the lumbar spinal cord, peaking 3 days after the injection, which corresponded with the onset of hind paw tactile allodynia. It is possible that some of the tactile allodynia resulting from Cx36 downregulation could be mediated through excitatory neuromodulators, such as glutamate and substance P. The Cx36 knockdown-evoked tactile allodynia was significantly attenuated by i.t. treatment with the N-methyl-D-aspartate glutamate receptor antagonist MK-801 but not the substance P receptor antagonist CP96345. Immunohistochemistry showed that Cx36 was colocalized with glycine transporter-2, a marker for inhibitory glycinergic spinal interneurons, but not with glutamate decarboxylase 67, a marker for inhibitory GABAergic spinal interneurons. The results indicate that spinal inhibition through glycinergic interneurons is reduced, leading to increased glutamatergic neurotransmission, as a result of Cx36 downregulation. The current data suggest that gap junction dysfunction underlies neuropathic pain and further suggest a novel target for the development of analgesics.

Long-term phase reorganization of conditioned food aversion memory in edible snail.

The specific features of memory reconsolidation in edible snails were studied over 30 days after learning of conditioned food aversion. Injections of a NMDA glutamate receptor antagonist MK-801 or protein synthesis inhibitor cycloheximide in combination with the conditioned food stimulus (reminder) on day 2 after learning were followed by the development of amnesia. Repeated training on day 10 after the induction of amnesia did not result in skill formation. Injections of MK-801 or cycloheximide and reminder by the 10th day after training had no effect on memory retention. Injections of MK-801 or cycloheximide and reminder by the 30th day after training were followed by the development of amnesia. In these experiments, memory was recovered after repeated training. Our results indicate that a complex phase reorganization of memory occurs over 30 days after learning. This process includes memory consolidation over the first days after training, stabilization and resistance to adverse factors after 10 days, and newly acquired ability for reconsolidation by the 30th day after training.

Kynurenic acid inhibits proliferation and migration of human glioblastoma T98G cells

BackgroundKynurenic acid (KYNA), tryptophan metabolite synthesized in the kynurenine pathway, is an endogenous antagonist of α-7 nicotinic receptor and all ionotropic glutamate receptors: N-methyl-d-aspartate (NMDA) receptor, α-amino-3-hydroxy-5-methyl-4-isoxasole propionate (AMPA) receptor and kainate receptor. The antiproliferative activity of KYNA toward colon and renal cancer cells has recently been discovered. The aim of the study was to verify whether human Glioblastoma tumors contain KYNA and if KYNA influences glioma cell proliferation and migration. MethodsKYNA content in Glioblastoma tumor samples was determined using HPLC. Proliferation of human glioblastoma T98G cells was measured by means of MTT and BrdU assays. Wound assay was used to evaluate the effect of KYNA on cancer cell migration. ResultsKYNA was detected in all tested Glioblastoma tumor samples (100.3±17.6pmol/g wet weight). In a series of experiments the antiproliferative activity of KYNA against T98G cells was revealed (IC50=1.3mM). Moreover, KYNA reversed the stimulatory effect of glutamate on glioma cell proliferation and enhanced antiproliferative effect of glutamate receptor antagonists MK801 and GYKI 52466. Next, KYNA at concentrations much lower than those needed to reduce cell proliferation elicited a prominent inhibitory effect on glioma cell motility. Moreover, co-incubation of temozolomide, a drug commonly used in antiglioblastoma therapy, with KYNA gave a superior effect than each of the substances applied alone. ConclusionsWe demonstrate the antiproliferative and antimigrative potential of KYNA against glioma cells in vitro.

The combination of glutamate receptor antagonist MK-801 with tamoxifen and its active metabolites potentiates their antiproliferative activity in mouse melanoma K1735-M2 cells

Recent reports suggest that N-methyl-d-aspartate receptor (NMDAR) blockade by MK-801 decreases tumor growth. Thus, we investigated whether other ionotropic glutamate receptor (iGluR) antagonists were also able to modulate the proliferation of melanoma cells. On the other hand, the antiestrogen tamoxifen (TAM) decreases the proliferation of melanoma cells, and is included in combined therapies for melanoma. As the efficacy of TAM is limited by its metabolism, we investigated the effects of the NMDAR antagonist MK-801 in combination with TAM and its active metabolites, 4-hydroxytamoxifen (OHTAM) and endoxifen (EDX). The NMDAR blockers MK-801 and memantine decreased mouse melanoma K1735-M2 cell proliferation. In contrast, the NMDAR competitive antagonist APV and the AMPA and kainate receptor antagonist NBQX did not affect cell proliferation, suggesting that among the iGluR antagonists only the NMDAR channel blockers inhibit melanoma cell proliferation. The combination of antiestrogens with MK-801 potentiated their individual effects on cell biomass due to diminished cell proliferation, since it decreased the cell number and DNA synthesis without increasing cell death. Importantly, TAM metabolites combined with MK-801 promoted cell cycle arrest in G1. Therefore, the data obtained suggest that the activity of MK-801 and antiestrogens in K1735-M2 cells is greatly enhanced when used in combination.

Role of glutamate receptors in the dorsal reticular nucleus in formalin-induced secondary allodynia

The role of glutamate receptors present in the medullary dorsal reticular nucleus (DRt) in the formalin test and formalin-induced secondary nociception was studied in rats. Secondary mechanical allodynia was assessed with von Frey filaments applied to the rat's hindpaw, and secondary thermal hyperalgesia was evaluated with the tail-immersion test. The selective glutamate receptor antagonists MK801 (N-methyl-D-aspartate receptor antagonist), 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) (AMPA/KA receptor antagonist) and A841720 (metabotropic glutamate 1 receptor antagonist) were injected into the DRt before or 6 days after formalin injection in the rat. In the formalin test, the three antagonists significantly reduced the number of flinches in both phases of the test. DRt microinjection of MK801 or A841720, but not of CNQX, reduced both secondary nociceptive behaviors. Moreover, pre-treatment with the three antagonists injected into the DRt prevented the development of secondary mechanical allodynia and secondary thermal hyperalgesia. Similarly, in these rats, the number of c-Fos-like immunoreactive neurons were markedly reduced in both the superficial and deep lamina of the dorsal horn. Our findings support the role of DRt as a pain facilitator in acute and chronic pain states, and suggest a key role of glutamate receptors during the development and maintenance of formalin-induced secondary allodynia.

Bacterial cytolysin during meningitis disrupts the regulation of glutamate in the brain, leading to synaptic damage.

Streptococcus pneumoniae (pneumococcal) meningitis is a common bacterial infection of the brain. The cholesterol-dependent cytolysin pneumolysin represents a key factor, determining the neuropathogenic potential of the pneumococci. Here, we demonstrate selective synaptic loss within the superficial layers of the frontal neocortex of post-mortem brain samples from individuals with pneumococcal meningitis. A similar effect was observed in mice with pneumococcal meningitis only when the bacteria expressed the pore-forming cholesterol-dependent cytolysin pneumolysin. Exposure of acute mouse brain slices to only pore-competent pneumolysin at disease-relevant, non-lytic concentrations caused permanent dendritic swelling, dendritic spine elimination and synaptic loss. The NMDA glutamate receptor antagonists MK801 and D-AP5 reduced this pathology. Pneumolysin increased glutamate levels within the mouse brain slices. In mouse astrocytes, pneumolysin initiated the release of glutamate in a calcium-dependent manner. We propose that pneumolysin plays a significant synapto- and dendritotoxic role in pneumococcal meningitis by initiating glutamate release from astrocytes, leading to subsequent glutamate-dependent synaptic damage. We outline for the first time the occurrence of synaptic pathology in pneumococcal meningitis and demonstrate that a bacterial cytolysin can dysregulate the control of glutamate in the brain, inducing excitotoxic damage.

Over-expectation generated in a complex appetitive goal-tracking task is capable of inducing memory reconsolidation

Discrepancies in an expected outcome have been demonstrated to result in modification of behaviour in both appetitive and aversive conditioning settings. In this study, we sought to establish whether overexpectation generated from compound conditioning with two previously rewarded stimuli was able to induce memory destabilisation and subsequent reconsolidation in a Pavlovian conditioned approach setting. It was shown that 4 days, but not 1 day, of overexpectation training was required to induce memory reconsolidation, and this was disrupted by application of the NMDA subtype of glutamate receptor antagonist MK-801 prior to overexpectation training, but not by MK-801 application 6 h post-training. These data provide evidence that the memories underlying Pavlovian conditioned approach do undergo reconsolidation and that such reconsolidation can be triggered by overexpectation. Therefore, the updating of appetitive conditioned stimulus and unconditioned stimulus associations underpinning conditioned responding in manners other than extinction training is likely achieved through memory reconsolidation.

Accumulation of autofluorescent storage material in brain is accelerated by ischemia in chloride channel 3 gene‐deficient mice

Autofluorescent storage material (ASM) is an aging pigment that accumulates during the normal course of senescence. Although the role of ASM has yet to be fully elucidated, ASM has been implicated in age-related neurodegeneration. In this study, we determined the level of ASM in chloride channel 3 (ClC-3) gene-deficient (KO) mice both in response to aging and following mild global ischemia. To understand the mechanism of action of the ASM, mice subjected to ischemia were treated with the cyclooxygenase (COX) inhibitor indomethacin or with the noncompetitive glutamate receptor antagonist MK-801. ClC-3 KO mice displayed age-related neurodegeneration of the neocortex as well as the hippocampus. The cortical layers in particular granular layers became thinner with aging. ASM accumulated in the brains of ClC-3 KO mice was increased seven- to 50-fold over that observed in the corresponding regions of their wild-type littermates. Young wild-type mice survived longer than age-matched ClC-3 KO mice after permanent global ischemia. However, in the case of older animals, the survival curves were similar. The ASM also increased four- to fivefold 10 days after mild global ischemia, an effect that was suppressed by treatment with indomethacin and MK-801. These results suggest that temporary ischemia might trigger a process similar to aging in the brain, mimicking the effect of age-related neurodegenerative diseases.

Common influences of non-competitive NMDA receptor antagonists on the consolidation and reconsolidation of cocaine-cue memory

Environmental stimuli or contexts previously associated with rewarding drugs contribute importantly to relapse among addicts, and research has focused on neurobiological processes maintaining those memories. Much research shows contributions of cell surface receptors and intracellular signaling pathways in maintaining associations between rewarding drugs (e.g., cocaine) and concurrent cues/contexts; these memories can be degraded at the time of their retrieval through reconsolidation interference. Much less studied is the consolidation of drug-cue memories during their acquisition. The present experiments use the cocaine-conditioned place preference (CPP) paradigm in rats to directly compare, in a consistent setting, the effects of N-methyl-D-aspartate (NMDA) glutamate receptor antagonists MK-801 and memantine on the consolidation and reconsolidation of cocaine-cue memories. For the consolidation studies, animals were systemically administered MK-801 or memantine immediately following training sessions. To investigate the effects of these NMDA receptor antagonists on the retention of previously established cocaine-cue memories, animals were systemically administered MK-801 or memantine immediately after memory retrieval. Animals given either NMDA receptor antagonist immediately following training sessions did not establish a preference for the cocaine-paired compartment. Post-retrieval administration of either NMDA receptor antagonist attenuated the animals' preference for the cocaine-paired compartment. Furthermore, animals given NMDA receptor antagonists post-retrieval showed a blunted response to cocaine-primed reinstatement. Using two distinct NMDA receptor antagonists in a common setting, these findings demonstrate that NMDA receptor-dependent processes contribute both to the consolidation and reconsolidation of cocaine-cue memories, and they point to the potential utility of treatments that interfere with drug-cue memory reconsolidation.

Activation of glutamate transport evokes rapid glutamine release from perisynaptic astrocytes

Stimulation of astrocytes by neuronal activity and the subsequent release of neuromodulators is thought to be an important regulator of synaptic communication. In this study we show that astrocytes juxtaposed to the glutamatergic calyx of Held synapse in the rat medial nucleus of the trapezoid body (MNTB) are stimulated by the activation of glutamate transporters and consequently release glutamine on a very rapid timescale. MNTB principal neurones express electrogenic system A glutamine transporters, and were exploited as glutamine sensors in this study. By simultaneous whole-cell voltage clamping astrocytes and neighbouring MNTB neurones in brainstem slices, we show that application of the excitatory amino acid transporter (EAAT) substrate d-aspartate stimulates astrocytes to rapidly release glutamine, which is detected by nearby MNTB neurones. This release is significantly reduced by the toxins L-methionine sulfoximine and fluoroacetate, which reduce glutamine concentrations specifically in glial cells. Similarly, glutamine release was also inhibited by localised inactivation of EAATs in individual astrocytes, using internal DL-threo-β-benzyloxyaspartic acid (TBOA) or dissipating the driving force by modifying the patch-pipette solution. These results demonstrate that astrocytes adjacent to glutamatergic synapses can release glutamine in a temporally precise, controlled manner in response to glial glutamate transporter activation. Since glutamine can be used by neurones as a precursor for glutamate and GABA synthesis, this represents a potential feedback mechanism by which astrocytes can respond to synaptic activation and react in a way that sustains or enhances further communication. This would therefore represent an additional manifestation of the tripartite relationship between synapses and astrocytes.

Effect of Chemical Stimulation of the Medial Frontal Lobe on the Micturition Reflex in Rats

We assessed the influence of the medial frontal lobe on micturition after chemical stimulation. We also examined the relation between the medial frontal lobe and the rostral pontine reticular formation, which has a strong inhibitory effect on micturition. A total of 35 female rats underwent continuous cystometry. Bladder activity changes were examined after physiological saline, glutamate, the glutamate receptor antagonist MK-801, noradrenaline or the adrenergic α-1 receptor antagonist naftopidil was injected in the medial frontal lobe. When glutamate was injected in the medial frontal lobe, MK-801 was also injected in the rostral pontine reticular formation. Glutamate injection in the medial frontal lobe prolonged the interval between bladder contractions while injection of the glutamate antagonist MK-801 shortened the interval. Glutamate injection in the medial frontal lobe just after MK-801 injection in the ipsilateral rostral pontine reticular formation also prolonged the interval between bladder contractions. However, after prior injection of MK-801 in the bilateral rostral pontine reticular formation glutamate injection in the medial frontal lobe did not influence cystometric parameters. Noradrenaline injection in the medial frontal lobe shortened the interval between bladder contractions while injection of its antagonist naftopidil prolonged the interval. Medial frontal lobe neurons excited by glutamate inhibited the micturition reflex via activation of the rostral pontine reticular formation by glutamatergic projection while medial frontal lobe neurons excited by noradrenaline facilitated the micturition reflex. Thus, the medial frontal lobe may be an important integration center for the initiation of micturition and urine storage mechanisms.