Alterations in nigral NMDA and GABAA receptor control of the striatal dopamine level after repetitive exposures to nitrogen narcosis

Nitrogen pressure exposure in rats results in decreased dopamine (DA) release at the striatal terminals of the substantia nigra pars compacta (SNc) dopaminergic neurons, demonstrating the narcotic potency of nitrogen. This effect is attributed to decreased excitatory and increased inhibitory inputs to dopaminergic neurons, involving a change in NMDA and GABA_A receptor function. We investigated whether repetitive exposures to nitrogen modify the excitatory and inhibitory control of the dopaminergic nigro-striatal pathway.We used voltammetry to measure dopamine levels in freely-moving rats, implanted with dopamine-sensitive electrodes in the striatum. NMDA/GABA_A receptor agonists (NMDA/muscimol) and antagonists (AP7/gabazine) were administered through a guide-cannula into the SNc, and their effects on striatal dopamine levels were measured under normobaric conditions, before and after five repetitive exposures to 1 MPa nitrogen.NMDA-mediated dopamine release was greater following repetitive exposures, AP7-mediated inhibition of glutamatergic input was blocked, suggesting that NMDA receptor sensitivity was increased and glutamate release reduced. Muscimol did not modify dopamine levels following repetitive exposures, whereas the effect of gabazine was greater after exposures than before. This suggested that interneuronal GABA_A receptors were desensitized, leading to an increased GABAergic input at dopaminergic cells. Thus, repetitive nitrogen exposure induced persistent changes in glutamatergic and GABAergic control of dopaminergic neurons, resulting in decreased activity of the nigrostriatal pathway.     

Increased brain D-amino acid oxidase (DAAO) activity in schizophrenia

D-serine has been shown to be a major endogenous coagonist of the N-methyl D-aspartate (NMDA) type of glutamate receptors. Accumulating evidence suggests that NMDA receptor hypofunction contributes to the symptomatic features of schizophrenia. d-serine degradation can be mediated by the enzyme d-amino acid oxidase (DAAO). An involvement of d-serine in the etiology of schizophrenia is suggested by the association of the disease with single nucleotide polymorphisms in the DAAO and its regulator (G72). The present study aims to further elucidate whether the DAAO activity is altered in schizophrenia. Specific DAAO activity was measured in postmortem cortex samples of bipolar disorder, major depression and schizophrenia patients, and normal controls (n=15 per group). The mean DAAO activity was two-fold higher in the schizophrenia patients group compared with the control group. There was no correlation between DAAO activity and age, age of onset, duration of disease, pH of the tissue and tissue storage time and no effect of gender, cause of death and history of alcohol and substance abuse. The group of neuroleptics users (including bipolar disorder patients) showed significantly higher D-amino acid oxidase activity. However, there was no correlation between the cumulative life-time antipsychotic usage and D-amino acid oxidase levels. In mice, either chronic exposure to antipsychotics or acute administration of the NMDA receptor blocker MK-801, did not change d-amino acid oxidase activity. These findings provide indications that D-serine availability in the nervous system may be altered in schizophrenia because of increased D-amino acid degradation by DAAO.     

Increased serum levels of glutamate in adult patients with autism

BACKGROUND: Precise mechanisms underlying the pathophysiology of autism are currently unknown. Given the major role of glutamate in brain development, we have hypothesized that glutamatergic neurotransmission plays a role in the pathophysiology of autism. In this study, we studied whether amino acids (glutamate, glutamine, glycine, D-serine, and L-serine) related to glutamatergic neurotransmission are altered in serum of adult patients with autism. METHODS: We measured serum levels of amino acids in 18 male adult patients with autism and age-matched 19 male healthy subjects using high-performance liquid chromatography. RESULTS: Serum levels (mean = 89.2 microM, S.D. = 21.5) of glutamate in the patients with autism were significantly (t = -4.48, df = 35, p < 0.001) higher than those (mean = 61.1 microM, S.D. = 16.5) of normal controls. In contrast, serum levels of other amino acids (glutamine, glycine, d-serine, l-serine) in the patients with autism did not differ from those of normal controls. There was a positive correlation (r = 0.523, p = 0.026) between serum glutamate levels and Autism Diagnostic Interview-Revised (ADI-R) social scores in patients. CONCLUSIONS: The present study suggests that an abnormality in glutamatergic neurotransmission may play a role in the pathophysiology of autism.     

In vitro effects of D-2-hydroxyglutaric acid on glutamate binding, uptake and release in cerebral cortex of rats

Neurological dysfunction is common in patients with D-2-hydroxyglutaric aciduria (DHGA). However, the mechanisms underlying the neuropathology of this disorder are far from understood. In the present study, we investigated the in vitro effects of D-2-hydroxyglutaric acid (DGA) at various concentrations (0.1-1.0 mM) on various parameters of the glutamatergic system, namely the basal and potassium-induced release of L-[3H]glutamate by synaptosomal preparations, Na(+)-dependent L-[3H]glutamate uptake by synaptosomal preparations and Na(+)-independent L-[3H]glutamate uptake by synaptic vesicles, as well as of Na(+)-independent and dependent L-[3H]glutamate binding to synaptic plasma membranes from cerebral cortex of male adult Wistar rats. We observed that DGA significantly increased synaptosomal L-[3H]glutamate uptake, without altering the other parameters. Although these findings do not support a direct excitotoxic action for DGA since the metabolite did not affect important parameters of the main neurotransmission system, they do not exclude a direct action of DGA on NMDA or other glutamate receptors. More comprehensive studies are therefore necessary to evaluate the exact role of DGA on neurotransmission.     

The role of glutamate on the action of antidepressants

Major depressive disorder (MDD) is a common, chronic, recurrent mental illness that affects millions of individuals worldwide. Currently available antidepressants are known to affect the monoaminergic (e.g., serotonin, norepinephrine, and dopamine) systems in the brain. Accumulating evidence suggests that the glutamatergic neurotransmission via the excitatory amino acid glutamate also plays an important role in the neurobiology and treatment of this disease. Clinical studies have demonstrated that the non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist ketamine has rapid antidepressant effects in treatment-resistant patients with MDD, suggesting the role of glutamate in the pathophysiology of treatment-resistant MDD. Furthermore, a number of preclinical studies demonstrated that the agents which act at glutamate receptors such as NMDA receptors, α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors and metabotropic glutamate receptors (mGluRs) might have antidepressant-like activities in animal models of depression. In this article, the author reviews the role of glutamate in the neuron-glia communication induced by potential antidepressants.     

A CSF and postmortem brain study of D-serine metabolic parameters in schizophrenia

Clinical trials demonstrated that D-serine administration improves schizophrenia symptoms, raising the possibility that altered levels of endogenous D-serine may contribute to the N-methyl D-aspartate receptor hypofunction thought to play a role in the disease. We hypothesized that cerebro-spinal fluid (CSF) D-serine levels are decreased in the patients due to reduced synthesis and/or increased degradation in brain. We now monitored amino acid levels in CSF from 12 schizophrenia patients vs. 12 controls and in postmortem parietal-cortex from 15 control subjects and 15 each of schizophrenia, major-depression and bipolar patients. In addition, we monitored postmortem brain serine racemase and D-amino acid oxidase protein levels by Western-blot analysis. We found a 25% decrease in D-serine levels and D/L-serine ratio in CSF of schizophrenia patients, while parietal-cortex D-serine was unaltered. Levels of L-serine, L-glutamine and L-glutamate were unaffected. Frontal-cortex (39%) and hippocampal (21%) serine racemase protein levels and hippocampal serine racemase/D-amino acid oxidase ratio (34%) were reduced. Hippocampal D-amino-acid-oxidase protein levels significantly correlated with duration of illness (r=0.6, p=0.019) but not age. D-amino acid oxidase levels in patients with DOI>20 years were 77% significantly higher than in the other patients and controls. Our results suggest that reduced brain serine racemase and elevated D-amino acid oxidase protein levels may contribute to the lower CSF D-serine levels in schizophrenia.     

Glutamatergic neurotransmission in schizophrenics

Glutamate is the most abundant amino acid in the brain, where it plays an important role as a well-established major excitatory neurotransmitter in the central nervous system. It has been suggested that reduced glutamate neurotransmission may be involved in the pathophysiology of schizophrenia. The glutamate hypothesis of schizophrenia postulates alterations in the glutamatergic system as an important neurobiochemical event in the pathophysiology of this group of psychotic disorders. An altered glutamate release from synaptosomes including a hypofunction of different glutamate receptors (i.e. NMDA receptors) from different brain areas have previously been reported. Furthermore, partial agonists at the glycine co-agonist site of the NMDA receptor might be a new approach in the treatment of schizophrenic symptoms but further studies are necessary to clarify the role and efficacy of these substances in schizophrenia. Changes in the glutamatergic cortico-striatal connections in schizophrenia could precipitate a potential perceptive overstimulation of the neocortex from thalamic input and an inhibiting influence of the striatum on the thalamus would modulate the information input of the cortex, thereby possibly counteracting the disturbed information processing which is relatively characteristic for schizophrenic psychoses.     

Modulatory effects of d-serine and sarcosine on NMDA receptor-mediated neurotransmission are apparent after stress in the genetically inbred BALB/c mouse strain

Abnormalities of NMDA receptor-mediated neurotransmission are involved in the pathophysiology of schizophrenia, Alzheimer's disease, substance abuse and seizure disorders. The NMDA receptor is implicated in schizophrenia because phencyclidine (PCP), a noncompetitive NMDA receptor antagonist, binds to a hydrophobic domain within the channel, precipitating a schizophreniform psychosis in susceptible persons. Pharmacological, environmental, and genetic variables alter NMDA receptor-mediated neurotransmission. Inbred mouse strains differ in their sensitivity to some of the behavioral effects of MK-801 (dizocilpine), a PCP analogue. The NMDA receptor complex in the BALB/c strain could reflect a unique stoichiometric combination of receptor subunits resulting in a higher proportion of the channels in the open configuration, a higher affinity of MK-801 for its hydrophobic channel domain, and/or a combination of the above. The BALB/c mouse strain, "stressed" mice, and behavioral consequences of MK-801 administration represent models of altered glutamatergic neural transmission. We were interested in examining the effect of stress on the modulatory properties of d-serine and sarcosine. d-Serine is a naturally occurring glycine agonist that modulates the ability of l-glutamate to influence the opening of the NMDA receptor-associated ionophore, and sarcosine is a naturally occurring glycine reuptake inhibitor. The data suggest that 24h after stress, d-serine and sarcosine interact synergistically to reduce MK-801's ability to antagonize electrically precipitated tonic hindlimb extension. Under conditions of stress, modulatory effects of d-serine and sarcosine on the antiseizure effect of MK-801 are observed that are not apparent in the nonstress condition. The results could be relevant to the development of glycinergic interventions for the treatment of neuropsychiatric disorders.     

Simultaneous quantification of D- vs. L-serine, taurine, kynurenate, phosphoethanolamine and diverse amino acids in frontocortical dialysates of freely-moving rats: differential modulation by N-methyl-D-aspartate (NMDA) and other pharmacological agents

This study describes a novel analytical method permitting simultaneous HPLC-fluorimetric quantification of multiple (15) d- and l-amino acids, kynurenate, taurine and phosphoethanolamine (a marker of membrane integrity) in microdialysates of prefrontal cortex of freely-moving rats. Levels of GABA were elevated by the transporter inhibitor, nipecotic acid, and by the transaminase inhibitor, vigabatrine. Supporting a neuronal origin, they were decreased by the GABA_B autoreceptor agonist, baclofen, yet unaffected by fluoroacetate which disrupts glial metabolism. Glutamate levels were elevated by the transporter inhibitor, l-trans-PDC, and mainly neuronal since they were not decreased by fluoroacetate, yet reduced by baclofen (which recruits GABA_B receptors on glutamatergic terminals) and elevated by the NMDA receptor antagonist, dizocilpine. By contrast, levels of glutamine were reduced by l-trans-PDC. Consistent with glial origin, they were unaffected by baclofen, yet reduced by fluoroacetate. Administration of d-serine selectively increased its levels over l-serine, and vice versa. d-serine modestly decreased levels of glycine, which were enhanced by administration of glycine itself and of the glycine transporter-1 inhibitor, sarcosine. Kynurenate levels were increased by its precursor, kynurenine, an effect abolished by the amino-transferase inhibitor, amino-oxyacetate. Taurine and the energy drink, Red Bull^®, selectively elevated levels of taurine, which were only slightly reduced by fluoroacetate. Finally, administration of NMDA increased levels of taurine, kynenurate and phosphoethanolamine, while reducing d-serine. These actions were abolished by the competitive NMDA receptor antagonist, CPP, which was inactive alone. This broad-based dialysis system should prove instructive for exploring actions of psychotropic drugs, and for characterising animal models of CNS disorders.     

Failure of glycine site NMDA receptor antagonists to protect againstl-2-chloropropionic acid-induced neurotoxicity highlights the uniqueness of cerebellar NMDA receptors

Cultured cerebellar granule cells and cerebellar slices from neonatal rats have been widely used to examine the biochemistry of excitatory amino acid-induced cell death mediated in part by the activation of NMDA receptors. However, the NMDA subunit stoichiometry, producing functional NMDA receptors is different in cultured granule cells, neonatal and adult rat cerebellum as compared to the NMDA receptors in forebrain regions. We have used thel-2-chloropropionic acid (l-CPA) (750 mg/kg) model of NMDA-medialed selective cerebellar granule cell necrosis in vivo to examine the role of the glycine binding site and possible effect of the NR2C subunit (which is largely expressed only in the cerebellum) on granule cell necrosis. The abilities of various NMDA receptor antagonists were examined in vivo to determine the relative contribution of both glutamate and glycine sites involved in thel-CPA-induced neurotoxicity. The potent neuroprotective, non-competitive NMDA receptor antagonist dizocilpine (MK-801) was compared with glutamate and glycine site NMDA antagonists. We have examined a number of markers for thel-CPA-induced granule cell necrosis. Thel-CPA-induced reduction in cerebellar aspartate and glutamate concentrations were used as markers of granule cell necrosis. We also measured the cerebellar water content and sodium concentrations as measures of thel-CPA-induced cerebellar edema that accompanies the granule cell necrosis. Finally the ability of the NMDA antagonists to attenuate thel-CPA-induced reductions in body weight gain and the prevention of the loss in hindlimb function using a behavioral measure of hindlimb retraction were examined. The potent glutamate antagonists, CPP and CGP40116 and dizocilpine prevented thel-CPA-induced locomotor dysfunction and granule cell necrosis as measured by their ability to preventl-CPA-induced reductions in aspartate and glutamate concentrations. CPR CGP40116 and dizocilpine also prevented the appearance of cerebellar edema followingl-CPA administration. In addition, dizocilpine, CPP and CGP40116 were able to partially prevent thel-CPA-induced loss in body weight over the 48 h experimental period. In contrast, none of the glycine partial agonists or antagonists, namely (±)HA-966,d-cycloserine, MDL-29951, DPCQ, MNQX or L-701 252 were able to prevent thel-CPA-induced loss in body weight,l-CPA-induced granule cell necrosis and behavioral disturbances when administered to rats. None of the NMDA antagonists had any effect on the cerebellar neurochemistry when injected alone or had any effect on animal behavior except for dizocilpine, CPP, CGP401 l6 and (±)HA-966 which resulted in a transient sedation for between three and five hours immediately following their administration. In conclusion, we demonstrate that NMDA open channel blockade and glutamate antagonists can provide full neuroprotection against thel-CPA-induced granule cell necrosis. The failure of the glycine partial agonists and antagonists to provide any neuroprotection againstl-CPA-induced neurotoxicity in the cerebellum contrasts with their neuroprotective efficacy in other animal models of excitatory amino acid-induced cell death in forebrain regions in vivo. We therefore suggest that the glycine site plays a lesser role in modulating NMDA receptor function in the cerebellum and may explain why cells expressing NMDA receptors composed of NR1/NR2C subunits are particularly resistant to excitatory amino acid-induced neurotoxicity.     

The emerging role of glutamate in the pathophysiology and treatment of schizophrenia

OBJECTIVE: Research has implicated dysfunction of glutamatergic neurotransmission in the pathophysiology of schizophrenia. This review evaluates evidence from preclinical and clinical studies that brain glutamatergic neurotransmission is altered in schizophrenia, may affect symptom expression, and is modulated by antipsychotic drugs. METHOD: A comprehensive review of scientific articles published over the last decade that address the role of glutamate in the pathophysiology of schizophrenia was carried out. RESULTS: Glutamatergic neurons are the major excitatory pathways linking the cortex, limbic system, and thalamus, regions that have been implicated in schizophrenia. Postmortem studies have revealed alterations in pre- and postsynaptic markers for glutamatergic neurons in several brain regions in schizophrenia. The N-methyl-D-aspartic acid (NMDA) subtype of glutamate receptor may be particularly important as blockade of this receptor by the dissociative anesthetics reproduces in normal subjects the symptomatic manifestations of schizophrenia, including negative symptoms and cognitive impairments, and increases dopamine release in the mesolimbic system. Agents that indirectly enhance NMDA receptor function via the glycine modulatory site reduce negative symptoms and variably improve cognitive functioning in schizophrenic subjects receiving typical antipsychotics. CONCLUSIONS: Dysfunction of glutamatergic neurotransmission may play an important role in the pathophysiology of schizophrenia, especially of the negative symptoms and cognitive impairments associated with the disorder, and is a promising target for drug development.     

Oxidative stress in schizophrenia: an integrated approach

Oxidative stress has been suggested to contribute to the pathophysiology of schizophrenia. In particular, oxidative damage to lipids, proteins, and DNA as observed in schizophrenia is known to impair cell viability and function, which may subsequently account for the deteriorating course of the illness. Currently available evidence points towards an alteration in the activities of enzymatic and nonenzymatic antioxidant systems in schizophrenia. In fact, experimental models have demonstrated that oxidative stress induces behavioral and molecular anomalies strikingly similar to those observed in schizophrenia. These findings suggest that oxidative stress is intimately linked to a variety of pathophysiological processes, such as inflammation, oligodendrocyte abnormalities, mitochondrial dysfunction, hypoactive N-methyl-d-aspartate receptors and the impairment of fast-spiking gamma-aminobutyric acid interneurons. Such self-sustaining mechanisms may progressively worsen producing the functional and structural consequences associated with schizophrenia. Recent clinical studies have shown antioxidant treatment to be effective in ameliorating schizophrenic symptoms. Hence, identifying viable therapeutic strategies to tackle oxidative stress and the resulting physiological disturbances provide an exciting opportunity for the treatment and ultimately prevention of schizophrenia.     

Spinal neuronal pathology associated with continuous intrathecal infusion ofN-methyl-d-aspartate in the rat

Summary Continuous intrathecal infusion ofN-methyl-d-aspartate (NMDA) at the level of the lumbar enlargement of the spinal cord in middle-aged rats produced dose-dependent toxicity of spinal cord neuronal systems. Toxicity was enhanced by coadministration of glycine, but was significantly reduced when NMDA was coadministered with the competitive inhibitordl-2-amino-5-phosphovaleric acid or the noncompetitive inhibitor MgSO₄. The toxic effects of NMDA were manifest most dramatically and at the lowest concentrations in the neuropil, while neuronal loss was obvious at higher concentrations. The distribution and intensity of reactive astrocytosis was consistent with the known regional and subcellular distribution of NMDA receptors in the spinal cord of rats. The increase in ribosomes and rough endoplasmic reticulum observed in anterior horn cells suggested an increase of cell metabolism reflecting either a nonspecific response to injury or a specific increase in cell metabolism secondary to sustained activation of NMDA receptors. The present studies implicate excitatory amino acid receptors of the NMDA type in producing toxicity to selected neuronal populations of the spinal cord. This model provides a system for studies of the protective effects and rescue of neuronal populations susceptible to the toxic effects of excitatory amino acids.Supported by the ALS Society of Canada     

No genetic association between SLC7A10 and Japanese patients with schizophrenia

Disrupted glutamatergic neurotransmission may be a pathophysiological feature in the brains from patients with schizophrenia, and glutamatergic amino acids including d-serine have been found to be involved in pathophysiology. Endogenous and exogenous d-serine have shown potential as biological markers for the pathophysiology of schizophrenia and especially as a therapeutic strategy in treatment-resistant schizophrenia (TRS). This is the first study investigating whether SLC7A10, a d-serine transporter gene, is associated with schizophrenia in Japanese patients.We investigated the association between schizophrenia in Japanese patients with SLC7A10 using six tag single nucleotide polymorphisms (SNPs). Results failed to show any association between Japanese schizophrenia and each individual SNP or with two-, three-, or four-window haplotype analyses. We also investigated whether SLC7A10 contributes to TRS in Japanese participants. Results showed no association.In conclusion, SLC7A10 had no apparent degree of association with schizophrenia as a candidate susceptibility gene in the disease per se.     

Down-regulation of glutaredoxin by estrogen receptor antagonist renders female mice susceptible to excitatory amino acid mediated complex I inhibition in CNS

beta-N-oxalyl-amino-L-alanine, (L-BOAA), an excitatory amino acid, acts as an agonist of the AMPA subtype of glutamate receptors. It inhibits mitochondrial complex I in motor cortex and lumbosacral cord of male mice through oxidation of critical thiol groups, and glutaredoxin, a thiol disulfide oxido-reductase, helps maintain integrity of complex I. Since incidence of neurolathyrism is less common in women, we examined the mechanisms underlying the gender-related effects. Inhibition of complex I activity by L-BOAA was seen in male but not female mice. Pretreatment of female mice with estrogen receptor antagonist ICI 182,780 or tamoxifen sensitizes them to L-BOAA toxicity, indicating that the neuroprotection is mediated by estrogen receptors. L-BOAA triggers glutathione (GSH) loss in male mice but not in female mice, and only a small but significant increase in oxidized glutathione (GSSG) was seen in females. As a consequence, up-regulation of gamma-glutamyl cysteinyl synthase (the rate-limiting enzyme in glutathione synthesis) was seen only in male mouse CNS but not in females. Both glutathione reductase and glutaredoxin that reduce oxidized glutathione and protein glutathione mixed disulfides, respectively, were constitutively expressed at higher levels in females. Furthermore, glutaredoxin activity in female mice was down-regulated by estrogen antagonist indicating its regulation by estrogen receptor. The higher constitutive expression of glutathione reductase and glutaredoxin could potentially confer neuroprotection to female mice.     

A flow-cytometric method to investigate glutamate-receptor-sensitivity in whole blood platelets - results from healthy controls and patients with schizophrenia

Hypofunction of glutamate receptors may contribute to the symptoms of schizophrenia. Human platelets express glutamate receptors and can serve as peripheral surrogate model for neuronal cells. Aim of this study was to establish a fast and sensitive flow-cytometric method to determine the glutamate-dependent kinetics of intracellular calcium ([Ca++]i) mobilization in platelets of schizophrenic patients. Glutamate stimulated [Ca++]i response was measured with a flow-cytometer in anti-CD-41a-labelled whole blood platelets of treated schizophrenic patients (n = 18) and controls (n = 18). In two control experiments the NMDA-receptor antagonist MK-801 and the dopamine antagonist amisulpride, respectively, were added to probes from healthy subjects. Stimulation with glutamate led dose-dependently to a mobilization of [Ca++]i in both healthy controls and patients. This effect was significantly reduced in patients. In vitro NMDA-antagonism inhibited the glutamate response, whereas dopamine-antagonism had no effect. Our flow-cytometric method allows to measure glutamate-receptor mediated [Ca++]i response in whole blood platelets, without requiring platelet rich preparations. The reduced glutamate-response in the patients was not explained by a direct inhibitory treatment effect. However, further studies with drug naive patients will be necessary to find out whether or not the observed hypoglutamergic function of platelets is endogenous to the disorder.     

Low cerebrospinal fluid glutamate and glycine in refractory affective disorder.

BACKGROUND: Glutamatergic dysregulation has been documented in schizophrenia but has received less systematic study in affective illness. METHODS: Cerebrospinal fluid (CSF) levels of the excitatory amino acids glutamate (Glu) and aspartate (Asp) and the N-methyl-D-aspartate (NMDA) receptor modulator, glycine (GLY) were measured by high performance liquid chromatography in 32 patients with refractory affective disorder (16 female/16 male, 12 bipolar I, 12 bipolar II, and 8 unipolar) and in 14 age-matched controls. RESULTS: There was a significant reduction in CSF glutamate and glycine in patients versus controls. A diagnosis by sex interaction was present for CSF glycine with lower levels in female patients compared to female controls. Levels of the excitatory amino acids were highly inter-correlated in patients, but not in controls. In patients studied after 6 weeks of lamotrigine, there was a trend for CSF glutamate levels to increase. CONCLUSIONS: These data suggest that in patients with refractory affective disorder, excitatory amino acids are dysregulated, as exemplified both by the decreased CSF glutamate and glycine and their high intercorrelation compared to controls. Further controlled study of glutamatergic dysregulation and its relationship to the pathophysiology of affective disorders and potential mechanism of action of mood stabilizers appears indicated.     

Hepatocyte growth factor promotes the number of PSD-95 clusters in young hippocampal neurons

Hepatocyte growth factor (HGF) and its receptor are expressed in various regions of the brain and have protective effects against excitotoxic injuries. However, their effects on synapse formation remain to be elucidated. To determine whether HGF has the ability to alter synaptic function during development, we investigated changes in the number of synapse detected by double immunostaining for NMDA receptor subunits and a presynaptic marker in cultured young hippocampal neurons. Whereas application of HGF increased the number of cluster of synapsin, a presynaptic protein, the clusters of NMDA receptor subunits NR1 and NR2B were not altered. Interestingly, colocalization of PSD-95, a scaffolding protein of the receptor, with synapsin was increased by HGF treatment without a change in the total amount of it. In addition, we investigated the expression of surface NMDA receptor, neuroligin, and neurexin, which were assessed by use of a cell-surface biotinylation assay. The application of HGF did not change the surface expression of these proteins. Furthermore, we determined the release of glutamate in response to depolarization. Treatment with HGF promoted depolarization-evoked release of glutamate. These results suggest that HGF modulates the expression of the scaffolding protein of the NMDA receptor at the synapse and promotes maturation of excitatory synapses in young hippocampal neurons.     

In vivo monitoring of extracellular glutamate in the brain with a microsensor

Recent discoveries have revealed that glutamatergic neurotransmission in the central nervous system is mediated by a dynamic interplay between neurons and astrocytes. To enhance our understanding of this process, the study of extracellular glutamate is crucial. At present, microdialysis is the most frequently used analytical technique to monitor extracellular glutamate levels directly in the brain. However, the neuronal and physiological origin of the detected glutamate levels is questioned as they do not fulfil the classical release criteria for exocytotic release, such as calcium dependency or response to the sodium channel blocker tetrodotoxine (TTX). It is hypothesized that an analytical technique with a higher spatial and temporal resolution is required. Glutamate microsensors provide a promising analytical solution to meet this requirement. In the present study, we applied a 10 micro m diameter hydrogel-coated glutamate microsensor to monitor extracellular glutamate levels in the striatum of anesthetized rats. To explore the potential of the microsensor, different pharmacological agents were injected in the vicinity of the sensor at an approximate distance of 100 micro m. It was observed that KCl, exogenous glutamate, kainate and the reuptake inhibitor DL-threo-beta-benzyloxyaspartate (DL-TBOA) increased the extracellular glutamate levels significantly. TTX decreased the basal extracellular glutamate levels approximately 90%, which indicates that the microsensor is capable of detecting neuronally derived glutamate. This is one of the first studies in which a microsensor is applied in vivo on a routine base, and it is concluded that microsensor research can contribute significantly to improve our understanding of the physiology of glutamatergic neurotransmission in the brain.