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Aberrant behaviors related to learning and memory in olfactory bulbectomized (OBX) mice have been documented in the previous studies. We reported that the impairment of long-term potentiation (LTP) of hippocampal CA1 regions from OBX mice was associated with down-regulation of CaM kinase II (CaMKII) and protein kinase C (PKC) activities. We now demonstrated that the nootropic drug, nefiracetam, significantly improved spatial reference memory-related behaviors as assessed by Y-maze and novel object recognition task in OBX mice. Nefiracetam also restored hippocampal LTP injured in OBX mice. Nefiracetam treatment restored LTP-induced PKCalpha (Ser657) and NR1 (Ser896) phosphorylation as well as increase in their basal phosphorylation in the hippocampal CA1 region of OBX mice. Likewise, nefiracetam improved LTP-induced CaMKIIalpha (Thr286) autophosphorylation and GluR1 (Ser831) phosphorylation and increased their basal phosphorylation. The enhancement of PKCalpha (Ser657) and CaMKIIalpha (Thr286) autophosphorylation by nefiracetam was inhibited by treatment with (+/-)-alpha-Methyl-(4-carboxyphenyl)glycine and DL-2-Amino-5-phosphonovaleric acid, respectively. The enhancement of LTP induced by nefiracetam is inhibited by treatment with 2-methyl-6-(phenylethynyl)-pyridine, but not by treatment with LY367385, suggesting that metabotropic glutamate receptor 5 (mGluR5) but not mGluR1 is involved in the nefiracetam-induced LTP enhancement. Taken together, nefiracetam ameliorates OBX-induced deficits in memory-related behaviors and impairment of LTP in the hippocampal CA1 region through activation of NMDAR and mGluR5, thereby leading to an increase in activities of CaMKIIalpha (Thr286) and PKCalpha (Ser657), respectively.

The role of neurotoxic non-protein amino acid beta-N-methylamino-L-alanine (L-BMAA) as a putative causative agent of Western pacific amyotrophic lateral sclerosis/Parkinsonism dementia complex (ALS/PDC) has recently been reinvigorated. In view of this data we have investigated the strength and mechanism of effect of L-BMAA in presence of 20 mmol/L bicarbonate (a cofactor for BMAA) on membrane potential of the Leech Haemopis sanguisuga. Our results show that L-BMAA has excitatory effect in bicarbonate containing solution, which is more potent than in nominally bicarbonate free solution. This potentiation by bicarbonate is L-BMAA specific, as it was not exhibited by beta-N-oxalylamino-L-alanine. The effect of L-BMAA was partially blocked by non-NMDA receptor antagonist CNQX. Application of L-BMAA caused a decrease in input membrane resistance, an increase of intracellular sodium activity, and a decrease of intracellular potassium activity. Present findings indicate that BMAA could initiate excitotoxicity through activation of non-NMDA ionotropic glutamate receptors.

In this review, we will summarize our ongoing studies on the functionality of both gamma-aminobutyric acid (GABA) and glutamate receptors expressed by undifferentiated neural progenitor cells isolated from embryonic rodent brains. Cells were cultured with growth factors for the formation of round spheres by clustered cells under floating conditions, whereas a reverse transcription polymerase chain reaction analysis revealed expression of mRNA for particular subtypes of different ionotropic and metabotropic GABA and glutamate receptors in undifferentiated progenitors and neurospheres. Moreover, sustained exposure to either GABAergic or glutamatergic agonists not only modulated the size of neurospheres formed, but also affected spontaneous and induced differentiation of neural progenitor cells into particular progeny cell lineages such as neurons and astroglia. Both GABA and glutamate could play a pivotal role in the mechanisms underlying proliferation for self-replication along with the determination of subsequent differentiation fate toward particular progeny lineages through activation of their receptor subtypes functionally expressed by undifferentiated neural progenitor cells. Accordingly, neurogenesis seems to be also under control by GABAergic and glutamatergic signaling in developing brains as seen with neurotransmission in adult brains.

Two pathways for glutamate toxicity have been described, receptor-mediated excitotoxicity and non-receptor mediated oxidative glutamate toxicity. Here, we show that two distinct forms of receptor-mediated primary cortical neuronal death exist, chronic and acute glutamate toxicity, and that these depend on exposure time. In vitro, neuronal sensitivity to chronic glutamate exposure increased as neurons matured and the initial plating medium contributed as well. In immature neurons, high concentrations of glutamate induced neuronal death. The chronic glutamate toxicity was independent of neuronal density, whereas increased density potentiated acute glutamate toxicity. Activation of ionotropic glutamate receptors (iGluRs) contributed to induction of chronic and acute glutamate toxicity at similar rates at DIV14. Inactivation of the metabotropic glutamate receptors (mGluRs) by AIDA increased neuronal sensitivity to chronic glutamate exposure but not to acute exposure. Neuronal death by acute toxicity was much faster than by chronic toxicity in which activation of mGluRs was involved. These results suggest that acute glutamate toxicity is quite different from chronic toxicity, in which activation of mGluRs is associated with resistance to glutamate toxicity.

Several studies have demonstrated that systemically administered N-methyl-d-aspartate (NMDA) receptor antagonists increase serotonin (5-HT) and glutamate release in the medial prefrontal cortex (mPFC). Previously we showed that the perfusion of clozapine in the mPFC prevented the MK-801-induced increase in extracellular glutamate and 5-HT whereas haloperidol blocked only the effect of MK-801 on glutamate. To study the contribution of different monoaminergic receptors (for which clozapine and haloperidol exhibit distinct affinities) to these effects, here we used in-vivo microdialysis to examine the role of local blockade of dopamine D2, 5-HT2A and alpha1-adrenergic receptors as well as agonism at dopamine D1 and 5-HT1A receptors in the mPFC on the increased efflux of glutamate and 5-HT elicited by MK-801. The results show that M100907 (5-HT2A antagonist), BAY x 3702 (5-HT1A agonist) and prazosin (alpha1-adrenergic antagonist) blocked the MK-801-induced increase of 5-HT and glutamate in the mPFC. However, raclopride, eticlopride (dopamine D2 antagonists) and SKF-38393 (dopamine D1 agonist) were able to prevent the increased efflux of glutamate (but not that of 5-HT) elicited by MK-801. We propose that D2 receptor antagonists and D1 agonists would act predominantly on a subpopulation of GABAergic interneurons of the mPFC, thus leading to an enhanced cortical inhibition that would prevent an excessive glutamatergic transmission. On the other hand, atypical antipsychotic drugs might further act upon 5-HT2A, 5-HT1A and alpha1-adrenoceptors present in pyramidal cells (including those projecting to the dorsal raphe nucleus), which would directly inhibit an excessive excitability of these cells.

An animal model of depression combining genetic vulnerability and early-life stress (ELS) was prepared by submitting the Flinders Sensitive Line (FSL) rats to a standard paradigm of maternal separation. We analysed hippocampal synaptic transmission and plasticity in vivo and ionotropic receptors for glutamate in FSL rats, in their controls Flinders Resistant Line (FRL) rats, and in both lines subjected to ELS. A strong inhibition of long-term potentiation (LTP) and lower synaptic expression of NR1 subunit of the NMDA receptor were found in FSL rats. Remarkably, ELS induced a remodelling of synaptic plasticity only in FSL rats, reducing inhibition of LTP; this was accompanied by marked increase of synaptic NR1 subunit and GluR2/3 subunits of AMPA receptors. Chronic treatment with escitalopram inhibited LTP in FRL rats, but this effect was attenuated by prior ELS. The present results suggest that early gene-environment interactions cause lifelong synaptic changes affecting functional and molecular aspects of plasticity, partly reversed by antidepressant treatments.

The present study examined the antinociceptive effects of a hydroalcoholic extract of Polygala paniculata in chemical and thermal behavioural models of pain in mice. The antinociceptive effects of hydroalcoholic extract was evaluated in chemical (acetic-acid, formalin, capsaicin, cinnamaldehyde and glutamate tests) and thermal (tail-flick and hot-plate test) models of pain or by biting behaviour following intratecal administration of both ionotropic and metabotropic agonists of excitatory amino acids receptors glutamate and cytokines such as interleukin-1beta (IL-1beta) and tumour necrosis factor-alpha (TNF-alpha) in mice. When given orally, hydroalcoholic extract (0.001-10 mg/kg), produced potent and dose-dependent inhibition of acetic acid-induced visceral pain. In the formalin test, the hydroalcoholic extract (0.0001-0.1 mg/kg orally) also caused significant inhibition of both the early (neurogenic pain) and the late (inflammatory pain) phases of formalin-induced licking. However, it was more potent and efficacious in relation to the late phase of the formalin test. The capsaicin-induced nociception was also reduced at a dose of only 1.0 mg/kg orally. The hydroalcoholic extract significantly reduced the cinnamaldehyde-induced nociception at doses of 0.01, 0.1 and 1.0 mg/kg orally. Moreover, the hydroalcoholic extract (0.001-1.0 mg/kg orally) caused significant and dose-dependent inhibition of glutamate-induced pain. However, only rutin, but not phebalosin or aurapten, isolated from P. paniculata, administered intraperitoneally to mice, produced dose-related inhibition of glutamate-induced pain. Furthermore, the hydroalcoholic extract (0.1-100 mg/kg orally) had no effect in the tail-flick test. On the other hand, the hydroalcoholic extract caused a significant increase in the latency to response at a dose of 10 mg/kg orally, in the hot-plate test. The hydroalcoholic extract (0.1 mg/kg orally) antinociception, in the glutamate test, was neither affected by intraperitoenal treatment of animals with l-arginine (precursor of nitric oxide, 600 mg/kg) and naloxone (opioid receptor antagonist, 1 mg/kg) nor associated with non-specific effects such as muscle relaxation or sedation. In addition, oral administration of hydroalcoholic extract produced a great inhibition of the pain-related behaviours induced by intrathecal injection of glutamate, N-methyl-D-aspartate (NMDA), IL-1beta and TNF-alpha, but not by alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA), kainate or trans-1-amino-1.3-cyclopentanediocarboxylic acid (trans-ACPD). Together, our results suggest that inhibition of glutamatergic ionotropic receptors, may account for the antinociceptive action reported for the hydroalcoholic extract from P. paniculata in models of chemical pain used in this study.

Polychlorinated biphenyls (PCBs), major environmental hormonally active agents, are metabolized into hydroxylated PCBs in the liver to facilitate excretion. Some of hydroxylated PCBs also have potencies disturbing endogenous hormonal activities at least in vitro. Hormonal activities of hydroxylated PCBs raise a possibility of their interfering with normal brain development which is strictly regulated by endogenous hormones. We investigated whether and how prenatal exposure to a congener of hydroxylated PCBs (4-OH-2',3,3',4',5'-penta CB; 4-OH-PCB106) having activities to disrupt thyroid hormone-dependent signals in vitro could perturb normal gene expression in the developing brain in vivo. Pregnant rats were exposed to 4-OH-PCB106 subcutaneously at the dose of 1.0mg/(kgday) from day 7 of gestation to postnatal day 1. Then three brain regions (cerebral cortex, hippocampus and striatum) were obtained from offspring on postnatal day 1 and subjected to further gene expression analyses. Comprehensive analyses of mRNA expression by oligo DNA microarrays and subsequent validations by quantitative RT-PCR revealed that prenatal exposure to 4-OH-PCB106 affected mRNA expression of glutamate receptors as well as that of thyroid hormone-responsive genes in region-specific manners. Concomitantly 4-OH-PCB106 exposure increased mRNA expression of genes related to exocytosis in the three brain regions. These results raise the possibility that prenatal exposure to some hydroxylated PCBs with thyroid hormone-disrupting potencies leads to abnormal brain development via perturbations on the expression of genes involved in glutamatergic neurotransmission.

Purpose: The anterior cingulate cortex (ACC)--which plays a role in pain, emotions and behavior--can generate epileptic seizures. To date, little is known on the neuronal mechanisms leading to epileptiform synchronization in this structure. Therefore, we investigated the role of excitatory and inhibitory synaptic transmission in epileptiform activity in this cortical area. In addition, since the ACC presents with a high density of opioid receptors, we studied the effect of opioid agonism on epileptiform synchronization in this brain region., Methods: We used field and intracellular recordings in conjunction with pharmacological manipulations to characterize the epileptiform activity generated by the rat ACC in a brain slice preparation., Results: Bath-application of the convulsant 4-aminopyridine (4AP, 50 microM) induced both brief and prolonged periods of epileptiform synchronization resembling interictal- and ictal-like discharges, respectively. Interictal events could occur more frequently before the onset of ictal activity that was contributed by N-methyl-D-aspartate (NMDA) receptors. Mu-opioid receptor activation abolished 4AP-induced ictal events and markedly reduced the occurrence of the pharmacologically isolated GABAergic synchronous potentials. Ictal discharges were replaced by interictal events during GABAergic antagonism; this GABA-independent activity was influenced by subsequent mu-opioid agonist application., Conclusions: Our results indicate that both glutamatergic and GABAergic signaling contribute to epileptiform synchronization leading to the generation of electrographic ictal events in the ACC. In addition, mu-opioid receptors appear to modulate both excitatory and inhibitory mechanisms, thus influencing epileptiform synchronization in the ACC.

We studied the effect of corticotropin-like intermediate lobe peptide (CLIP) on presynaptic NMDA receptors and postsynaptic GABA, NMDA, and AMPA receptors in rat brain. CLIP inhibited presynaptic and postsynaptic NMDA receptors, but potentiated postsynaptic GABA and AMPA receptors. Our results indicate that CLIP modulates function of ionotropic receptors for glutamate and GABA.

The treatment of schizophrenia for the last half century has been with dopamine (DA) D(2) receptor blockers, implicating a hyperdopamine basis for psychosis. However, a 2007 report found that the glutamate agonist LY404039 was effective in schizophrenia, suggesting a hypoglutamate state for the illness. Although phencyclidine psychosis also supports a hypoglutamate cause, assessing the basic and clinical findings shows that phencyclidine has DA D(2) agonist actions as well. Accurate Dreiding models of phencyclidine and the LY glutamate agonists precisely fit the known tetrahedral model of the D(2) receptor that accommodates all DA agonists. A further view is that metabotropic glutamate agonists also exert D(2) agonism, and their antipsychotic doses (about 100 mg/d) are predicted by their dissociation constants (about 20 nM) for D(2). Hence, the clinical antipsychotic action of a glutamate agonist may depend on its ability to interfere with DA neurotransmission by its DA partial agonism.

The preference for foods or fluids in rats is partly dependent on its postingestive consequences. Many studies have investigated postingestive effect of high caloric substances, such as carbohydrate or fat. In this study, we examined postingestive effect of L-glutamate at the preferable concentration using conditioned flavor preference paradigm. Adult male rats with chronic intragastric (IG) cannula were trained to drink a flavored solution (conditioned stimulus; CS+) paired with IG infusion of nutrient solution and another flavored solution (CS-) with IG distilled water infusion on alternate days. The nutrient solution was 60mM monosodium L-glutamate, sodium chloride or glucose. Before and after conditioning, rats received 30min two-bottle choice tests for CS+ and CS- solution. All groups exhibited no significant preference for CS+ in pre-test period. By the last half of conditioning period, intake of CS+ solution was significantly higher than that of CS- in MSG group, but not in NaCl and glucose groups. After conditioned, the MSG group showed significantly higher intake and preference for CS+ solution (69.9%), while the NaCl and glucose group did not show any significant intake and preference for CS+ solution (50.9%, 43.5%, respectively). These results indicate that the amino acid L-glutamate at a preferable concentration has a positive postingestive effect as demonstrated by its ability to condition a flavor preference. The mechanism(s) for this positive effect could be through a direct effect on gut Glu receptors rather than the provision of calories or glucose from metabolized Glu; Further studies are needed to test these hypotheses.

In the brain stem glycine is associated with multiple sensory and visceral regulations, being involved in, for instance, cardiovascular, respiratory and auditory functions. We here studied the mechanisms of the release of preloaded [(3)H]glycine from mouse brain stem slices in a superfusion system. A depolarizing concentration of K(+) ions (50 mM) evoked glycine release, but in the absence of Ca(2+) the effect was attenuated, indicating that a part of the evoked release represents Ca(2+)-dependent exocytosis. The Ca(2+)-independent release was enhanced by omission of Na(+) and Cl(-). The stimulatory effect of extracellular glycine confirmed the involvement of transporters functioning in a reverse direction. A part of the release is mediated by Na(+) and Cl(-) channels, since it was inhibited by the inhibitors of these, riluzole and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonate, respectively. Glycine release was potentiated by the activation of protein kinase C and diminished by increasing cyclic guanosine monophosphate levels with a phosphodiesterase inhibitor, zaprinast. The release was also modulated by the phospholipase inhibitor quinacrine and the tyrosine kinase inhibitor genistein. Adenosine A(1) receptors likewise regulate glycine release, since it was enhanced by their agonist R(-)N(6)-(2-phenylisopropyl)adenosine, which effect was blocked by the antagonist 8-cyclopentyl-1,3-dipropylxanthine. The ionotropic glutamate receptor agonists N-methyl-D: -aspartate, kainate and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate failed to have any effects contrary to their effects in higher brain regions, e.g., in the hippocampus. The group I and III metabotropic glutamate receptor agonists (S)-3,5-dihydroxyphenylglycine and O-phospho-L: -serine, respectively, increased the release in a receptor-mediated manner. Glycine release in the brain stem was also markedly enhanced by cell-damaging conditions, including hypoxia, hypoglycemia and ischemia.

Levels of ionotropic glutamate (Glu) N-methyl-d-aspartic acid (NMDA), 2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propionic acid (AMPA), and kainic acid (KA) receptors in forebrain regions of juvenile rats (age 42 days) were quantified after 3 weeks of treatment with three different doses of risperidone (0.3, 1.0 and 3.0 mg/kg) and compared findings to those in adult rats treated with risperidone (3.0 mg/kg/day) previously. Risperidone (at 0.3 mg/kg/day) did not alter levels of three ionotropic Glu receptors in all brain regions examined. Risperidone (at 1.0 and 3.0 mg/kg/day) significantly decreased NMDA binding in caudate-putamen of juvenile and adult animals. In contrast, the same two doses of risperidone decreased NMDA receptors in nucleus accumbens of juveniles and not adults. Risperidone (at 1.0 and 3.0 mg/kg/day) increased AMPA receptors in medial prefrontal cortex and caudate-putamen of juvenile animals, whereas risperidone (at 3.0 mg/kg) increased AMPA receptors in caudate-putamen and hippocampus of adults. Kainate receptors were not altered by any dose of risperidone in any brain region examined in developing and mature animals. The findings indicate that risperidone exerts dose-dependent effects on Glu receptor subtypes in developing animals, and that Glu receptor responses to repeated administration of risperidone are different in juvenile animals than adults.

Chronic heart failure (CHF) is characterized by increased sympathetic tone. The glutamatergic input in the rostral ventrolateral medulla (RVLM), which is a key region involved in sympathetic outflow, seems not to be involved in the generation of sympathetic tone in the normal state. The aim of this study was to determine the role of the RVLM glutamate receptors in the generation of sympathetic tone in CHF. CHF was produced by coronary artery ligation. Bilateral microinjection of the glutamate receptor antagonist kynurenic acid, the N-methyl-d-aspartate (NMDA) receptor antagonist D-2-amino-5-phosphonopentanoate, or the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione into the RVLM dose-dependently reduced resting blood pressure and renal sympathetic nerve activity in CHF but not in sham rats. Picoinjection of kynurenic acid (100 pmol in 5 nL) significantly decreased the basal discharge by 47% in 25 RVLM presympathetic neurons in CHF rats. In contrast, kynurenic acid had no effect on the discharge in all 22 of the RVLM presympathetic neurons tested in sham rats. These data suggest that upregulated glutamate receptors, including NMDA and non-NMDA, in the RVLM are involved in tonic control of elevated sympathetic tone in CHF.

NG2-glia are a substantial population of cells in the central nervous system (CNS) that can be identified by their specific expression of the NG2 chondroitin sulphate (CSPG). NG2-glia can generate oligodendrocytes, but it is unlikely this is their only function; indeed, they may be multipotent neural stem cells. Moreover, NG2-glia are a highly reactive cell type and a major function is to help form the axon growth inhibitory glial scar in response to CNS injury. The factors that regulate these diverse behaviours of NG2-glia are not fully resolved, but NG2-glia express receptors to the neurotransmitter glutamate, which has known potent effects on other glia. Here, we have examined the actions of glutamate receptor activation on NG2-glia in the rat optic nerve, a typical CNS white matter tract that does not contain neuronal cell bodies. Glutamate induces an increase in [Ca(2+)](i) in immuno-identified NG2-glia in situ and in vitro. In addition, we examined the effects of glutamate receptor activation in vivo by focal injection of the glutamate receptor agonist kainate into the optic nerve; saline was injected in controls. Changes in glial and axonal function were determined at 7 days post injection (dpi), by immunohistochemistry and electrophysiological measurement of the compound action potential (CAP). Injection of kainate resulted in a highly localized 'injury response' in NG2-glia, marked by dense labelling for NG2 at the lesion site, as compared to astrocytes, which displayed a more extensive reactive astrogliosis. Furthermore, injection of kainate resulted in an axonal conduction block. These glial and axonal changes were not observed following injection of saline vehicle. In addition, we provide evidence that endogenous glutamate induces calcium-dependent phosphorylation of extracellular signal-regulated kinases (ERK1/2), which may provide a potential mechanism by which glutamate-mediated changes in raised intracellular calcium could regulate the observed gliosis. The results provide evidence that activation of AMPA-kainate type ionotropic glutamate receptors evoke raised calcium in NG2-glia and induces an injury response in NG2-glia.

Withdrawal (WD) anxiety is a significant factor contributing to continued alcohol abuse in alcoholics. This anxiety is extensive, long-lasting, and develops well after the obvious physical symptoms of acute WD. The neurobiological mechanisms underlying this prolonged WD-induced anxiety are not well understood. The basolateral amygdala (BLA) is a major emotional center in the brain and regulates the expression of anxiety. New evidence suggests that increased glutamatergic function in the BLA may contribute to WD-related anxiety following chronic ethanol exposure. Recent evidence also suggests that kainate-type ionotropic glutamate receptors are inhibited by intoxicating concentrations of acute ethanol. This acute sensitivity suggests potential (KA-R) contributions by these receptors to the increased glutamatergic function seen during chronic exposure. Therefore, we examined the effect of chronic intermittent ethanol (CIE) and WD on KA-R-mediated synaptic transmission in the BLA of Sprague-Dawley rats. Our study showed that CIE, but not WD, increased synaptic responses mediated by KA-Rs. Interestingly, both CIE and WD occluded KA-R-mediated synaptic plasticity. Finally, we found that BLA field excitatory postsynaptic potential responses were increased during CIE and WD via a mechanism that is independent of glutamate release from presynaptic terminals. Taken together, these data suggest that KA-Rs might contribute to postsynaptic increases in glutamatergic synaptic transmission during CIE and that the mechanisms responsible for the expression of KA-R-dependent synaptic plasticity might be engaged by chronic ethanol exposure and WD.

Chronic intermittent hypoxia (CIH), as occurs in sleep apnea, impairs baroreflex-mediated reductions in heart rate (HR) and enhances HR responses to electrical stimulation of vagal efferent. We tested the hypotheses that HR responses to activation of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and N-methyl-D-aspartate (NMDA) receptors in the nucleus ambiguous (NA) are reduced in CIH-exposed rats and that this impairment is associated with degeneration of glutamate receptor (GluR)-immunoreactive NA neurons. Fischer 344 rats (3-4 mo) were exposed to room air (RA) or CIH for 35-50 days (n = 18/group). At the end of the exposures, AMPA (4 pmol, 20 nl) and NMDA (80 pmol, 20 nl) were microinjected into the same location of the left NA (-200 microm to +200 microm relative to caudal end of area postrema; n = 6/group), and HR and arterial blood pressure responses were measured. In addition, brain stem sections at the level of -800, -400, 0, +400, and +800 microm relative to obex were processed for AMPA and NMDA receptor immunohistochemistry. The number of NA neurons expressing AMPA receptors and NMDA receptors (NMDARs) was quantified. Compared with RA, we found that after CIH 1) HR responses to microinjection of AMPA into the left NA were reduced (RA -290 +/- 30 vs. CIH -227 +/- 15 beats/min, P < 0.05); 2) HR responses to microinjection of NMDA into the left NA were reduced (RA -302 +/- 16 vs. CIH -238 +/- 27 beats/min, P < 0.05); and 3) the number of NMDAR1, AMPA GluR1, and AMPA GluR2/3-immunoreactive cells in the NA was reduced (P < 0.05). These results suggest that degeneration of NA neurons expressing GluRs contributes to impaired baroreflex control of HR in rats exposed to CIH.

We have examined the effect of the peripheral application of glutamate and capsaicin to deep craniofacial tissues in influencing the activation and peripheral sensitization of deep craniofacial nociceptive afferents. The activity of single trigeminal nociceptive afferents with receptive fields in deep craniofacial tissues were recorded extracellularly in 55 halothane-anesthetized rats. The mechanical activation threshold (MAT) of each afferent was assessed before and after injection of 0.5 M glutamate (or vehicle) and 1% capsaicin (or vehicle) into the receptive field. A total of 68 afferents that could be activated by blunt noxious mechanical stimulation of the deep craniofacial tissues (23 masseter, 5 temporalis, 40 temporomandibular joint) were studied. When injected alone, glutamate and capsaicin activated and induced peripheral sensitization reflected as MAT reduction in many afferents. Following glutamate injection, capsaicin-evoked activity was greater than that evoked by capsaicin alone, whereas following capsaicin injection, glutamate-evoked responses were similar to glutamate alone. These findings indicate that peripheral application of glutamate or capsaicin may activate or induce peripheral sensitization in a subpopulation of trigeminal nociceptive afferents innervating deep craniofacial tissues, as reflected in changes in MAT and other afferent response properties. The data further suggest that peripheral glutamate and capsaicin receptor mechanisms may interact to modulate the activation and peripheral sensitization in some deep craniofacial nociceptive afferents.

Previous studies indicated that a long-term decrease in the activity of ionotropic glutamate receptors induces cholinergic activity in rat and mouse hypothalamic neuronal cultures. Here we studied whether a prolonged inactivation of ionotropic glutamate receptors also induces cholinergic activity in hippocampal neurons. Receptor activity was chronically suppressed in rat hippocampal primary neuronal cultures with two proportionally increasing sets of concentrations of NMDA plus non-NMDA receptor antagonists: 100 microM/10 microM AP5/CNQX (1X cultures) and 200 microM/20 microM AP5/CNQX (2X cultures). Using calcium imaging we demonstrate that cholinergic activity does not develop in these cultures. Instead, network-driven glutamate-dependent activity, that normally is detected in hyper-excitable conditions, reappears in each culture group in the presence of these antagonists and can be reversibly suppressed by higher concentrations of AP5/CNQX. This activity is mediated by non-NMDA receptors and is modulated by NMDA receptors. Further, non-NMDA receptors, the general level of glutamate receptor activity and CaMK-dependent signaling are critical for development of this network-driven glutamatergic activity in the presence of receptor antagonists. Using electrophysiology, western blotting and calcium imaging we show that some neuronal parameters are either reduced or not affected by chronic glutamate receptor blockade. However, other parameters (including neuronal excitability, mEPSC frequency, and expression of GluR1, NR1 and betaCaMKII) become up-regulated and, in some cases, proportionally between the non-treated, 1X and 2X cultures. Our data suggest recovery of the network-driven glutamatergic activity after chronic glutamate receptor blockade. This recovery may represent a form of neuronal plasticity that compensates for the prolonged suppression of the activity of glutamate receptors.

The design and synthesis of four pyrrolidine scaffolds that are structurally related to the known ionotropic glutamate receptor antagonist, (-)-kaitocephalin, is described. Additionally, preliminary results of the biological evaluation of these compounds are disclosed.

Positive and negative reinforcing systems are part of the mechanism of drug dependence. Drugs with abuse potential may change the manner of response to negative emotional stimuli, activate positive emotional reactions and possess primary reinforcing properties. Catecholaminergic and peptidergic processes are of importance in these mechanisms. Current research needs to understand the types of adaptations that underlie the particularly long-lived aspects of addiction. Presently, glutamate is candidate to play a role in the enduring effects of drugs of abuse. For example, it participates in the chronic pathological changes of corticostriatal terminals produced by methamphetamine. At the synaptic level, a link between over-activation of glutamate receptors, [C(a2+)](i) increase and neuronal damage has been clearly established leading to neurodegeneration. Thus, neurodegeneration can start after an acute over-stimulation whose immediate effects depend on a diversity of calcium-activated mechanisms. If sufficient, the initial insult results in calcification and activation of a chronic on-going process with a progressive loss of neurons. At present, long-term effects of drug dependence underlie an excitotoxicity process linked to a polysynaptic pathway that dynamically regulates synaptic glutamate. Retaliatory mechanisms include energy capability of the neurons, inhibitory systems and cytoplasmic calcium precipitation as part of the neuron-glia interactions. This paper presents an integrated view of these molecular and cellular mechanisms to help understand their relationship and interdependence in a chronic pathological process that suggest new targets for therapeutic intervention.

It has been proposed that there is an "apparent monosynaptic" connection between gastric vagal afferent nerve terminals and inhibitory projection neurons in the nucleus tractus solitarius (NTS) and that two efferent parallel pathways from the dorsal motor nucleus of the vagus (DMV) influence peripheral organs associated with these reflexes (6). The purpose of our study was to verify the validity of these views as they relate to basal control of gastric motility. To test the validity of a direct connection of vagal afferent terminals (known to release l-glutamate) directly impacting second-order projection neurons, we evaluated the effect of GABA(A) receptor blockade in the area of the medial subnucleus of the tractus solitarius (mNTS) on gastric motility. Microinjection of bicuculline methiodide into the mNTS produced robust decreases in gastric motility (-1.6 +/- 0.2 mmHg, P < 0.05, n = 23), which were prevented by cervical vagotomy and by pretreatment with kynurenic acid microinjected into the mNTS. Kynurenic acid per se had no effect on gastric motility. However, after GABA(A) receptor blockade in the mNTS, kynurenic acid produced a robust increase in gastric motility. To test for the contribution of two parallel efferent DMV pathways, we assessed the effect of either intravenous atropine methylbromide or N(G)-nitro-l-arginine methyl ester on baseline motility and on decreases in gastric motility induced by GABA(A) receptor blockade in the mNTS. Only atropine methylbromide altered baseline motility and prevented the effects of GABA(A) receptor blockade on gastric motility. Our data demonstrate the presence of intra-NTS GABAergic signaling between the vagal afferent nerve terminals and inhibitory projection neurons in the NTS and that the cholinergic-cholinergic excitatory pathway comprises the functionally relevant efferent arm of the vagovagal circuit.

We used long-range organotypic cultures of auditory nuclei in the chick hindbrain to test the development of glutamate receptor activity in auditory neurons growing in a tissue environment that includes early deprivation of peripheral glutamatergic input, subsequent to removal of the otocyst. Cultures started at embryonic day (E)5, and lasted from 6 h to 15 days. Neuronal migration, clustering and axonal extension from the nucleus magnocellularis (NM) to the nucleus laminaris (NL) partially resembled events in vivo. However, the distinctive laminar organization of the NL was not observed. Glutamate receptor (GluR) activity was tested with optical recordings of intracellular Ca2+ in the NM. alpha-Amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA)/kainate receptors had Ca2+ responses with a time course similar to that in control slices. Peak amplitude, however, was significantly lower. N-methyl-D-aspartate (NMDA)-mediated Ca2+ responses were higher in 2-day cultures (E5 + 2d) than in E7 explant controls, returning later to control values. Metabotropic GluRs did not elicit Ca2+ responses at standard agonist doses. Blocking NMDA or AMPA/kainate receptors with specific antagonists for 10 days in culture did not limit neuronal survival. Blocking metabotropic GluRs resulted in complete neuronal loss. Thus, ionotropic GluRs are not required for NM neuronal survival. However, their activity during development is affected when neurons grow in an in vitro environment that includes prevention of arrival of peripheral glutamatergic input.

Acylpolyamine-type spider toxins are known to be potent and specific blockers against glutamate receptors (GluRs). The present study describes the syntheses and biological activities of several fluorescent-labeled analogs related to a Madagascar Joro spider toxin NPTX-594 to analyze visually the unknown interaction between spider toxins and GluRs.

The present study investigates changes in red nucleus (RN) neuronal activity and the role of glutamate receptors (GluRs) after simulated microgravity (tail-suspension) in the rat using single-unit recording and microinjection. The results showed that tail-suspension for 3, 7, and 14 days could induce a significant decrease in spontaneous firing rate of RN neurons in a time-dependent manner. Unilateral microinjection of glutamate into the RN significantly increased the firing rate of RN neurons, but the increased firing rate was significantly reduced following tail-suspension time. Microinjection of the NMDA receptor antagonist MK-801 or the non-NMDA receptor antagonist DNQX into the RN blocked this excitatory effect induced by glutamate. However, microinjection of the metabotropic glutamate receptor (mGluR) antagonist (+/-)-MCPG into the RN had no effect. These results suggest that simulated microgravity can reduce excitability of RN neurons following a functional impairment of glutamate receptors. NMDA and non-NMDA receptors, but not mGluRs, are involved in the mediation of glutamate-evoked excitation of RN neurons. The decrease in excitability of RN neurons may be involved in simulated microgravity-induced muscle atrophy.

It is becoming clear that astrocytes are active participants in synaptic functioning and exhibit properties, such as the secretion of classical transmitters, previously thought to be exclusively neuronal. Whether these similarities extend to the release of neuropeptides, the other major class of transmitters, is less clear. Here we show that cortical astrocytes can synthesize both native and foreign neuropeptides and can secrete them in a stimulation-dependent manner. Reverse transcription-PCR and mass spectrometry indicate that cortical astrocytes contain neuropeptide Y (NPY), a widespread neuronal transmitter. Immunocytochemical studies reveal NPY-immunoreactive (IR) puncta that colocalize with markers of the regulated secretory pathway. These NPY-IR puncta are distinct from the synaptic-like vesicles that contain classical transmitters, and the two types of organelles are differentially distributed. After activation of metabotropic glutamate receptors and the release of calcium from intracellular stores, the NPY-IR puncta fuse with the cell membrane, and the peptide-containing dense cores are displayed. To determine whether peptide secretion subsequently occurred, exocytosis was monitored from astrocytes expressing NPY-red fluorescent protein (RFP). In live cells, after activation of glutamate receptors, the intensity of the NPY-RFP-labeled puncta declined in a step-like manner indicating a regulated release of the granular contents. Because NPY is a widespread and potent regulator of synaptic transmission, these results suggest that astrocytes could play a role in the peptidergic modulation of synaptic signaling in the CNS.

Repeated cocaine administration induces numerous neuroadaptations. Cocaine controls many neurotransmitter systems, including glutamate, GABA, dopamine, serotonin, and noradrenalin. Cocaine exposure modifies neuron morphology. Peptide systems such as the orexins probably play a key role in cocaine's long-term effects. The opioid system may be involved in cocaine's long-term effects. The complexity and the multiplicity of the neuroadaptations observed with cocaine may explain the current absence of effective pharmacotherapy.

The aim of the present study was to determine whether various glutamate receptor antagonists could affect ethanol withdrawal-induced anxiety-like behavior measured in the elevated plus-maze test in rats. In our study, memantine (8 and 12 mg/kg), a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist, did not show any effect on ethanol withdrawal anxiety. Acamprosate (NMDA and metabotropic glutamate5 (mGlu5) receptor antagonist), at a dose of 400 mg/kg showed anxiolytic-like effect, thus increasing the percent of time spent in open arms and open arms entries. Antagonists of group I mGlu receptors, such as MTEP ([(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine, mGlu5 receptor) or EMQMCM (3-ethyl-2-methyl-quinolin-6-yl-(4-methoxy-cyclohexyl)-methanone methanesulfonate, mGlu1 receptor), caused similar effects to acamprosate. In contrast to acamprosate and MTEP, EMQMCM (5 mg/kg) elevated the ethanol withdrawal-induced decrease in locomotion. When given alone to the saline-treated group, EMQMCM indicated anxiolytic-like effect. Our results imply a crucial role of mGlu5 receptor in an anxiety-like effect of ethanol withdrawal because MTEP (a selective mGlu5 receptor antagonist) and acamprosate (which also indirectly inhibits mGlu5 receptor) attenuated ethanol withdrawal anxiety-like behavior without influence on ethanol withdrawal hypolocomotion and did not show any effect in the saline-treated groups. However, difference in anxiolytic-like potency between both these group I mGlu receptors antagonists may be due to the recent experimental design. Therefore, taking into account a positive correlation between ethanol withdrawal-induced anxiety and relapse to ethanol drinking, our results suggest that mGlu receptor antagonists of group I (similarly to acamprosate) could prevent relapse to drinking and, therefore they might be useful in therapy of alcoholism.

Noradrenaline is known to modulate memory formation in the mammalian hippocampus. We have examined how noradrenaline and selective beta-adrenoceptor (AR) agonists affect memory consolidation and how antagonists inhibit memory consolidation in the avian hippocampus. Injection of selective beta-AR agonists and antagonists at specific times within 30 min of a weakly or strongly reinforced, single-trial, bead discrimination learning test in 1-day-old chicks allowed us to determine the pattern of beta-AR involvement in hippocampal memory processing. Different beta-AR subtypes were recruited in temporal sequence after learning in the order beta(1), beta(3), and beta(2.) We provide evidence that the effect of manipulation of beta(1)-ARs by selective agonists and antagonists within 2.5 min of training parallels the action of NMDA receptor agonists and antagonists. Activation of beta(3)- and beta(2)-ARs facilitated memory but utilized different mechanisms: beta(3)-ARs by stimulating glucose uptake and metabolism, and beta(2)-ARs by increasing the breakdown of glycogen--with both metabolic events occurring in astrocytes and affecting intermediate memory. The different receptors are activated at different times within the lifetime of labile memory and within 30 min of learning. We have defined separate roles for the three beta-ARs in memory and demonstrated that the avian hippocampus is involved in learning and memory in much the same way as the hippocampus in the mammalian brain.

Although the function of the kainate receptors in the brain is still not clear, they are increasingly defined as targets in the development of new classes of anti-epileptics. The thienopyrimidines described in this report were tested for their antagonistic effect at the kainate receptor subtypes GluR5 and GluR6. The highest effectiveness was obtained by a 4-ethoxy-thieno[2,3-d]pyrimidin with an IC50 = 68 microM at the GluR6 receptor.

Considerable research into the neurobiology of cocaine addiction has shed light on the role of glutamate. Findings from models of relapse to cocaine-seeking indicate that the glutamatergic system is critically involved, as glutamate levels in the nucleus accumbens increase during reinstatement and glutamate receptor activation is necessary for reinstatement to drug-seeking. Thus, it would seem beneficial to block the increased glutamate release, but full antagonists of ionotropic glutamate receptors produce undesirable side effects. Therefore, modulation of glutamatergic transmission would be advantageous and provide novel pharmacotherapeutic avenues. Pharmacotherapies have been developed that have the potential to modulate excessive glutamatergic transmission through ionotropic and metabotropic (mGluR) glutamate receptors. Compounds that modulate glutamatergic transmission through ionotropic glutamate receptors include the non-competitive N-methyl-D-aspartic acid antagonists, amantadine and memantine, and the partial N-methyl-D-aspartic acid agonist d-cycloserine. They have shown promise in preclinical models of cocaine addiction. The mGluR2/3 agonist LY379268 is effective in inhibiting cocaine seeking in preclinical animal models and could decrease stress-induced relapse due to its anxiolytic effects. Similarly, the mGluR1/5 antagonists, 2-methyl-6-(phenylethynyl)pyridine and 3-[2-methyl-4-thiazolyl)ethynyl]pyridine, have shown to be effective in preclinical models of cocaine addiction. The cysteine pro-drug, N-acetylcysteine, restores the inhibitory tone on presynaptic glutamate receptors and has been effective in reducing cue-induced craving and cocaine use in humans. Furthermore, anticonvulsants, such as topiramate or lamotrigine, have shown efficacy in treating cocaine dependence or reducing relapse in humans. Future pharmacotherapy may focus on manipulating signal transduction proteins and pathways, which include Homer/N-methyl-D-aspartic acid complexes, to provide effective treatment for cocaine addiction.

Ethnopharmacological Relevance: Searsia dentata and Searsia pyroides are used in traditional South African medicine to treat convulsions and epilepsy. Previous studies have demonstrated that extracts of these plants comprise compounds that bind to the flumazenil-sensitive site on the GABA(A) receptor. However, their use as anticonvulsant medicinal plants cannot be adequately explained by these findings., Aims: The aim of this study was to examine the possible involvement of the glutamatergic system of extracts from the plants., Materials and Methods: The mouse cortical wedge preparation was used for functional characterization of the extracts. The affinity towards the NMDA and the AMPA receptor was investigated using classical [(3)H]-GP39653 and [(3)H]-AMPA binding assays, respectively., Results: The extracts of Searsia dentata and Searsia pyroides inhibited the spontaneous epileptiform discharges in mouse cerebral cortical slices with ED(50) of 0.62 and 1.67mg dry extract/mL, respectively. Both extracts displaced [(3)H]-GP39653 binding and significantly inhibited the NMDA-induced response during co-administration in cortical slices., Conclusion: In this study, the NMDA receptor antagonistic effect of the crude ethanolic extracts of these two South African medicinal plants was demonstrated.

L-Glutamate and 5'-ribonucleotides such as guanosine-5'-monophosphate (GMP) and inosine-5'-monophosphate (IMP) elicit a unique taste called 'umami' that is distinct from the tastes of sweet, salty, sour, and bitter. For umami, like sweet and bitter compounds, taste signaling is initiated by binding of tastants to G-protein-coupled receptors (GPCR) in taste bud cells. To date, several GPCRs for umami compounds have been identified in taste cells, including the heterodimer T1R1/T1R3, and truncated type 1 and 4 metabotropic glutamate receptors missing most of the N-terminal extracellular domain (taste-mGluR4 and truncated-mGluR1). Apparently contradictory data in T1R3 knock-out (KO) mouse models have been reported. One study showed that behavioral preference and taste nerve responses to umami stimuli in T1R3-KO mice were totally abolished, suggesting that T1R1/T1R3 is a sole receptor for umami taste. The other reported reduced but not abolished responses to umami in T1R3-KO mice, suggesting existence of multiple receptors for umami taste. In this paper, we summarized the data from recent studies that further addressed this issue by using different experimental techniques. Some of the studies provided additional evidence for the existence of umami receptor systems mediated by mGluR1 and mGluR4 in addition to T1R1/T1R3. It is proposed that the signal mediated by the pathway involving T1R1/T1R3 may play a different role from that derived from the mGluRs. The former occurs mainly in the anterior tongue, and plays a major role in preference behavior, whereas the latter occurs mainly in the posterior tongue and contributes to behavioral discrimination between umami and other taste compounds.

The supratrigeminal region (SupV) receives abundant orofacial sensory inputs and descending inputs from the cortical masticatory area and contains premotor neurons that target the trigeminal motor nucleus (MoV). Thus it is possible that the SupV is involved in controlling jaw muscle activity via sensory inputs during mastication. We used voltage-sensitive dye, laser photostimulation, patch-clamp recordings, and intracellular biocytin labeling to investigate synaptic transmission from the SupV to jaw-closing and jaw-opening motoneurons in the MoV in brain stem slice preparations from developing rats. Electrical stimulation of the SupV evoked optical responses in the MoV. An antidromic optical response was evoked in the SupV by MoV stimulation, whereas synaptic transmission was suppressed by substitution of external Ca2+ with Mn2+. Photostimulation of the SupV with caged glutamate evoked rapid inward currents in the trigeminal motoneurons. Gramicidin-perforated and whole cell patch-clamp recordings from masseter motoneurons (MMNs) and digastric motoneurons (DMNs) revealed that glycinergic and GABAergic postsynaptic responses evoked in MMNs and DMNs by SupV stimulation were excitatory in P1-P4 neonatal rats and inhibitory in P9-P12 juvenile rats, whereas glutamatergic postsynaptic responses evoked by SupV stimulation were excitatory in both neonates and juveniles. Furthermore, the axons of biocytin-labeled SupV neurons that were antidromically activated by MoV stimulation terminated in the MoV. Our results suggest that inputs from the SupV excite MMNs and DMNs through activation of glutamate, glycine, and GABAA receptors in neonates, whereas glycinergic and GABAergic inputs from the SupV inhibit MMNs and DMNs in juveniles.

This study was designed to delineate the medullary and spinal pathways mediating the cardiovascular responses to cold pressor test (CPT) and to identify neurotransmitters in these pathways. Experiments were done in barodenervated, urethane-anesthetized, male Wistar rats. The CPT was performed by immersing the limbs and ventral half of the body of the rat in ice-cold water (0.5 degrees C) for 2 min. CPT elicited an immediate increase in mean arterial pressure (MAP), heart rate (HR), and greater splanchnic nerve activity (GSNA). Bilateral blockade of ionotropic glutamate receptors (iGLURs) in the rostral ventrolateral medullary pressor area (RVLM) significantly attenuated the CPT-induced responses. Bilateral blockade of gamma-aminobutyric acid (GABA) receptors, but not iGLURs, in the nucleus ambiguus (nAmb) significantly reduced the CPT-induced increases in HR, but not MAP. Blockade of spinal iGLURs caused a significant reduction in CPT-induced increases in MAP and GSNA, whereas the increases in HR were reduced to a lesser extent. Combination of the blockade of spinal iGLURs and bilateral vagotomy or intravenous atropine almost completely blocked CPT-induced tachycardia. Midcollicular decerebration significantly reduced CPT-induced increases in MAP and HR. These results indicated that: 1) CPT-induced increases in MAP, HR, and GSNA were mediated by activation of iGLURs in the RVLM and spinal cord, 2) activation of GABA receptors in the nAmb also contributed to the CPT-induced tachycardic responses, and 3) brain areas rostral to the brain stem also participated in the CPT-induced pressor and tachycardic responses.

Dark neurons have plagued the interpretation of brain tissue sections, experimentally and clinically. Seen only when perturbed but living tissue is fixed in aldehydes, their mechanism of production is unknown. Since dark neurons are seen in cortical biopsies, experimental ischemia, hypoglycemia, and epilepsy, we surmised that glutamate release and neuronal transmembrane ion fluxes could be the perturbation leading to dark neuron formation while the fixation process is underway. Accordingly, we excised biopsies of rat cortex to simulate neurosurgical production of dark neurons. To ascertain the role of glutamate, blockade of N-methyl-D-aspartate (NMDA) and non-NMDA receptors was done prior to formaldehyde fixation. To assess the role of transmembrane sodium ion (and implicitly, water) fluxes, tetraethylammonium (TEA) was used. Blockade of NMDA receptors with MK-801 and non-NMDA receptors with the quinoxalinediones (CNQX and NBQX) abolished dark neuron formation. More delayed exposure of the tissue to the antagonist, CNQX, by admixing it with the fixative directly, allowed for some production of dark neurons. Aminophosphonoheptanoate (APH), perhaps due to its polarity, and TEA, did not prevent dark neurons, which were abundant in control formaldehyde fixed material unexposed to either receptor or ion channel antagonists. The results demonstrate a role for the pharmacologic subtypes of glutamate receptors in the pathogenetic mechanism of dark neuron formation. Our results are consistent with the appearance of dark neurons in biopsy where the cerebral cortex has been undercut, and rendered locally ischemic and hypoglycemic, as well as in epilepsy, hypoglycemia, and ischemia, all of which lead to glutamate release. Rather than a pressure-derived mechanical origin, we suggest that depolarization, glutamate release or receptor activation are more likely mechanisms of dark neuron production.

Obsessive-compulsive disorder (OCD) is a frequent disorder and glutamate seems to be an important transmitter. The effect of glutamate depends on the level as well as the effect mediated through glutamate receptors. The level is altered in brain areas that are involved in OCD. Genes encoding receptors, enzymes involved in the metabolism and transport proteins are located in regions that are related to OCD. Pharmacogenetic studies examining the genetic variations in the glutaminate system are lacking. The aim was to examine the current knowledge on the glutaminate system in the development and maintenance of OCD.

The anxiolytic-like activity of magnesium in mice during the elevated plus maze (EPM) has been demonstrated previously. In the present study, we examined the involvement of NMDA/glutamate receptor ligands on the magnesium effect on the EPM. We demonstrated that low, ineffective doses of NMDA antagonists (the competitive NMDA antagonist CGP 37849, 0.3 mg/kg; an antagonist of the glycineB sites, L-701,324, 1 mg/kg; a partial agonist of the glycineB sites, D-cycloserine, 2.5 mg/kg; and the non-competitive NMDA antagonist MK-801, 0.05 mg/kg) administered together with an ineffective dose of magnesium (10 mg/kg) evoked a significant increase in the percentage of time spent in the open arm of the maze (an index of anxiety). Moreover, magnesium-induced anxiolytic-like activity (20 mg/kg) was antagonized by D-serine (100 nmol/mouse), an agonist of glycineB site of the NMDA receptor complex. The present study demonstrates the involvement of the NMDA/glutamate pathway in the magnesium anxiolytic-like activity in the EPM in mice, and that this activity particularly involves the glycineB sites.

The clinical efficacy of current antidepressant therapies is unsatisfactory; antidepressants induce a variety of unwanted effects, and, moreover, their therapeutic mechanism is not clearly understood. Thus, a search for better and safer agents is continuously in progress. Recently, studies have demonstrated that zinc and magnesium possess antidepressant properties. Zinc and magnesium exhibit antidepressant-like activity in a variety of tests and models in laboratory animals. They are active in forced swim and tail suspension tests in mice and rats, and, furthermore, they enhance the activity of conventional antidepressants (e.g., imipramine and citalopram). Zinc demonstrates activity in the olfactory bulbectomy, chronic mild and chronic unpredictable stress models in rats, while magnesium is active in stress-induced depression-like behavior in mice. Clinical studies demonstrate that the efficacy of pharmacotherapy is enhanced by supplementation with zinc and magnesium. The antidepressant mechanisms of zinc and magnesium are discussed in the context of glutamate, brain-derived neurotrophic factor (BDNF) and glycogen synthase kinase-3 (GSK-3) hypotheses. All the available data indicate the importance of zinc and magnesium homeostasis in the psychopathology and therapy of affective disorders.

Liberation of zinc from intracellular stores contributes to oxidant-induced neuronal injury. However, little is known regarding how endogenous oxidant systems regulate intracellular free zinc ([Zn(2+)](i)). Here we simultaneously imaged [Ca(2+)](i) and [Zn(2+)](i) to study acute [Zn(2+)](i) changes in cultured rat forebrain neurons after glutamate receptor activation. Neurons were loaded with fura-2FF and FluoZin-3 to follow [Ca(2+)](i) and [Zn(2+)](i), respectively. Neurons treated with glutamate (100 microM) for 10 min gave large Ca(2+) responses that did not recover after termination of the glutamate stimulus. Glutamate also increased [Zn(2+)](i), however glutamate-induced [Zn(2+)](i) changes were completely dependent on Ca(2+) entry, appeared to arise entirely from internal stores, and were substantially reduced by co-application of the membrane-permeant chelator TPEN during the glutamate treatment. Pharmacological maneuvers revealed that a number of endogenous oxidant producing systems, including nitric oxide synthase, phospholipase A(2), and mitochondria all contributed to glutamate-induced [Zn(2+)](i) changes. We found no evidence that mitochondria buffered [Zn(2+)](i) during acute glutamate receptor activation. We conclude that glutamate-induced [Zn(2+)](i) transients are caused in part by [Ca(2+)](i)-induced reactive oxygen species that arises from both cytosolic and mitochondrial sources.

In the present study, we describe GABAA receptor-mediated tonic inhibitory currents in the substantia gelatinosa (SG) region of rat spinal trigeminal nucleus pars caudalis (Vc). The GABA(A) receptor-mediated tonic currents were identified by bath-application of the GABAA receptor antagonists, picrotoxin (1mM), SR95531 (100microM) and bicuculline (100microM). All three antagonists completely blocked outward spontaneous (phasic) inhibitory postsynaptic currents, but only picrotoxin and bicuculline induced a significant (>5pA) inward shift of holding currents at a holding potential (Vh) of 0mV in 60-70% of SG neurons, revealing the existence of tonic outward currents. The tonic currents were resistant to further the blockades of glycine receptors or those in addition to glutamate receptors and voltage-dependent sodium channels. An acute bath-application of THDOC (0.1microM), the stress-related neurosteroid, did enhance tonic currents, but only in a small population of SG neurons. In addition, slices incubated with THDOC for 30min increased the probability of neurons with significant tonic currents. The GABAergic tonic inhibition demonstrated in this study may play a significant role in the sensory processing system of the Vc.

( S)-Glutamic acid (Glu) is the major excitatory neurotransmitter in the central nervous system (CNS) activating the plethora of ionotropic Glu receptors (iGluRs) and metabotropic Glu receptors (mGluRs). In this paper, we present a chemo-enzymatic strategy for the enantioselective synthesis of five new Glu analogues 2a- f ( 2d is exempt) holding a functionalized substituent in the 4-position. Nine Glu analogues 2a- j are characterized pharmacologically at native 2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA), kainic acid (KA), and N-methyl- d-aspartic acid (NMDA) receptors in rat synaptosomes as well as in binding assays at cloned rat iGluR5-7 subtypes. A detailed in silico study address as to why 2h is a high-affinity ligand at iGluR5-7 ( K i = 3.81, 123, 57.3 nM, respectively), while 2e is only a high affinity ligand at iGluR5 ( K i = 42.8 nM). Furthermore, a small series of commercially available iGluR ligands are characterized in iGluR5-7 binding.

The transmission of rhythmic drive to respiratory motoneurons in vitro is critically dependent on glutamate acting primarily on non-NMDA receptors. We determined whether both non-NMDA and NMDA receptors contribute to respiratory drive transmission at respiratory motoneurons in the intact organism, both in the state of anesthesia and in the same animals during natural behaviors. Twenty-seven rats were implanted with electroencephalogram and neck electrodes to record sleep-wake states and genioglossus and diaphragm electrodes for respiratory muscle recordings. Microdialysis probes were inserted into the hypoglossal motor nucleus (HMN). Under anesthesia, non-NMDA or NMDA receptor antagonism significantly decreased respiratory-related genioglossus activity, indicating a contribution of each receptor to respiratory drive transmission at the HMN. However, despite the presence of respiratory-related genioglossus activity in the same rats across sleep-wake states, neither non-NMDA receptor antagonism at the HMN nor glutamate uptake inhibition had any effect on respiratory-related genioglossus activity. These results showed that, compared with anesthesia, respiratory drive transmission through the non-NMDA receptor is low in the behaving organism. In contrast, glutamate uptake inhibition increased tonic genioglossus activity in wakefulness and non-rapid-eye-movement sleep, indicating a functional endogenous glutamatergic modulation of tonic, but not respiratory, motor tone. Such an effect on tonic drive may contribute to the suppression of both tonic and respiratory-related genioglossus activity in wakefulness and sleep with NMDA receptor antagonism at the HMN. These data do not refute previous identification of a glutamatergic (mostly non-NMDA receptor activating) respiratory drive to hypoglossal motoneurons, but this mechanism is more prominent in anesthetized or in vitro preparations.

Glutamate neurotoxicity is exacerbated when energy metabolism is impaired. In vitro studies show that neuronal death in these conditions is related to mitochondrial dysfunction, ATP depletion, and the loss of calcium homeostasis. We have recently observed that, in vivo, enhancement of glutamate toxicity elicited by previous mitochondrial inhibition does not involve severe ATP depletion, suggesting the involvement of other processes. Factors such as the activation of different proteases may determine the extent and type of cell death. Protease activation might be triggered by internal or external factors, such as mitochondrial damage or the activation of a particular glutamate receptor subtype. In the present study we aimed to investigate whether moderate inhibition of mitochondrial metabolism facilitates glutamate toxicity through caspase-3 or calpain activation, as well as the contribution of NMDA and non-NMDA glutamate ionotropic receptors to this activation. Rats were pre-treated with a subtoxic dose of 3-NP and 4 h later intrastriatally injected with glutamate. Results show that neither of these treatments alone (3-NP or Glu) or in combination (3-NP+Glu) activated caspase-3. Conversely, calpain activity is induced after glutamate injection both in intact and 3-NP pre-treated rats. Inhibition of calpain activity by MDL-28170 significantly prevented striatal damage. NMDA and non-NMDA receptors contributed equally to calpain activation and to the induction of neuronal death. Results suggest that enhancement of glutamate toxicity due to inhibition of mitochondrial metabolism in vivo, does not recruit caspase-dependent apoptosis but favors calpain activation through the stimulation of both subtypes of glutamate ionotropic receptors.

DPKQDFMRFamide, the most abundant FMRFamide-like peptide in Drosophila melanogaster, has been shown previously to enhance contractions of larval body wall muscles elicited by nerve stimulation and to increase excitatory junction potentials (EJPs). The present work investigated the possibility that this peptide can also stimulate muscle contraction by a direct action on muscle fibers. DPKQDFMRFamide induced slow contractions and increased tonus in body wall muscles of Drosophila larvae from which the central nervous system had been removed. The threshold for this effect was approximately 10(-8)M. The increase in tonus persisted in the presence of 7x10(-3)M glutamate, which desensitized postsynaptic glutamate receptors. Thus, the effect on tonus could not be explained by enhanced release of glutamate from synaptic terminals and, thus, may represent a postsynaptic effect. The effect on tonus was abolished in calcium-free saline and by treatment with L-type calcium channel blockers, nifedipine and nicardipine, but not by T-type blockers, amiloride and flunarizine. The present results provide evidence that this Drosophila peptide can act postsynaptically in addition to its apparent presynaptic effects, and that the postsynaptic effect requires influx through L-type calcium channels.

Clinical evidence demonstrates altered glutamatergic neurotransmission in patients suffering from obsessive-compulsive disorder (OCD). We examined the effects of glutamate-related drugs on marble-burying behavior, which is an animal model of OCD. The uncompetitive N-methyl-d-aspartate (NMDA) antagonists memantine (10 mg/kg, i.p.) and amantadine (30 mg/kg, i.p.) significantly inhibited marble-burying behavior without affecting locomotor activity in mice. Similarly, the uncompetitive NMDA receptor antagonist 5R,10S-(+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d)cyclohepten-5,10-imine hydrogen maleate (MK-801, 0.3 mg/kg, i.p.) inhibited marble-burying behavior. However, MK-801 at the same dose markedly increased locomotor activity. By contrast, the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor antagonist 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide (NBQX) and the glutamate release inhibitor riluzole showed no effect on marble-burying behavior and significant suppression of locomotor activity. MK-801 (0.3 mg/kg, i.p.) and memantine (10 mg/kg, i.p.) significantly disrupted prepulse inhibition as an operational measure of sensorimotor gating. By contrast, amantadine (30 mg/kg, i.p.) did not affect prepulse inhibition. These findings suggest that amantadine could be a useful drug for the treatment of OCD.

Opioid and excitatory amino acid receptors contribute to morphine dependence, but there are no studies of their role in heroin dependence. Thus, mice injected with acute or chronic heroin doses in the present study were pretreated with one of the following selective antagonists: 7-benzylidenenaltrexone (BNTX), naltriben (NTB), nor-binaltorphimine (nor-BNI; delta1, delta2, and kappa opioid receptors, respectively), MK-801, or LY293558 (NMDA and AMPA excitatory amino acid receptors, respectively). Naloxone-precipitated withdrawal jumping frequency, shown here to be a reliable index of heroin dependence magnitude, was reduced by BNTX or NTB in mice injected with both acute and chronic heroin doses. In contrast, nor-BNI did not alter jumping frequencies in mice injected with an acute heroin dose but significantly increased them in mice receiving chronic heroin injections. Continuous MK-801 or LY293558 infusion, but not injection, reduced jumping frequencies during withdrawal from acute heroin treatment. Their delivery by injection was nonetheless effective against chronic heroin dependence, suggesting mechanisms not simply attributable to NMDA or AMPA blockade. These data indicate that whereas delta1, delta2, NMDA, and AMPA receptors enable acute and chronic heroin dependence, kappa receptor activity limits the dependence liability of chronic heroin. With the exception of delta1 receptors, the apparent role of these receptors to heroin dependence is consistent with their contribution to morphine dependence, indicating that there is substantial physiological commonality underlying dependence to both heroin and morphine. The ability of kappa receptor blockade to differentially alter acute and chronic dependence supports previous assertions from studies with other opioids that acute and chronic opioid dependence are, at least in part, mechanistically distinct. Elucidating the substrates contributing to heroin dependence, and identifying their similarities and differences with those of other opioids such as morphine, may yield effective treatment strategies to the problem of heroin dependency.

Experiments were done in urethane-anesthetized, barodenervated, male Wistar rats. Chemical stimulation of the hypothalamic paraventricular nucleus (PVN) by unilateral microinjections of N-methyl-D-aspartic acid (NMDA) elicited increases in mean arterial pressure (MAP) and greater splanchnic nerve activity (GSNA). The increases in the MAP and GSNA induced by chemical stimulation of the PVN were significantly exaggerated by bilateral microinjections of D(-)-2-amino-7-phosphono-heptanoic acid (D-AP7) and 2,3-dioxo-6-nitro-1,2,3,4-tetrahydro-benzo[f]quinoxaline-7-sulfonamide disodium (NBQX) (ionotropic glutamate receptor antagonists) into the medial subnucleus of the nucleus tractus solitarius (mNTS). These results were confirmed by single unit recordings; i.e. excitation of mNTS barosensitive neurons caused by chemical stimulation of the ipsilateral PVN was blocked by application of D-AP7 and NBQX to these neurons. Bilateral microinjections of D-AP7 and NBQX into the mNTS elicited pressor responses which were significantly attenuated by inhibition of PVN neurons by bilateral microinjections of muscimol. Unilateral microinjections of fluorogold into the mNTS resulted in bilateral retrograde labeling of the PVN neurons. Unilateral microinjections of biotinylated dextran amine into the PVN resulted in anterograde labeling of axons and terminals in the mNTS bilaterally and the labeled terminals exhibited vesicular glutamate transporter-2 immunoreactivity. These results indicated that 1) a tonically active glutamatergic bilateral projection from the PVN to the mNTS exists; 2) bilateral blockade of ionotropic glutamate receptors in the mNTS exaggerates the increases in MAP and GSNA, but not heart rate, to the chemical stimulation of the PVN; and 3) this projection may serve as a restraint mechanism for excitatory cardiovascular effects of PVN stimulation.