Adenosine A2A receptors inhibit the N-methyl-D-aspartate component of excitatory synaptic currents in rat striatal neurons

The effects of the adenosine A(2A) receptor agonist 2-p-(2-carboxyethyl)phenethyl-amino-5'-N-ethylcarboxamidoadenosine (CGS 21680) on currents mediated by excitatory amino acid receptors were examined in rat striatal brain slices. In a Mg(2+)-free superfusion medium, CGS 21680 decreased the amplitude of excitatory postsynaptic currents (EPSCs) in about 70% of striatal neurons. The inhibitory effect of CGS 21680 disappeared both in the presence of the adenosine A(2A) receptor antagonist 8-(3-chlorostyryl) caffeine and the NMDA receptor antagonist 2-amino-5-phosphonopentanoic acid (AP-5). NMDA-induced currents were also depressed by CGS 21680 in a subset of striatal cells, whereas alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA)-induced currents were not affected. The results suggest that adenosine A(2A) receptor agonists inhibit the NMDA component of the EPSC.     

Blockade of glutamate transporters leads to potentiation of NMDA receptor current in layer V pyramidal neurons of the rat prefrontal cortex via group II metabotropic glutamate receptor activation

Membrane currents of layer V pyramidal cells in slices of the rat prefrontal cortex (PFC) were recorded with the patch-clamp technique. In an Mg(2+)-free superfusion medium l-trans-pyrrolidine-2,4-dicarboxylic acid (PDC), a preferential blocker of astrocytic glutamate transporters, caused inward current due to the activation of NMDA receptors. The blockade of conducted action potentials by tetrodotoxin did not interfere with this effect. ATP was inactive when given alone and potentiated the NMDA-induced current in an Mg(2+)-containing but not Mg(2+)-free superfusion medium. Agonists of group I ((S)-3,5-dihydroxyphenylglycine; DHPG) and II ((1R,4R,5S,6R)-4-amino-2-oxabicyclo[3.1.0]hexane-4,6-dicarboxylic acid; LY 379268) metabotropic glutamate receptors (mGluRs) also potentiated responses to NMDA, whereas the group III mGluR agonist l-(+)-2-amino-4-phosphonobutyric acid (l-AP4) did not affect them. In contrast to ATP, PDC evoked inward current in the absence but not in the presence of external Mg(2+), when given alone, and facilitated the NMDA effect Mg(2+)-independently. The PDC-induced facilitation of NMDA responses was blocked by group II ((2S)-2-amino-2-[(1S,2S)-2-carboxycycloprop-1-yl]-3-(xanth-9-yl) propanoic acid; LY 341495), but not group I ((RS)-1-aminoindan-1,5-dicarboxylic acid; AIDA) or III (alpha-methyl-3-methyl-4-phosphonophenylglycine; UBP 1112) mGluR antagonists. In conclusion, the blockade of astrocytic glutamate uptake by PDC may lead to a stimulation of group II mGluRs, while the triggering of exocytotic glutamate release from astrocytes by ATP may cause activation of group I mGluRs, both situated postsynaptically at layer V PFC pyramidal cells. Either group of mGluRs may interact with NMDA receptors in a positive manner.     

Multiple P2Y receptors mediate contraction in guinea pig mesenteric vein

Vasoconstrictor responses to exogenous adenine and pyrimidine nucleotides were measured in endothelium-denuded segments of guinea pig mesenteric vein and compared with responses in mesenteric artery. The rank order of potency for nucleotides in veins was: 2-MeSADP = 2-MeSATP > UTP > ATPgammaS = alpha,betaMeATP > UDP = ATP > ADP > beta,gamma-D-MeATP = beta,gamma-L-MeATP. In contrast 2-MeSADP, UTP, and UDP were inactive in arteries, and the rank order of potency of other nucleotides differed; that is, alpha,betaMeATP > beta, gamma-D-MeATP > beta,gamma-L-MeATP = ATPgammaS = 2-MeSATP > ATP > ADP. In veins, UTP, ATP, and 2-MeSATP were more efficacious contractile agents than alpha,beta MeATP. In addition, the ability to desensitize responses to these nucleotides and inhibit them with various blockers differed. The response to alpha,betaMeATP in veins exhibited rapid desensitization and was inhibited by pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid tetrasodium (PPADS) and suramin. The response to 2-MeSATP in veins did not desensitize; nor was it inhibited by prior alpha,betaMeATP desensitization, but it was inhibited by PPADS, suramin, and the selective P2Y(1) receptor antagonist adenosine 3',5'-bisphosphate (ABP, 10-100 microM). Responses to ATP and UTP in veins did not desensitize and were not inhibited by PPADS, suramin, ABP, or alpha, betaMeATP desensitization. In conclusion, our results suggest that venous contraction to a variety of nucleotides is mediated in large part by P2Y receptors including P2Y(1) receptors and an UTP-preferring P2Y receptor. A small component of contraction also appears to be mediated by P2X(1) receptors. This receptor profile differs markedly from that of mesenteric arteries in which P2X(1) receptors predominate.     

Cocaine-seeking by rats: regulation, reinforcement and activation

RATIONALE: Previous experiments have shown that P2 receptor activation increases the release of dopamine in the mesolimbic mesocortical system. OBJECTIVE: In order to investigate the functional correlates of dopaminergic stimulation, EEG and behavioural responses to injection of the P2 receptor agonist 2-methylthio ATP (2-MeSATP) into the nucleus accumbens (NAc) of rats were investigated. METHODS: EEG electrodes were positioned into the NAc together with the guide cannula for intracerebral injection. Behavioural analysis was performed in an open field cage and was evaluated by a video activity measurement system. Rats were assigned to separate groups that were given artificial cerebrospinal fluid (aCSF) or drug treatment. RESULTS: 2-MeSATP significantly extended the period of locomotor activity in the novel environment. The quantitative EEG was characterized by an elevation of the power in the alpha-range and a decrease in power in the delta range. The P2 receptor antagonists reactive blue 2 but not pyridoxalphosphate-6-azophenyl-2'4'-disulphonic acid (PPADS) also enhanced locomotion when given alone, and elevated the alpha-1 and beta-2 bands. Both antagonists abolished the locomotor and EEG responses to 2-MeSATP. The dopamine D1 receptor antagonist SCH 23390 and the D2/D3 receptor antagonist sulpiride did not alter locomotor activity when given either alone or in combination. Only sulpiride and especially sulpiride in combination with SCH 23390 prevented the effect of 2-MeSATP. Sulpiride produced a selective increase in the alpha-1 band of the power spectrum whereas SCH 23390 elevated the power of the alpha-1, alpha-2 and beta-1 activities. Neither antagonist inhibited the effect of 2-MeSATP on the EEG when applied separately; however, the co-administration of SCH 23390 and sulpiride abolished the 2-MeSATP-induced alteration of power distribution. After a 6-hydroxydopamine (6-OHDA)-induced lesion of the accumbal dopaminergic terminals, 2-MeSATP failed to enhance the locomotor activity and to induce the characteristic EEG changes. CONCLUSIONS: The observed alterations in open field behaviour and quantitative EEG after injection of 2-MeSATP into the NAc may be mostly due to P2 receptor-mediated dopamine release and subsequent receptor activation.     

Dissociable effects of antagonism of NMDA and AMPA/KA receptors in the nucleus accumbens core and shell on cocaine-seeking behavior.

The purpose of the present experiment was to investigate the involvement of NMDA and AMPA/KA receptors in the nucleus accumbens core and shell in the control over cocaine-seeking behavior by drug-associated cues. Rats were trained under a second-order schedule of reinforcement for cocaine with five infusions of cocaine being available in each daily session. The NMDA receptor antagonist AP-5 and the AMPA/KA receptor antagonist LY293558 were infused directly into the core or shell. LY293558 infused into the core produced a dose-dependent decrease in responding during both the first, cocaine-unaffected interval and also after cocaine had been self-administered in subsequent intervals. By contrast, AP-5 infused into the core had no effect on responding. Infusion of AP-5 into the shell had the limited effect of decreasing responding during the second interval only. There were no effects of LY293558 infused into the shell. These results indicate that NMDA and AMPA receptor-mediated glutamate transmission in the core and shell are dissociably involved in cocaine-seeking behavior controlled in part by drug-associated cues.     

Electrophysiological study of the mechanism of mexidol action

It has been found that mexidol (5 mM) significantly (96 +/- 2%) depressed excitatory postsynaptic current caused by step depolarization in neurons of medial vestibular nucleus of medulla oblongata slices in young (aged 13 - 17 days) male albino rats. In addition, mexidol (2,5 - 5 mM) depressed by 94 +/- 3% excitatory postsynaptic current caused by Shaffer collaterals stimulation of CA1 pyramidal neurons of hippocampal slices in young rats. Complex MK-801 (non-competitive antagonist of NMDA receptors), in contrast to CNQX (competitive AMPA receptor antagonist), considerably decreased the depressant effect of the drug in both brain structures. Therefore, the central favorable effect of mexidol can be mediated by ion mechanisms with glutamate- and GABA-ergic components, primarily by the inhibition of ion currents through NMDA receptor complex.     

γ-Hydroxybutyrate inhibits excitatory postsynaptic potentials in rat hippocampal slices

γ-Hydroxybutyrate (GHB) has been shown to mimic different central actions of ethanol, to suppress alcohol withdrawal syndrome, and to reduce alcohol consumption both in rats and in humans. The aim of the present study was to determine if GHB shared with alcohol the ability to inhibit glutamate action at both NMDA and AMPA/kainate receptors. The NMDA or the AMPA/kainate receptors-mediated postsynaptic potentials were evoked in CA1 pyramidal neurons by stimulation of Schaffer-collateral commissural fibers in the presence of CGP 35348, bicuculline to block the GABA_B and GABA_A receptors, and 10 μM 6,7-dinitroquinoxaline-2,3-dione (DNQX) or 30 μM -2-amino-5-phosphonovalerate (d-APV) to block AMPA/kainate or NMDA receptors, respectively. GHB (600 μM) produced a depression of both NMDA and AMPA/kainate receptors-mediated excitatory postsynaptic potentials with recovery on washout. The GHB receptors antagonist, NCS-382, at the concentration of 500 μM had no effect per se on these responses but prevented the depressant effect of GHB (600 μM) on the NMDA and AMPA/kainate-mediated responses. In the paired-pulse experiments, GHB (600 μM) depressed the amplitude of the first and the second evoked AMPA/kainate excitatory postsynaptic potentials, and significantly increased the paired-pulse facilitation (PPF). These results suggest that GHB inhibits excitatory synaptic transmission at Schaffer-collateral commissural–pyramidal neurons synapses by decreasing the probability of release of glutamate.     

Potentiation of excitatory serotonergic responses by MK-801 in the medial prefrontal cortex

New atypical antipsychotics show a greater affinity to serotonergic rather than to dopamine receptors, suggesting that serotonin (5-HT) has a major role in the pathophysiology and treatment of schizophrenia. The goal of this study was to characterise the response of pyramidal neurons in the medial prefrontal cortex (mPFC) to 5-HT and NMDA before and after administration of the NMDA receptor antagonist, MK-801 (dizocilpine), a well-validated pharmacological model of psychosis. mPFC pyramidal (glutamatergic) neurons were recorded in urethane-anaesthetised rats. The responses to NMDA and 5-HT were assessed using in vivo electrophysiology and microiontophoresis. The 5-HT_2A/2C antagonist ritanserin and the 5-HT_1A antagonist WAY100635 were used to block 5-HT responses. MK-801 decreased the NMDA-induced excitatory responses and increased NMDA-evoked burst activity among mPFC pyramidal neurons. Three subpopulations of pyramidal cells were identified according to their responses to 5-HT: excitation (33%), inhibition (40%) and non-response (27%). The inhibitory responses were blocked by WAY100635 in 100% of cases, but not by ritanserin; the excitatory responses were blocked by ritanserin in 75% of cases, but not by WAY100635. The administration of MK-801 potentiated the firing rate of excitatory responses but did not modify the inhibitory responses induced by microiontophoretic application of 5-HT. These results suggest that MK-801 modifies 5-HT synapses in the mPFC by potentiating the excitatory 5-HT_2A/2C responses and attenuating NMDA excitations. These data indicate that 5-HT excitatory transmission is selectively impaired at the mPFC level in this pharmacological model of schizophrenia.     

Interaction between intrathecal morphine and glutamate receptor antagonists in formalin test

The analgesic interaction between intrathecally administered morphine and the NMDA receptor antagonist, ((+/-)-2-amino-5-phosphonopentanoic acid; AP-5), the NMDA receptor glycine site antagonist, (5-nitro-6,7-dichloro-2,3-quinoxaline dion; ACEA 1021), or the AMPA (alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid) receptor antagonist (ACEA 2752) in the formalin test was investigated with a rat model of chronic lumbar intrathecal catheterization. After obtaining dose-response curves for each agent, combinations of morphine and AP-5, ACEA 1021 or ACEA 2752 were tested for their effect on the number of flinches in the formalin test and for associated side-effects, such as motor disturbances, flaccidity, and agitation/allodynia. Using isobolographic analyses, a potent analgesic synergy was observed with decreased side-effects between morphine and ACEA 2752 or AP-5. ACEA 1021 increased the analgesic effect of low-dose morphine. Spinal mu-opioid receptor activation and NMDA or AMPA receptor antagonism showed a synergistic antinociception against tonic pain. These results suggest an important direction in the management of inflammatory pain.     

The effects of purine compounds on the isolated aorta of the frog Rana temporaria.

1. In the isolated aorta of the frog, Rana temporaria, adenosine concentration-dependently, endothelium-independently relaxed adrenaline pre-constricted vessels. None of the adenosine analogues including D-5''-(N-ethylcarboxamide) adenosine (NECA), R- and S-N6-(2-phenylisopropyl) adenosine (R-and S-PIA) and 2-chloroadenosine (2-CA), or the more selective A1, A2 and A3 agonists cyclopentyladenosine (CPA), CGS 21680 and N6-(3-iodobenzyl) adenosine-5''-N-methylcarboxamide (IB-MECA) respectively, had any effect. 2. The non-selective adenosine antagonist, 8-p-sulphophenyl-theophylline (8-pSPT; 30 microM) failed to inhibit adenosine relaxations, as did NG-nitro-L-arginine methyl ester (L-NAME; 0.1 mM) and indomethacin (30 microM). 3. Adenosine 5''-triphosphate (ATP), alpha, beta-methylene ATP (alpha, beta-MeATP), beta, gamma-methylene ATP (beta, gamma-MeATP), 2-methylthio ATP (2-MeSATP) and uridine 5''-triphosphate (UTP) all concentration-dependently contracted the frog aorta. ATP and alpha, beta-MeATP were equipotent and more potent than UTP and beta, gamma-MeATP; 2-MeSATP had little activity. 4. The P2-purinoceptor antagonist, suramin (0.1 mM) inhibited contractions to alpha, beta-MeATP but not to ATP. Pyridoxalphosphate-6-azophenyl-2'',4''-disulphonic acid (PPADS; 30 microM) also inhibited contractions to alpha, beta-MeATP but not to ATP. Contractions to ATP were, however, inhibited by indomethacin (30 microM). 5. In conclusion, in the frog aorta there appears to be a novel subclass of P1-purinoceptor mediating vasodilatation, although like the A3 subclass it is not blocked by methylxanthines; a P2-purinoceptor mediates vasconstriction which resembles a P2x subtype, based on the agonist potency of alpha, beta-MeATP being more potent than 2-MeSATP (UTP has moderate activity) and PPADS is an effective antagonist. There is no evidence for the presence of a P2y-purinoceptor, mediating vasodilatation, in this preparation.     

Excitatory amino acid receptor-mediated responses in periaqueductal gray neurons are increased during ethanol withdrawal

The ventrolateral periaqueductal gray (PAG) is critical for propagation in the neuronal network for ethanol withdrawal (ETX) seizures, and ethanol is known to alter glutamate effects. This study evaluated changes in glutamate antagonist effects on PAG neurophysiology in brain slices from rats treated with ethanol in vivo. Spontaneous action potentials were rare in control PAG neurons but common during ETX. Spontaneous excitatory postsynaptic potential (EPSP) frequency was increased during ETX, and an AMPA antagonist, 6,7-dinitroquinoxaline-2,3-dione (DNQX) was more effective in suppressing this activity than an NMDA antagonist, 2-amino-7-phosphonoheptanoate (AP7). EPSPs evoked by stimulation of dorsolateral PAG were decreased by AP7 or DNQX in ETX and control neurons. EPSPs of ETX neurons were significantly less sensitive than controls to blockade by AP7 and DNQX. Paired-pulse facilitation of EPSPs was significantly increased during ETX, but paired-pulse inhibition occurred in controls. Thus, PAG hyperexcitability during ETX results from alterations of both NMDA and AMPA receptor-mediated neurotransmission, which may contribute importantly to ETX seizures. These results differ from previous findings in the seizure-initiating site for ETX seizures, inferior colliculus (IC), where NMDA receptor-mediated mechanisms dominate excitability increases during ETX. This dichotomy may be related to the different role played by IC and PAG in the ETX seizure network.     

Effects of 17beta-estradiol on chemically induced long-term depression

In this study, we have investigated the effects of 17beta-estradiol (E2) on chemically induced long-term depression (LTD). LTD was induced by a brief application of N-methyl-D-aspartate (NMDA) or (R,S)-3,5-dihydroxyphenylglycine (DHPG), a group I metabotropic glutamate receptor agonist. Bath application of E2 alone potentiated population excitatory postsynaptic potentials. This potentiation was readily reversed by washing with control saline. The effect of E2 on NMDA-induced LTD was a conversion of LTD to long-term potentiation (LTP). An application of NMDA in the presence of E2 induced LTP. The induction of LTP was inhibited by an inhibitor of calcium/calmodulin dependent protein kinase (CaMKII). The results suggest that E2 potentiates NMDA receptor function and induces an increase in postsynaptic Ca2+ concentration. An increase in postsynaptic Ca2+ concentration activates CaMKII, leading to LTP. In contrast to NMDA-induced LTD, an application of DHPG in the presence of E2 induced significantly larger LTD. The results suggest that E2 potentiates an as yet unidentified process(es) in inducing LTD by an application of DHPG. The effects of E2 both on NMDA-induced and DHPG-induced LTD were suppressed by an estrogen receptor antagonist.     

Functional interactions between presynaptic NMDA receptors and metabotropic glutamate receptors co-expressed on rat and human noradrenergic terminals

BACKGROUND AND PURPOSE: Electrophysiological studies described potentiation of NMDA receptor function by metabotropic glutamate receptors (mGluRs) of group I occurring postsynaptically. Since release-enhancing NMDA receptors exist on noradrenergic terminals and group I mGluRs have recently been identified on these nerve endings, we have investigated if NMDA receptor-mGluR interactions also can occur at the presynaptic level. EXPERIMENTAL APPROACH: Rat hippocampus and human neocortex synaptosomes were labelled with [(3)H]noradrenaline and superfused with mGluR agonists and antagonists. NMDA-evoked [(3)H]noradrenaline release was produced by removal of external Mg(2+) or by simultaneous application of NMDA and AMPA in Mg(2+)-containing solutions. KEY RESULTS: The mGluR1/5 agonist 3,5-DHPG, inactive on its own, potentiated both the release of [(3)H]noradrenaline elicited by AMPA/NMDA/glycine and that evoked by NMDA/glycine following Mg(2+) removal. The effect of 3,5-DHPG on the AMPA/NMDA/glycine-induced release was insensitive to the mGluR1 antagonist CPCCOEt, but it was abolished by the mGluR5 antagonist MPEP; moreover, it was potentiated by the mGluR5 positive allosteric modulator DFB. When NMDA receptors were activated by Mg(2+) removal, both mGluR5 and mGluR1 contributed to the evoked release, the mGluR-mediated release being blocked only by CPCCOEt and MPEP in combination. Experiments with human neocortex synaptosomes show NMDA receptor-mGluR interactions qualitatively similar to those observed in rodents. CONCLUSIONS AND IMPLICATIONS: Group I mGluRs, both of the mGluR1 and mGluR5 subtypes, co-localize with NMDA receptors on noradrenergic terminals of rat hippocampus and human neocortex. Depending on the mode of activation, NMDA receptors exert differential permissive roles on the activation of presynaptic mGluR1 and mGluR5.     

Topiramate antagonizes NMDA- and AMPA-induced seizure-like activity in planarians

The mechanism of anticonvulsant action of topiramate includes inhibition of glutamate-activated ion channels. The evidence is most convincing for direct inhibitory action at the ionotropic AMPA (α-Amino-3-hydroxy-5-methylisoxazole-4-propionic acid) and kainate ((2S,3S,4S)-3-(Carboxymethyl)-4-prop-1-en-2-ylpyrrolidine-2-carboxylic acid) glutamate receptor subtypes. Less direct connection has been made to the NMDA (N-Methyl-d-aspartate) subtype. In the present study, we demonstrate that NMDA and AMPA produce concentration-dependent seizure-like activity in planarians, a type of flatworm which possesses mammalian-like neurotransmitters. In contrast, planarians exposed to the inhibitory amino acid, glycine, did not display pSLA. For combination experiments, topiramate significantly reduced planarian seizure-like activity (pSLA) produced by NMDA or AMPA. Additionally, NMDA-induced pSLA was antagonized by either an NMDA receptor antagonist (MK-801) or AMPA receptor antagonist (DNQX), thus suggesting that NMDA-induced pSLA was mediated by NMDA and non-NMDA receptors. The present results provide pharmacologic evidence of a functional inhibitory action of topiramate on glutamate receptor activity in invertebrates and provide a sensitive, quantifiable end-point for studying anti-seizure pharmacology.     

Regulation of AMPA receptor dephosphorylation by glutamate receptor agonists

Phosphorylation of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor subunit GluR1 at Ser(845) enhances AMPA channel activity. This study demonstrates that Ser(845) is rapidly dephosphorylated upon AMPA receptor activation in nucleus accumbens slices. AMPA-induced dephosphorylation at Ser(845) was blocked by CNQX, an AMPA receptor antagonist, by nifedipine, an L-type Ca(2+) channel antagonist, or by cyclosporin A, a calcineurin inhibitor. N-methyl-D-aspartate (NMDA) treatment also decreased phosphorylation of Ser(845), an effect that was blocked by MK-801, an NMDA receptor antagonist, but not by nifedipine. Accumbens neurons are enriched for dopamine- and cyclic AMP (cAMP)-regulated phosphoprotein, Mr 32,000 (DARPP-32), a potent inhibitor of protein phosphatase 1 (PP1) when phosphorylated by PKA (at Thr(34)). We tested the hypothesis that the AMPA/KA or NMDA-stimulated dephosphorylation of DARPP-32 via calcineurin, leading to increased PP1 activity and dephosphorylation of GluR1. AMPA or NMDA treatment decreased phospho-Thr(34)-DARPP-32 levels, effects that were blocked by receptor antagonists, or cyclosporin A. However, dephosphorylation of Ser(845) mediated by AMPA or NMDA receptors was unaffected in DARPP-32/inhibitor-1 knockout mice. These data suggest that AMPA- or NMDA-induced dephosphorylation of GluR1 at Ser(845) occurs by a mechanism that is independent of DARPP-32 and PP1, but involves activation of calcineurin. Thus, Ca(2+)-dependent dephosphorylation of GluR1 may serve as a negative feedback mechanism for the regulation of AMPA receptor activity in neurons.     

Topiramate antagonizes NMDA- and AMPA-induced seizure-like activity in planarians

The mechanism of anticonvulsant action of topiramate includes inhibition of glutamate-activated ion channels. The evidence is most convincing for direct inhibitory action at the ionotropic AMPA (α-Amino-3-hydroxy-5-methylisoxazole-4-propionic acid) and kainate ((2S,3S,4S)-3-(Carboxymethyl)-4-prop-1-en-2-ylpyrrolidine-2-carboxylic acid) glutamate receptor subtypes. Less direct connection has been made to the NMDA (N-Methyl-d-aspartate) subtype. In the present study, we demonstrate that NMDA and AMPA produce concentration-dependent seizure-like activity in planarians, a type of flatworm which possesses mammalian-like neurotransmitters. In contrast, planarians exposed to the inhibitory amino acid, glycine, did not display pSLA. For combination experiments, topiramate significantly reduced planarian seizure-like activity (pSLA) produced by NMDA or AMPA. Additionally, NMDA-induced pSLA was antagonized by either an NMDA receptor antagonist (MK-801) or AMPA receptor antagonist (DNQX), thus suggesting that NMDA-induced pSLA was mediated by NMDA and non-NMDA receptors. The present results provide pharmacologic evidence of a functional inhibitory action of topiramate on glutamate receptor activity in invertebrates and provide a sensitive, quantifiable end-point for studying anti-seizure pharmacology.     

N-methyl-D-aspartate-induced neuronal activation is selectively modulated by sigma receptors

The effects of two high-affinity sigma ligands, DTG (1,3-di(2-tolyl)guanidine) and haloperidol, on the activation of dorsal hippocampus pyramidal neurons induced by microiontophoretic application of N-methyl-D-aspartate (NMDA) were assessed electrophysiologically. Low doses of DTG (0.5-3 micrograms/kg i.v.) potentiated the NMDA response. This effect of DTG was blocked by haloperidol (10 micrograms/kg i.v.), but not by spiperone, a potent dopamine antagonist with low affinity for sigma receptors. These results suggest that sigma receptors modulate the NMDA-induced neuronal activation.     

Activation of the spinal sigma-1 receptor enhances NMDA-induced pain via PKC- and PKA-dependent phosphorylation of the NR1 subunit in mice

BACKGROUND AND PURPOSE: Previously we demonstrated that the spinal sigma-1 receptor (Sig-1 R) plays an important role in pain transmission, although the exact mechanism is still unclear. It has been suggested that Sig-1 R agonists increase glutamate-induced calcium influx through N-methyl-D-aspartate (NMDA) receptors. Despite data suggesting a link between Sig-1 Rs and NMDA receptors, there are no studies addressing whether Sig-1 R activation directly affects NMDA receptor sensitivity. EXPERIMENTAL APPROACH: We studied the effect of intrathecal (i.t.) administration of Sig-1 R agonists on protein kinase C (PKC) and protein kinase A (PKA) dependent phosphorylation of the NMDA receptor subunit NR1 (pNR1) as a marker of NMDA receptor sensitization. In addition, we examined whether this Sig-1 R mediated phosphorylation of NR1 plays an important role in sensory function using a model of NMDA-induced pain. KEY RESULTS: Both Western blot assays and image analysis of pNR1 immunohistochemical staining in the spinal cord indicated that i.t. injection of the Sig-1 R agonists, PRE-084 or carbetapentane dose dependently enhanced pNR1 expression in the murine dorsal horn. This increased pNR1 expression was significantly reduced by pretreatment with the specific Sig-1 R antagonist, BD-1047. In another set of experiments Sig-1 R agonists further potentiated NMDA-induced pain behaviour and pNR1 immunoreactivity and this was also reversed with BD-1047. CONCLUSIONS AND IMPLICATIONS: The results of this study suggest that the activation of spinal Sig-1 R enhances NMDA-induced pain via PKC- and PKA-dependent phosphorylation of the NMDA receptor NR 1 subunit.     

Ethanol inhibition of synaptically evoked kainate responses in rat hippocampal CA3 pyramidal neurons.

Many studies have demonstrated that intoxicating concentrations of ethanol (10-100 mM) can selectively inhibit the component of glutamatergic synaptic transmission mediated by N-methyl-D-aspartate (NMDA) receptors while having little or no effect on excitatory synaptic transmission mediated by non-NMDA receptors [i.e., alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and/or kainate (KA) receptors]. However, until the recent development of highly selective AMPA receptor antagonists, it was not possible to assess the relative contribution of AMPA and KA receptors to non-NMDA receptor-mediated synaptic transmission or to determine whether these glutamate receptor subtypes differed in their sensitivity to ethanol. In the present experiments, we used the highly selective AMPA receptor antagonist LY 303070 to pharmacologically isolate KA receptor-mediated excitatory postsynaptic currents (EPSCs) in rat hippocampal CA3 pyramidal neurons and tested their sensitivity to ethanol. Concentrations of ethanol as low as 20 mM significantly and reversibly depressed KA EPSCs. Ethanol also inhibited KA currents evoked by direct pressure application of KA in the presence of LY 303070, suggesting that this inhibition was mediated by a postsynaptic action. In contrast, ethanol had no effect on AMPA EPSCs in these cells, even at the highest concentration tested (80 mM). Ethanol significantly inhibited NMDA EPSCs in these neurons, but these responses were less sensitive to ethanol than KA EPSCs. These results suggest that in addition to its well-described depressant effect on NMDA receptor-mediated synaptic transmission, ethanol has an even greater inhibitory effect on glutamatergic synaptic transmission mediated by KA receptors in rat hippocampal CA3 pyramidal neurons.     

High potency of the orally-active NMDA-receptor antagonist CGP 40 116 in inhibiting excitatory postsynaptic potentials of rat basolateral amygdala neurones in vitro

Conventional intracellular recordings were used to monitor postsynaptic potentials of basolateral amygdala neurones (BLA) in brain slices comprising the BLA, the entorhinal cortex (EC) and the hippocampus, in which the EC-BLA connections were preserved. Stimulation of the BLA with a bipolar electrode elicited complex postsynaptic potentials consisting of α-amino-3-hydroxy-5-methyl-isoxazoleproprionic acid (AMPA) receptor-mediated fast excitatory postsynaptic potentials (fast EPSPs), γ-amino-butyric acid (GABA_A) receptor-mediated fast inhibitory postsynaptic potentials (fast IPSPs) and GABA_B receptormediated slow IPSPs. Bath application of 10 μM of the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2, 3-dione and of 10 μM of the GABA_A receptor antagonist bicuculline methiodide (BMI) revealed a N-methyl-D-aspartate (NMDA) receptor-mediated slow EPSPs, which was occasionally followed by a GABA_B receptormediated slow IPSPs. Under these conditions, the log concentration-response curve for -(E)-2-amino-4-methyl-5-phosphono-3-pentanoic acid (CGP 40 116), a newly developed drug with proposed NMDA-receptor antagonist properties, was compared to that obtained with the ‘classic’ antagonist D(−)-2-amino-5-phosphonopentanoic acid ( -AP5) in inhibiting the NMDA-mediated postsynaptic potentials. CGP 40 116 ( ₅₀: 130 nM) was over 30 times more potent than -AP5 ( ₅₀: 4100 nM) in reducing NMDA-mediated slow EPSP. In conclusion, the present study indicates that CGP 40 116, a new orally-active NMDA antagonist, shows a very high potency on NMDA receptors in the amygdala and may therefore be a valuable tool for studying the behavioural effect of NMDA-receptor mediated transmission in this structure.