Regulation of cholecystokinin mRNA content in rat striatum: a glutamatergic hypothesis.

Changes in the cholecystokinin (CCK) mRNA content in rat striatum after the administration of specific glutamate and dopamine (DA) receptor agonists and antagonists were investigated. MK-801 (1 mg/kg i.p.), a selective noncompetitive N-methyl-D-aspartate (NMDA)-sensitive glutamatergic receptor antagonist, but not 6-cyano-7-nitroquinoxaline-2,3-dione (1.1-9.2 micrograms i.c.v.), a competitive non-NMDA glutamatergic receptor antagonist, produced a time- and dose-dependent decrease in striatal CCK mRNA. The maximum inhibition (50%) was observed after a daily treatment for 1 week with MK-801 (1 mg/kg). The activation of NMDA receptors by a single injection of NMDA (1.4 micrograms i.c.v.) elicited an 80% increase in CCK mRNA in rat striatum 8 hr after the injection. These data suggest that glutamate exerts a tonic regulation on striatal CCK mRNA, mainly through NMDA-sensitive glutamatergic receptors. B-HT 920, a DA D2 receptor agonist and benztropine, a DA uptake blocker, increased striatal CCK mRNA. This increase was partially blocked by the concomitant administration of MK-801. Moreover, the DA receptor antagonist haloperidol, at a dose that per se failed to change CCK mRNA (0.3 mg/kg i.p.), partially blocked the increase in CCK mRNA elicited by NMDA. Similarly, the NMDA effect was attenuated in rats with a 6-hydroxydopamine-induced nigrostriatal lesion. Our findings suggest that in rat striatum a complex DA-glutamate interaction tonically regulates CCK expression via D2 and/or NMDA receptor activation.     

Met-enkephalin release from slices of the rat striatum and globus pallidus: stimulation by excitatory amino acids.

The present study evaluated the effects of high K+ and four excitatory amino acids (EAAs) on the release of met-enkephalin-like immunoreactivity (ME-i.r.) from slices of the rat striatum and globus pallidus. High K+ (15-50 mM) increased the release of ME-i.r. in a concentration-dependent manner in both regions, the release response in the globus pallidus being consistently greater than in the striatum. This release was highly Ca(++)-dependent and was significantly enhanced in the absence of external Mg++. D-2-Amino-7-phosphonoheptanoic acid (0.5 mM), a competitive N-methyl-D-aspartate (NMDA) receptor antagonist, did not alter this enhanced action of K+, suggesting that the activation of NMDA receptors by an endogenous agonist did not contribute to the enhancement. Exposure of pallidal or striatal slices to four EAA receptor agonists, NMDA, L-glutamate, kainate (KA) and quisqualate, increased the release of ME-i.r. above the base line, an effect that was Ca(++)-dependent. Both L-glutamate and NMDA, at concentrations of 1 and 5 mM, produced a graded increase in the ME-i.r. release, but a higher concentration (10 mM) produced a lower release. In both regions the NMDA (5 mM)-evoked release was effectively inhibited by Mg++ (1.2 mM), 3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) (5 microM), a competitive NMDA receptor antagonist and thienylcyclohexylpiperidine (10 microM), a noncompetitive NMDA receptor antagonist. Tetrodotoxin (0.3 microM), a Na+ channel blocker, did not affect the NMDA-evoked release of ME-i.r. in the striatum, but decreased it by 52% in the globus pallidus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regulation of striatal enkephalin turnover in rats receiving antagonists of specific dopamine receptor subtypes

The hypothesis that striatal dopamine regulates enkephalin (ENK) synthesis is supported by the increase of striatal proenkephalin mRNA and ENK after intranigral injection of 6-hydroxydopamine. In order to elucidate which dopamine receptor subtype is operative in the regulation of the dynamic state of ENK, the effect of drugs that block D-1 or D-2 receptor selectively was studied. Daily administration of 140 mumol/kg s.c. of the D-2 antagonist I-sulpiride twice daily for 2 weeks produces a 30% decrease in the content of striatal proenkephalin mRNA and ENK. In contrast, a 50% increase was observed after 2 weeks of treatment with the D-1 antagonist SCH 23390 at 74 nmol/kg s.c. three times a day. Hence, it can be inferred that the endogenous activation of D-1 tonically decreases striatal ENK synthesis. Removal of this neurally mediated regulation either by a specific pharmacologic blockage of D-1 or by lesioning with 6-hydroxydopamine increases the biosynthesis of ENK. The increase of ENK biosynthesis elicited by denervation with 6-hydroxydopamine cannot be due to the endogenous activation of D-2 receptors and must be due to the inactivation of the tonic inhibition exerted by D-1 receptors.     

Role of adenosine and N-methyl-D-aspartate receptors in mediating haloperidol-induced gene expression and catalepsy.

Acute blockade of dopamine D(2) receptors by the typical antipsychotic drug haloperidol leads to alterations in neuronal gene expression and behavior. In the dorsolateral striatum, the levels of mRNA for the immediate-early gene c-fos and the neuropeptide gene neurotensin/neuromedin N (NT/N) are significantly increased by haloperidol. An acute behavioral response to haloperidol is catalepsy, considered to be a rodent correlate of some of the immediate extrapyramidal motor side effects seen in humans. Several lines of evidence suggest a link between neurotensin induction in the dorsolateral striatum and catalepsy. We hypothesize that both striatal gene induction and catalepsy elicited by haloperidol arise from the combined effect of excitatory adenosinergic and glutamatergic inputs acting at adenosine A(2A) and N-methyl-D-aspartate (NMDA) receptors, respectively. In agreement with our previous reports, adenosine antagonists reduced haloperidol-induced c-fos and neurotensin gene expression as well as catalepsy. In agreement with other reports, the noncompetitive NMDA receptor antagonist MK-801 also reduced gene expression and catalepsy in response to haloperidol. The competitive NMDA receptor antagonist LY235959 decreased haloperidol-induced catalepsy. We show here that blocking both A(2A) and NMDA receptors simultaneously in conjunction with haloperidol resulted in a combined effect on gene expression and behavior that was greater than that for block of either receptor alone. Both c-fos and NT/N mRNA levels were reduced, and catalepsy was completely abolished. These results indicate that the haloperidol-induced increases in c-fos and NT gene expression in the dorsolateral striatum and catalepsy are driven largely by adenosine and glutamatergic inputs acting at A(2A) and NMDA receptors.     

Amino acid metabolism in neurodegenerative diseases]

Although various neurological diseases occur in patients with inborn error of metabolism of amino acids, amino acids also act as neurotransmitters. Glutamic acid, aspartic acid and glycine play roles as an excitatory neurotransmitter, but exert a neurodegenerative effect in case of the excessive release. Extensive studies have recently been performed on glutamate receptors, especially N-methyl-D-aspartate (NMDA) receptor in the hippocampus. Alzheimer brain shows a decreased number of NMDA receptors in the frontal cortex. The parkinsonian changes caused by MPTP is abolished by the administration of a NMDA antagonist. gamma-Aminobutyric acid (GABA) acts as an inhibitory amino acid. The content of GABA is low in the striatum of patients with Huntington's disease. The number of NMDA receptor is decreased also in Huntington striatum. These observations may give a clue for the prevention of various neurodegenerative diseases.     

Tachykinin gene expression in rat limbic nuclei: modulation by dopamine antagonists

The content and nature of the preprotachykinin (PPT; i.e., substance P/neurokinin A-encoding) messenger RNAs (mRNAs) present in rat brain striatum and limbic tissues were determined by RNA protection experiments. The rank order of PPT mRNA concentration was striatum greater than nucleus accumbens much greater than bed nucleus of the stria terminalis greater than hypothalamus, amygdala and septum. The proportion of beta-(full length) to gamma-(minus exon 4) PPT mRNA was invariant (40/60) among the tissues tested. Because these brain regions receive prominent dopaminergic innervations, the effects of repeated treatment with dopamine antagonists (antipsychotic drugs) on PPT gene expression were assessed. The prototypical dopamine antagonists haloperidol and chlorpromazine decreased striatal PPT mRNA, had no effect on PPT mRNA in the nucleus accumbens or bed nucleus of the stria terminalis, and increased septal PPT mRNA levels. In contrast, the atypical antipsychotic drugs clozapine and l-sulpiride did not alter striatal or septal PPT mRNA, but increased PPT mRNA content in the nucleus accumbens and bed nucleus. The correlation between the effects of typical and atypical antipsychotic drugs on rat striatal and limbic PPT gene expression and their clinical side effects and therapeutic efficacy is discussed.     

Competitive and noncompetitive antagonists at N-methyl-D-aspartate receptors protect against methamphetamine-induced dopaminergic damage in mice.

The administration of methamphetamine (METH) to experimental animals results in damage to nigrostriatal dopaminergic neurons. We have demonstrated previously that the excitatory amino acids may be involved in this neurotoxicity. For example, several compounds which bind to the phenyclidine site within the ion channel linked to the N-methyl-D-aspartate (NMDA) receptor protected mice from the METH-induced loss of neostriatal tyrosine hydroxylase activity and dopamine content. The present study was conducted to characterize further the role of the excitatory amino acids in mediating the neurotoxic effects of METH. The administration of three or four injections of METH (10 mg/kg) every 2 hr to mice produced large decrements in neostriatal dopamine content (80-84%) and in tyrosine hydroxylase activity (65-74%). A dose-dependent protection against these METH-induced decreases was seen with two noncompetitive NMDA antagonists, ifenprodil and SL 82.0715 (25-50 mg/kg/injection), both of which are thought to bind to a polyamine or sigma site associated with the NMDA receptor complex, and with two competitive NMDA antagonists, CGS 19755 (25-50 mg/kg/injection) and NPC 12626 (150-300 mg/kg/injection). Moreover, an intrastriatal infusion of NMDA (0.1 mumol) produced a slight but significant loss of neostriatal dopamine which was potentiated in mice that also received a systemic injection of METH. The results of these studies strengthen the hypothesis that the excitatory amino acids play a critical role in the nigrostriatal dopaminergic damage induced by METH.     

Regulation of the mesocorticolimbic dopamine system by glutamic acid receptor subtypes

Glutamic acid and excitatory amino acids specific for the glutamate receptor subtypes were microinjected into the A10 region of the rat. Glutamate produced an increase in motor behavior that was antagonized by pretreatment with the dopamine D2 receptor antagonist, haloperidol. This motor stimulant effect was produced by kainate, but not by N-methyl-D-aspartate (NMDA) or quisqualic acid. By using in vivo dialysis it was found that dopamine release in the nucleus accumbens and locomotor activity were enhanced by glutamate injection into the A10 region. Whereas glutamate was found to increase the postmortem concentration of dopamine metabolites in the medial prefrontal cortex, nucleus accumbens and A10 region, NMDA selectively increased dopamine metabolism in the prefrontal cortex, and kainate produced increases in the nucleus accumbens and A10 region. When glutamate and the NMDA receptor antagonist, 3-[(+/-)-2-carboxypiperazine-4-yl)propyl-1-phosphonic acid (CPP) were coadministered, CPP selectively abolished the effect of glutamate on medial prefrontal cortical dopamine metabolites. A physiological role for the NMDA receptor modulation of A10 dopamine neurons was shown by intra-A10 pretreatment with CPP antagonism of mild footshock-induced increase in dopamine metabolites in the prefrontal cortex. These data argue that glutamate is a regulatory transmitter of A10 dopamine neurons, and that the NMDA receptor subtype modulates neurons projecting to the prefrontal cortex whereas the kainate subtype modulates mesoaccumbens neurons.     

Characterization of the inhibition of excitatory amino acid-induced neurotransmitter release in the rat striatum by phencyclidine-like drugs

In the present study, the authors found that, in Mg++-free buffer, N-methyl-D-aspartate (NMDA) was able to evoke the Ca++-dependent and tetrodotoxin-sensitive release of striatal acetylcholine (ACh), presumably via interaction with receptors on cholinergic interneurons. In Mg++-free buffer containing pargyline, NMDA also evoked a Ca++-dependent and tetrodotoxin-sensitive release of striatal [3H]dopamine (DA). Phencyclidine (PCP) and physiological concentrations of Mg++ (1.2 mM) also inhibited ACh release evoked by L-glutamate, L-aspartate and DL-homocysteate, but not ACh release evoked by the glutamate analogs quisqualate and kainate, suggesting that PCP is selective for the magnesium-sensitive, NMDA-preferring glutamate-aspartate receptor subtype. Comparison of PCP inhibition of NMDA-stimulated ACh and DA release with that produced by the competitive NMDA antagonist 2-amino-5-phosphonovalerate indicates that PCP is probably not altering release by a direct action on the NMDA recognition site. The ability of 2-amino-5-phosphonovalerate, but not PCP, to prevent desensitization of NMDA-induced ACh release is consistent with this interpretation. Binding studies did, however, reveal a reduction in the apparent affinity of the PCP binding site by high concentrations of NMDA. This may suggest an allosteric link between the PCP-sigma receptor and the NMDA-type glutamate-aspartate receptor. The receptors mediating excitatory amino acid-induced DA release were somewhat less selective than those on cholinergic neurons in their sensitivity to both Mg++ and PCP. Structure-activity-relationship studies suggested that the inhibition off ACh and DA release evoked by NMDA involves biding to the PCP-sigma receptor.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intrastriatal injection of a selective metabotropic excitatory amino acid receptor agonist induces contralateral turning in the rat.

The consequence of in vivo activation of the phosphoinositide-coupled (metabotropic) excitatory amino acid (EAA) receptor subtype was investigated. We report that unilateral intrastriatal injection of 1S,3R-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD), a selective metabotropic EAA receptor agonist, produced turning behavior (rotations) contralateral to the site of injection. This effect peaked at 5 to 8 hr after injection and was dose-related (EC50 = 0.59 mumol), producing a maximal effect at 1 mumol (32 +/- 4 rotations per 5 min). 1S,3R-ACPD-induced rotations were not mimicked by intrastriatal injection of vehicle (2 microliters of normal saline) or up to 2 mumol of 1R,3S-ACPD, the inactive ACPD isomer at the metabotropic EAA receptor. The selective competitive N-methyl-Daspartate receptor antagonist LY27461 4 (up to 5 mg/kg i.p) did not significantly affect 1S,3R-ACPD-induced rotations. However, coinjection of the metabotropic EAA receptor antagonist L-2-amino-3-phosphonopropionic acid (1 mumol) significantly reduced 1S,3R-ACPD-induced contralateral rotations. 1S,3R-ACPD at a dose which produced maximal contralateral rotations did not produce any loss of striatal gamma-aminobutyric acid neurons as indexed by glutamic acid decarboxylase enzyme activity in the injected striatum. In contrast to 1S,3R-ACPD, a dose of N-methyl-D-aspartate (0.2 mumol), which only very modestly induces contralateral rotations results in highly significant neuronal degeneration (50% loss of glutamic acid decarboxylase activity), and is associated with other excitatory behaviors such as clonic convulsions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enhancement of spinothalamic neuron responses to chemical and mechanical stimuli following combined micro-iontophoretic application of N-methyl-D-aspartic acid and substance P.

A role for sensitization of nociceptors in the generation of primary hyperalgesia is well documented. More recent work has begun to define a role of an increased excitability of neurons within the spinal cord in the generation of secondary hyperalgesia. The present study demonstrates increased responses of primate spinothalamic neurons following co-administration of N-methyl-D-aspartic acid (NMDA) and substance P (SP) by micro-iontophoresis. Wide dynamic range and high threshold STT neurons in laminae I-VI showed an increased frequency of discharges following application of NMDA which was characterized by a slow onset to peak discharge rate and a slow return to background levels of discharge. Combined application of NMDA with SP resulted in an enhancement of responses to NMDA that often long outlasted the administration of SP. This increase in response of the cells to NMDA was not produced by repeated application of NMDA alone or following combined application of NMDA with an SP analog. NMDA responses were reduced or prevented in all cases by co-application of an NMDA-receptor antagonist. Finally, long-lasting potentiation of NMDA responses by SP was paralleled by enhanced responses to mechanical stimulation of skin. It is proposed that a mechanism involving the combined synaptic release of excitatory amino acids and peptides leads to secondary hyperalgesia.     

Activation of excitatory amino acid receptors may mediate the folate-induced stimulation of locomotor activity after bilateral injection into the rat nucleus accumbens

Folic acid (FA) and 5-formyltetrahydrofolic acid (FTHF) have been shown previously to produce a marked stimulation of locomotor activity after bilateral injection into the rat nucleus accumbens. This study was designed to determine whether the hypermotility response produced by the folates is mediated through the activation of excitatory amino acid receptors in the nucleus accumbens. Although FA stimulated locomotor activity, pteroic acid, a congener of FA that lacks the glutamate moiety, was ineffective, suggesting that the glutamate portion of the molecule is essential for the hypermotility response. The N-methyl-D-aspartic acid (NMDA) receptor antagonists, D-alpha-aminoadipic acid, DL-alpha-epsilon-diaminopimelic acid and MgCl2, at doses that attenuated NMDA-induced hypermotility, were ineffective in decreasing the folate-induced hypermotility response. This behavioral observation is consistent with the biochemical observation that the folates, at a 1 mM concentration, were unable to stimulate the release of [3H]acetylcholine from striatal slices, a model system that is sensitive to the activation of NMDA receptors. In contrast to the ineffectiveness of the NMDA antagonists in inhibiting the response to the folates, the antagonist, glutamic acid diethylester, which inhibited the response to quisqualic acid, but not NMDA, also inhibited the response to both FA and FTHF. Two recently characterized dipeptides, gamma-D-glutamylaminomethylsulfonic acid and gamma-D-glutamyltaurine, antagonized the stimulation of locomotor activity produced by quisqualic acid, FA and FTHF. However, these dipeptides also inhibited the response to NMDA, suggesting that these compounds are not able to distinguish between quisqualate and NMDA receptors in the nucleus accumbens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pharmacological characterization of LY233053: a structurally novel tetrazole-substituted competitive N-methyl-D-aspartic acid antagonist with a short duration of action

This study reports the activity of a structurally novel excitatory amino acid receptor antagonist, LY233053 [cis-(+-)-4-[(2H-tetrazol-5-yl)methyl]piperidine-2-carboxylic acid], the first tetrazole-containing competitive N-methyl-D-aspartic acid (NMDA) antagonist. LY233053 potently inhibited NMDA receptor binding to rat brain membranes as shown by the in vitro displacement of [3H] CGS19755 (IC50 = 107 +/- 7 nM). No appreciable affinity in [3H]alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) or [3H]kainate binding assays was observed (IC50 values greater than 10,000 nM). In vitro NMDA receptor antagonist activity was further demonstrated by selective inhibition of NMDA-induced depolarization in cortical wedges (IC50 = 4.2 +/- 0.4 microM vs. 40 microM NMDA). LY233053 was effective after in vivo systemic administration in a number of animal models. In neonatal rats, LY233053 selectively blocked NMDA-induced convulsions (ED50 = 14.5 mg/kg i.p.) with a relatively short duration of action (2-4 hr). In pigeons, LY233053 potently antagonized (ED50 = 1.3 mg/kg i.m.) the behavioral suppressant effects of 10 mg/kg of NMDA. However, a dose of 160 mg/kg, i.m., was required to produce phencyclidine-like catalepsy in pigeons. In mice, LY233053 protected against maximal electroshock-induced seizures at lower doses (ED50 = 19.9 mg/kg i.p.) than those that impaired horizontal screen performance (ED50 = 40.9 mg/kg i.p.). Cholinergic and GABAergic neuronal degenerations after striatal infusion of NMDA were prevented by single or multiple i.p. doses of LY233053. In summary, the antagonist activity of LY233053 after systemic administration demonstrates potential therapeutic value in conditions of neuronal cell loss due to NMDA receptor excitotoxicity. The relatively short duration of action of LY233053 may make this compound particularly advantageous as a neuroprotective agent in the treatment of acute conditions such as cerebral ischemia.     

Role of nitric oxide in the genesis of excitatory amino acid-induced seizures from the deep prepiriform cortex

The role of nitric oxide (NO) in the genesis of motor and electrocortical seizures elicited by administration of excitatory amino acid agonists into the deep prepiriform cortex (DPC) has been evaluated. Motor and electrocortical seizures occurred in rats receiving unilateral microinjections into the DPC of either N-methyl-D-aspartate (NMDA, 5 and 10 nmol) or kainate (KA, 100 pmol). The selective NMDA receptor antagonist 2-amino-7-phosphonoheptanoate (APH), when microinjected into DPC, prevented the development of seizures induced by both NMDA and KA injected in the same site. In addition, methylene blue (20 nmol, which prevents activation of soluble guanylate cyclase) or NG-monomethyl-L-arginine (NMMA, 40 nmol; a specific inhibitor of nitric oxide synthesis), when microinjected into DPC 15 min prior to either NMDA or KA, significantly protected against seizures elicited by both excitatory amino acid agonists. These data confirm the role of excitatory amino acid transmission in the genesis of seizures elicited from the deep prepiriform cortex. They further suggest that activation of excitatory amino acid receptors within the DPC leads to the release of a substance which shares properties with EDRF/NO and contributes to the genesis of seizure activity in this area.     

Inhibition of the N-methyl-d-aspartate receptor unmasks the antinociception of endogenous opioids in the periphery

Although N-methyl-d-aspartate (NMDA) receptor antagonists potentiate antinociceptive effects induced by various exogenous opioids at the spinal, supraspinal, or peripheral level, less is known regarding the interaction between NMDA and endogenous opioids in antinociception. We therefore assessed the effects of NMDA receptor antagonists on endogenous opioids in antinociception at the peripheral level by testing the ability of the locally administered receptor antagonists to modify pain-related behavior induced by carrageenan injection into the knee joint. The NMDA receptor antagonist AP-5 or the exogenous opioid morphine was injected intra-articularly before carrageenan injection and 5 h after carrageenan injection, respectively. We evaluated whether intra-articular injection of the opioid receptor antagonist naloxone reversed the analgesic effect of AP-5. In addition, we tested the effects of AP-5 on carrageenan-induced levels of the β-endorphin protein in dorsal root ganglia (DRG), saphenous nerve and synovial membrane. We found that AP-5 prevented and morphine reversed carrageenan-induced pain-related behavior. Intriguingly, injection of naloxone 5 h after carrageenan injection reversed the antinociceptive effects of AP-5 pre-treatment, although naloxone alone had no effect on carrageenan-induced pain-related behavior. Western blots showed that AP-5 pre-treatment followed by carrageenan injection resulted in a higher level of β-endorphin protein in the DRG and saphenous nerve, but not in the synovial membrane, than that observed following saline control treatment. These results suggest that inhibition of the NMDA receptor unmasks antinociception induced by endogenous opioids at the peripheral level, partly through the increased protein level of the endogenous μ-opioid peptide β-endorphin in DRG and saphenous nerve.     

Muscimol, gamma-aminobutyric acidA receptors and excitatory amino acids in the mouse spinal cord

These experiments examined the effects of intrathecally administered gamma-aminobutyric acid (GABA) agonists on the effects of intrathecally administered excitatory amino acid (EAA) agonists: N-methyl-D-aspartic acid (NMDA), quisqualic acid and kainic acid. We have found that muscimol, a GABAA receptor agonist, but not baclofen, a GABAB receptor agonist, dose-dependently inhibited caudally directed biting and scratching behavior induced by all three EAA agonists. This nonselective blockade of the expression of effects mediated by all three types of EAA receptor is in marked contrast to the selective blockade of NMDA effects seen previously in the case of mu opioids and phencyclidine receptor agonists. Inhibition by muscimol was blocked with the GABAA receptor antagonist, bicuculline. Decreased latency or hyperalgesia in the tail-flick test, found previously to be induced selectively by NMDA and blocked by an NMDA receptor antagonist, was similarly affected by muscimol but not baclofen, each given intrathecally. However, muscimol prolonged the tail-flick latency only after presentation of NMDA suggesting a possible antinociceptive effect of GABAA agonists in the presence of agonists at NMDA receptors. This study together with the preceding paper resolves GABA-mediated spinal antinociception into two components: a GABAA agonist selectively blocks nociception involving EAA receptors whereas a GABAB agonist selectively blocks substance P spinal activity (the preceding paper).     

Compensatory activation of substance P biosynthesis by L-dihydroxyphenylalanine in striatonigral neurons of neonatal dopaminergic denervated rats.

Previous studies have shown that L-dihydroxyphenylalanine (L-DOPA) administration to adult rats that had been subjected to dopamingergic denervation with 6-hydroxydopamine during early postnatal period, led to a marked decrease in the levels of striatonigral substance P (SP). The present study examined the hypothesis of whether there is a compensatory activation of SP biosynthesis in order to replenish the L-DOPA-induced SP depletion. Three-day old Sprague-Dawley rat pups were lesioned with 6-hydroxydopamine and were challenged with L-DOPA at about 60 days of age. The animals were sacrificed 75 min, 6 hr or 24 hr after the L-DOPA administration. The levels of SP (in the striatum and substantia nigra) were determined by radioimmunoassay. The abundance of SP-encoding preprotachykinin (PPT) messenger RNA (mRNA) in the striatum was determined by Northern blot analysis and the abundance of individual PPT-mRNAs (alpha, beta and gamma) was determined by nuclease protection assays. Concentrations of dopamine and its acid metabolite, dihydroxyphenyl acetic acid were determined by high-pressure liquid chromatography with electrochemical detection. At the 75 min time point, L-DOPA produced a greater decrease in SP levels in the striatum and the substantia nigra, than that observed with lesion alone. The SP levels recovered to lesioned-control levels by 6 hr and remained at this level at the 24-hr time point. This recovery was accompanied by a marked increase in striatal SP-encoding PPT-mRNA abundance at 6 hr; mRNA levels were below lesioned-control value at 24 hr. There were no differential changes in the individual SP-encoding PPT-mRNAs.(ABSTRACT TRUNCATED AT 250 WORDS)     

N-methyl-D-aspartate receptor blockade inhibits cardiac inflammation in the Mg2+-deficient rat

Elevated plasma levels of the neuropeptide substance P (SP) precede the perivascular inflammatory infiltrate seen in hearts of Mg(2+)-deficient (MgD) animals. The N-methyl-d-aspartate (NMDA) receptor is found in neurons, and activation of this receptor participates in SP release; under normal circumstances, this release can be blocked by Mg(2+). Therefore, we reasoned that blockade of the NMDA receptor with dizolcipine maleate (a noncompetitive NMDA receptor antagonist) would prevent SP release from C-fibers due to MgD. In this study, animals were implanted with slow-release pellets containing dizolcipine or placebo and were fed with diet sufficient in Mg(2+) or deficient with only 9% of USDA-recommended Mg(2+). SP immunostaining of dorsal root ganglia showed a time-dependent depletion of SP in the MgD animals, with a dramatic decrease of SP by week 2; this depletion was prevented by pretreatment with dizolcipine maleate. The significant increase in plasma prostaglandin E(2) levels during MgD was prevented by dizolcipine, and the loss of total red blood cell glutathione content was significantly attenuated by NMDA blockade after 3 weeks of MgD (p < 0.01 versus controls). Immunohistochemical and Western blot analyses of ventricular tissue demonstrated that NMDA receptor blockade abolished MgD-related increase of endothelium adhesion molecule CD54 (weeks 1 and 2; p < 0.05), and of monocyte/macrophage surface protein CD11b expression (week 3; p < 0.05). We conclude that NMDA receptor blockade with dizolcipine maleate prevented SP depletion and reduced perivascular inflammatory infiltrates, thus decreasing cardiac injury due to Mg(2+) deficiency.     

电磁脉冲对大鼠脑海马N-甲基-D-天冬氨酸受体活性的影响

背景电磁脉冲是一种高能非电离辐射,它涵盖的频谱极宽,对电子仪器具有极强的破坏力,并具有一定的生物损伤效应.本课题组前期实验发现,电磁脉冲可造成实验大鼠学习记忆能力的显著下降,并且影响其海马长时程增强效应的形成.目的观察电磁脉冲照射对大鼠海马组织中氨基酸类神经递质含量和N-甲基-D-天冬氨酸受体的影响.设计随机对照的实验,方差分析.单位军事医学科学院放射与辐射医学研究所.材料实验于2004-01/03在军事医学科学院放射与辐射医学研究所完成.实验选用雄性Wistar大鼠32只,随机分为电磁脉冲组26只和对照组6只.方法电磁脉冲组大鼠经电磁脉冲照射(6×104V/m,脉冲上升时间20ns,脉宽30μs,频率2.5脉冲/min,作用2 min)后即刻,3,6,24,48 h麻醉状态下断头取脑,剥离海马;用高效液相色谱法测定氨基酸含量,并以3H标记的谷氨酸为配基进行放射性配基-受体结合实验.对照组麻醉处死前不做任何处理.主要观察指标①各组大鼠海马组织兴奋性氨基酸和抑制性氨基酸含量的变化.②各组大鼠海马N-甲基-D-天冬氨酸受体的最大结合量和平衡解离常数的变化.结果32只大鼠全部进入结果分析.①电磁脉冲照射后即刻、3 h、6 h可见天冬氨酸的含量显著升高[峰值(17.25±1.63)μmol/L],明显高于对照组[(10.56±1.5)μmol/L,P＜0.05],谷氨酸含量也于上述3个时间点升高[峰值(13.67±0.95)μmol/L],显著高于对照组[(6.94±1.1)μmol/L,P＜0.05],两者含量均于照射后24 h渐趋恢复,48 h接近正常水平.3抑制性氨基酸(甘氨酸、牛磺酸、γ-氨基丁酸)的含量也于电磁脉冲照射后不同时间点升高,并于48 h恢复.谷氨酸/γ-氨基丁酸比值于照后即刻明显升高(P＜0.05),照后24 h明显下降,48 h恢复至接近对照组.②电磁脉冲照射后即刻大鼠海马中N-甲基-D-天冬氨酸受体的平衡解离常数开始下降,照后3 h下降最显著(P＜0.05),6 h渐有恢复,48 h恢复至正常水平;受照大鼠海马中N-甲基-D-天冬氨酸受体的最大结合量于照后3 h和6 h显著下降(P＜0.05),24 h渐有恢复,照后48 h明显升高,超过正常组水平(P＜0.05).结论电磁脉冲照射导致大鼠海马组织中兴奋性氨基酸含量及谷氨酸/γ-氨基丁酸比值升高;同时N-甲基-D-天冬氨酸受体的亲和力升高及受体密度下降.提示实验动物认知障碍可能与兴奋性氨基酸的过度释放和N-甲基-D-天冬氨酸受体功能改变有关.     

Activation of brainstem N-methyl-D-aspartate receptors is required for the analgesic actions of morphine given systemically

The analgesic actions of opioids are in large part mediated by activation of brainstem pain modulating neurons that depress nociceptive transmission at the level of the dorsal horn. The present study was designed to characterize the contribution of N-methyl-D-aspartate (NMDA)- and non-NMDA-mediated excitatory transmission within the rostral ventromedial medulla (RVM) to the activation of brainstem inhibitory output neurons and analgesia produced by systemic morphine administration. The NMDA receptor antagonist D-2-amino-5-phosophonopentanoic acid (AP5), the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione disodium (CNQX) or saline was infused into the RVM of lightly anesthetized rats while recording the activity of identified pain modulating neurons: 'off-cells', thought to inhibit nociceptive transmission, and 'on-cells', thought to facilitate nociception. Nociceptive responsiveness (tail flick latency) was not affected by either antagonist. AP5, but not CNQX, attenuated or blocked activation and disinhibition of off-cells and the antinociception produced by systemically administered morphine. Reflex-related discharge of on-cells was unaffected by AP5, but significantly attenuated by CNQX. The present results highlight two important aspects of RVM pain modulatory circuits. First, morphine given systemically produces its analgesic effect at least in part by recruiting an NMDA-mediated excitatory process to activate off-cells within the RVM. This excitatory process may play a role in the analgesic synergy produced by simultaneous mu-opioid activation at different levels of the neuraxis. Second, reflex-related activation of on-cells is mediated by a non-NMDA receptor, and this activation does not appear to play a significant role in regulating reflex responses to acute noxious stimuli. Excitatory amino acid-mediated excitation thus has at least two distinct roles within the RVM, activating off-cells and on-cells under different conditions.