Neuronal N-methyl-D-aspartate Research Articles

AMPA glutamate receptor antagonism reduces neurologic injury after hypothermic circulatory arrest

Pharmacologic inhibition of the N-methyl-D-aspartate (NMDA) glutamate receptor can reduce the neurologic injury associated with hypothermic circulatory arrest; however, other receptor subtypes, such as the alpha-amino-3-hydroxy-5-methylisoazole-4-propionic acid/kainate or AMPA/kainate subtype, may predominate in the adult brain. In this experiment, a selective AMPA antagonist, NBQX, was used in a canine survival model of hypothermic circulatory arrest. Twelve male dogs (20 to 25 kg) were placed on closed-chest cardiopulmonary bypass, subjected to 2 hours of hypothermic circulatory arrest at 18 degrees C, and rewarmed on cardiopulmonary bypass. All were mechanically ventilated and monitored for 20 hours before extubation and survived for 3 days. Six dogs received NBQX beginning 2 hours after arrest (3 mg/kg for 3 hours then 1.5 mg/kg for 2 hours). Control dogs received vehicle only. Neurologic recovery was assessed every 12 hours using a species-specific behavior scale that yielded a neurodeficit score ranging from 0 (normal) to 500 (brain dead). After sacrifice at 72 hours, brains were examined by receptor autoradiography and histologically for patterns of selective neuronal necrosis and scored blindly from 0 (normal) to 100 (severe injury). Dogs given NBQX had better neurologic function compared with controls (neurodeficit score, 58.6 +/- 15 versus 204 +/- 30; p < 0.004) and had less neuronal injury (18.2 +/- 3 versus 52.5 +/- 6; p < 0.004). Densitometric receptor autoradiography revealed preservation of neuronal NMDA receptor expression only in dogs given NBQX. These results suggest that antagonism of the non-NMDA glutamate receptor AMPA may be neuroprotective in adults after hypothermic circulatory arrest.

Phencyclidine inhibits epinephrine-stimulated platelet aggregation independently of high affinity [formula omitted] (NMDA)-type glutamatereceptors

The psychotomimetic analgesic phencyclidine (PCP), which binds to a high affinity site on the neuronal N- methyl- d-aspartate (NMDA)-sensitive glutamate receptor, has previously been found to bind to platelets with high affinity and to specifically delay the onset of epinephrine-stimulated platelet aggregation (Jamieson et al. (1992) Biochem. J. 285, 35–39). We have now shown that the rank of binding affinities of 14 synthetic PCP analogs at the high affinity binding site on platelets does not parallel the rank order of their affinities in binding to rat brain membranes, indicating that the high affinity PCP binding site in platelets is distinct from the neuronal NMDA receptor. The order of potency of six of these analogs in delaying the onset of epinephrine-stimulated platelet aggregation also did not parallel the rank order of their binding affinities for platelet or brain binding sites. These data indicate that the ability of PCP analogs to inhibit epinephrine-stimulated aggregation is not related to their ability to bind to the high affinity platelet PCP binding site. Furthermore, (+)MK-801, which binds to the same high affinity binding site in neurons as does PCP, failed to inhibit epinephrine-stimulated platelet aggregation, further suggesting that the site at which PCP acts in platelets is not related to the NMDA-type glutamate receptor. Further studies showed that 5-HT 2 receptors and effects on platelet secretion are not involved in PCP-mediated inhibition of epinephrine-induced platelet aggregation.

Chapter 30 Nitric oxide: cellular regulation and neuronal injury

A variety of mechanisms have been proposed for NO neurotoxicity, including inhibition of mitochondrial enzymes such as cis-aconitase, inhibition of the mitochondria1 electron transport chain, inhibition of ribonucleotide reductase, DNA damage, mono-adenosine diphosphate (ADP)-ribosylation, and S-nitrosylation of glyceraldehyde-3-phosphate dehydrogenase. Excessive stimulation of N-methyl-D-aspartate (NMDA) by glutamate leads to excess production of NO, which is toxic to neurons under a variety of experimental paradigms and pathological conditions. When excessive amounts of NO are formed in response to the actions of the excitatory neurotransmitter, glutamate, acting at NMDA receptors, NO mediates components of neuronal killing NMDA toxicity acting via NO may account for neural damage associated with acquired immunodeficiency syndrome (AIDS) dementia as well as neural damage in cerebral infarction. Strategies aimed at decreasing the formation of NO or identifying targets of NO'S action may have clinical relevance for the development of agents which can prevent or reverse these degenerative disorders.

Effects of in vitro ethanol and fetal ethanol exposure on glutathione stimulation of N-methyl-D-aspartate receptor function.

The present studies investigated the effects of glutathione (GSH; gamma-glutamylcysteinylglycine) and its oxidized form (GSSG) on neuronal N-methyl-D-aspartate (NMDA) receptor activation in both acute and chronic preparations of ethanol exposure. It was demonstrated using fura-2-loaded dissociated brain cells from newborn rat pups that both GSH and GSSG (0-4 mM) produced concentration-dependent increases in intracellular calcium similar to those produced by NMDA and other agonists of the NMDA receptor. GSH-stimulated calcium entry was not inhibited by low intoxicating concentrations of ethanol, which contrasts with ethanol's typical inhibitory effect on NMDA-stimulated receptor activation. Behavioral studies in adult rats demonstrated that ethanol-induced sleep times were significantly decreased when 10 microliters of GSSG (20 mM) were administered intracerebroventricularly approximately 5 min before an intraperitoneal injection of 20% (w/v) ethanol (3 g/kg). These findings suggest that the less potent effect of ethanol on GSH-stimulated calcium entry as well as the reduction in ethanol-induced sleep times may be related to the presence of glycine in the peptide. The glycine found in GSH may activate the glycine site and block or reduce ethanol's action on this site. It appears that although GSH may play an important role in the activation of the NMDA receptor, this action does not involve a process that is sensitive to acute ethanol exposure. In contrast, when rat pups were chronically exposed to ethanol via prenatal exposure before the fura-2 preparation, increases in NMDA- and GSH-stimulated calcium entry were significantly decreased relative to those in pair-fed and ad libitum-fed controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Molecular cloning, functional expression, and pharmacological characterization of an N-methyl-D-aspartate receptor subunit from human brain.

A cDNA encoding a full-length N-methyl-D-aspartate (NMDA) receptor subunit 1, hNR1, was isolated from a human brain cDNA library. The hNR1 cDNA encodes an open reading frame of approximately 2.7 kb that shares high homology with the rat brain NMDA receptor subunit 1 and the mouse zeta 1 subunit. The hNR1 sequence, however, diverges from the rodent and murine homologs near the C terminus, suggesting that they represent alternatively spliced messages of the same gene. Oocytes injected with cRNA synthesized from the hNR1 cDNA express glutamate and NMDA-activated currents in the presence of glycine. Currents are blocked by the NMDA-receptor-specific antagonists 2-amino-5-phosphovaleric acid and 7-chlorokynurenate, and the open channel blockers MK-801 and phencyclidine, by Mg2+ ions in a voltage-dependent manner, and by Zn2+. Expressed hNR1 homomeric receptor channels exhibit the high Ca2+ permeability characteristic of neuronal NMDA receptors. Therefore, the cDNA clone hNR1 codes for a human brain NMDA receptor subunit cognate to the rodent and murine brain NR1 subunits.

Pharmacological characterization of the glutamate receptor in cultured astrocytes

Cultured astrocytes from neonatal rat cerebral hemispheres are depolarized by the excitatory neurotransmitter glutamate. In this study we have used selective agonists of different neuronal glutamate receptor subtypes, namely, the N-methyl-D-aspartate (NMDA), kainate, and quisqualate type, to characterize pharmacologically the glutamate receptor in astrocytes. The agonists of the neuronal quisqualate receptor, alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionic acid (AMPA) and quisqualate, depolarized the membrane. Kainate, an agonist of the neuronal kainate receptor, depolarized astrocytes more effectively than quisqualate. Combined application of kainate and quisqualate depolarized astrocytes to a level which was intermediate to that evoked by quisqualate and kainate individually. Agonists activating the neuronal NMDA receptor, namely NMDA and quinolinate, were ineffective. Application of NMDA did not alter the membrane potential even in combination with glycine or in Mg2+-free solution, conditions under which neuronal NMDA receptor activation is facilitated. The nonselective agonists L-cysteate, L-homocysteate, and beta-N-oxalylamino-L-alanine (BOAA) mimicked the effect of glutamate. Dihydrokainate, a blocker of glutamate uptake, did not, and several antagonists of neuronal glutamate receptors only slightly affect the glutamate response. These findings suggest that astrocytes express one type of glutamate receptor which is activated by both kainate and quisqualate, lending further support to the notion that cultured astrocytes express excitatory amino acid receptors which have some pharmacological similarities to their neuronal counterparts.

Rat brain N-methyl-D-aspartate receptors expressed in Xenopus oocytes.

N-methyl-D-aspartate (NMDA) activates a class of excitatory amino acid receptor involved in a variety of plastic and pathological processes in the brain. Quantitative study of the NMDA receptor has been difficult in mammalian neurons, because it usually exists with other excitatory amino acid receptors of overlapping pharmacological specificities. Xenopus oocytes injected with messenger RNA isolated from primary cultures of rat brain have now been used to study NMDA receptors. The distinguishing properties of neuronal NMDA receptors have been reproduced in this amphibian cell, including voltage-dependent block by magnesium, block by the NMDA receptor antagonist D-2-amino-5-phosphonovaleric acid, and potentiation by glycine. This preparation should facilitate the quantitative study of the regulation of NMDA receptor activation and serve as a tool for purification of the encoding messenger RNA.