Dynamic changes of excitatory amino acid receptors in the rat hippocampus following transient cerebral ischemia

The changes in excitatory amino acid receptor ligand binding induced by transient cerebral ischemia were studied in the rat hippocampal subfields. Ten minutes of ischemia was induced by common carotid artery occlusion combined with hypotension, and the animals were allowed variable periods of recovery ranging from 1 day to 4 weeks. The binding of 3H-AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) to quisqualate receptors, 3H-kainic acid (KA) to kainate receptors, and 3H-glutamate to N-methyl-D-aspartate (NMDA) receptors as determined by quantitative autoradiography. One week following ischemia the CA1 region of the hippocampus displayed a severe (90%) dendrosomatic lesion with preservation of presynaptic terminals. This was associated with a 60% decrease in AMPA binding and a 25% decrease in glutamate binding to NMDA receptors. At 4 weeks postischemia, both AMPA and NMDA sites were greatly reduced. Although the dentate gyrus granule cells are resistant to an ischemic insult of this magnitude, this region showed marked changes in receptor binding. One week following ischemia, the AMPA and NMDA binding decreased by approximately 40 and 20%, respectively. Following 2 weeks of recovery, the NMDA binding was not significantly different from control level, while the AMPA binding remained depressed up to 4 weeks postischemia. The high density of KA binding sites in the inner molecular layer of the dentate gyrus was unaffected by the ischemic insult, despite an extensive degeneration of cells in the hilus of dentate gyrus which projects glutamatergic afferents to this area.(ABSTRACT TRUNCATED AT 250 WORDS)     

A quantitative estimate of the contribution made by various receptor categories to the depolarizations evoked by some excitatory amino acids in the olfactory cortex

A study has been undertaken to assess the percentage contributions made by N-methyl-D-aspartate (NMDA), kainate and quisqualate receptors to the composite depolarizations evoked by L-cysteate, L-cysteinesulphinate, L-homocysteate and S-sulpho-L-cysteine in the rat olfactory cortex slice. The percentage contribution made by NMDA receptors, which was quantified by measuring the reduction in agonist responses in the presence of the highly selective NMDA receptor antagonist 2-amino-5-phosphonopentanoate (0.1 mM), was: L-homocysteate, 73%; S-sulpho-L-cysteine, 65%; L-cysteate, 42% and L-cysteinesulphinate, 30%. Responses mediated by NMDA, kainate and quisqualate receptors were abolished by a 'desensitization' procedure involving repeated application of a mixture containing high concentrations of the selective agonists followed by perfusion of the non-selective receptor antagonist cis-2,3-piperidine dicarboxylate (5 mM). Following this procedure, responses to L-homocysteate and S-sulpho-L-cysteine were almost abolished and simple calculation gave the contribution of kainate plus quisqualate receptors to the agonist responses as: L-cysteinesulphinate, 46%; L-cysteate, 34%; S-sulpho-L-cysteine, 28% and L-homocysteate, 23%. However, approximately 24% of the composite depolarizations evoked by L-cysteate and L-cysteinesulphinate was mediated by a mechanism not involving NMDA, kainate or quisqualate receptors, neither did it reflect possible electrogenic uptake of the amino acids nor an interaction with 2-amino-4-phosphonobutyrate receptors. It is suggested that this fraction of the depolarizations evoked by L-cysteate and L-cysteinesulphinate might be due to a non-receptor-mediated release of K+ or, perhaps, to activation of an as yet unidentified receptor category.     

Some effects of excitatory amino acid receptor antagonists on synaptic transmission in the rat olfactory cortex slice

A study has been made of the effects of a series of excitatory amino acid receptor antagonists on the field potentials evoked on electrical stimulation of the lateral olfactory tracts of olfactory cortex slices perfused in vitro. The antagonists studied included (+/-)-2-amino-5-phosphonovaleric acid, a potent, specific antagonist of N-methyl-D-aspartate (NMDA) receptors, gamma-D-glutamylglycine, an antagonist of NMDA and kainate receptors and (+/-)-cis-2,3-piperidine dicarboxylic acid and 2-amino-4-phosphonobutyric acid, drugs which in addition to antagonizing NMDA and kainate receptors also block responses to quisqualic acid. From the patterns of effects of the drugs it is proposed that quisqualate and NMDA but not kainate receptors are involved in mediating excitatory transmission in the olfactory cortex; quisqualate receptors are located at the lateral olfactory tract - superficial pyramidal cell synapse whereas NMDA receptors are present at the synapses of the superficial pyramidal cell collaterals with the deep pyramidal cell dendrites and/or at the synapses of the pyramidal cell collaterals and inhibitory interneurones. The results are discussed in terms of possible presynaptic and/or postsynaptic sites of antagonist action.     

Long-term potentiation in the hippocampal CA1 area and dentate gyrus plays different roles in spatial learning

NMDA receptor-dependent long-term potentiation (LTP) at hippocampal synapses has been considered a crucial component of the cellular basis for learning and memory. This form of LTP occurs in excitatory synapses in both the CA1 area and the dentate gyrus in the hippocampus. However, differential roles of LTP in these areas have not yet been identified. To address this issue, we enhanced the degree of LTP by expressing Ca2+-permeable AMPA receptors at either hippocampal CA1 or dentate gyrus synapses using Sindbis viral vectors (SINs) encoding both green fluorescent proteins and unedited GluR2 (GluR2Q) subunits, and examined their effects on rat spatial learning. The viral vectors were locally injected into the 8-week-old-rat brain in vivo bilaterally. The postsynaptic expression of Ca2+-permeable AMPA receptors enhanced the degree of LTP, and induced NMDA receptor-independent LTP in the presence of the NMDA receptor antagonist in SIN-infected regions in both CA1 and dentate gyrus in hippocampal slice preparations. However, the regional expression of Ca2+-permeable AMPA receptors caused opposite behavioural consequences on the Morris water maze task: rats with SIN-infected CA1 pyramidal cells showed shorter escape latency and better probe test performance, whereas those with SIN-infected dentate gyrus granule cells showed impaired performance. Thus, it was demonstrated that CA1 and dentate gyrus synapses play different functional roles in spatial learning despite their similar mechanism for LTP induction.     

Kindling enhances kainate receptor-mediated depression of GABAergic inhibition in rat granule cells

Several lines of evidence indicate a substantial contribution of kainate receptors to temporal lobe seizures. The activation of kainate receptors located on hippocampal inhibitory interneurons was shown to reduce GABA release. A reduced GABA release secondary to kainate receptor activation could contribute to an enhanced seizure susceptibility. As the dentate gyrus serves a pivotal gating function in the spread of limbic seizures, we tested the role of kainate receptors in the regulation of GABA release in the dentate gyrus of control and kindled animals. Application of glutamate (100 micro m) in the presence of the NMDA receptor antagonist d-APV and the AMPA receptor antagonist, SYM 2206 caused a slight depression of evoked monosynaptic inhibitory postsynaptic currents (IPSCs) in control, but a substantial decrease in kindled dentate granule cells. The observation that kainate receptor activation altered paired-pulse depression and reduced the frequency of TTX-insensitive miniature IPSCs without affecting their amplitude is consistent with a presynaptic action on the inhibitory terminal to reduce GABA release. In kindled preparations, neither glutamate (100 micro m) nor kainate (10 micro m) applied in a concentration known to depolarize hippocampal interneurons led to an increase of the TTX-sensitive spontaneous IPSC frequency nor to changes of the postsynaptic membrane properties. Consistently, the inhibitory effect on evoked IPSCs was not affected by the presence of the GABAB receptor antagonist, CGP55845A, thus excluding a depression by an enhanced release of GABA acting on presynaptic GABAB receptors. The enhanced inhibition of GABA release following presynaptic kainate receptor activation favours a use-dependent hyperexcitability in the epileptic dentate gyrus.     

Acidic Amino Acids and Self-stimulation of the Prefrontal Cortex in the Rat: A Pharmacological Study

The effects of intraventricular and intracortical microinjections of acidic amino acid antagonists on self-stimulation (SS) of the medial prefrontal cortex (MPC) were investigated. Self-stimulation was measured by depressing a lever in a standard chamber. Spontaneous motor activity of the animal and SS of the contralateral non-injected MPC were used as control for non-specific effects of the drugs. Intraventricular microinjections of gamma-d-glutamylglycine (DGG), an antagonist of NMDA, kainate and quisqualate receptors, or 2-amino-5-phosphonovalerate (AP-5), a specific antagonist of NMDA receptors, produced a dose-related decrease of SS in the MPC. Spontaneous motor activity of the animal was not significantly affected. Unilateral microinjections into the medial prefrontal cortex of DGG or AP-5 produced a decrease of SS in the ipsilateral side while no effects were found on the contralateral MPC. On the contrary, intraventricular microinjections of gamma-d-glutamyltaurine (Glu-tau), an antagonist with more relative affinity for kainate and quisqualate receptors, produced a dose-related decrease of both self-stimulation and spontaneous motor activity of the rats. Moreover, intracortical microinjections of Glu-tau had no effect on self-stimulation of this cortical area. These results suggest that acidic amino acids through NMDA, but not kainate or quisqualate, receptors could be part of the neurochemical substrate underlying SS of the MPC in the rat.     

Effects of adrenal steroid manipulations and repeated restraint stress on dynorphin mRNA levels and excitatory amino acid receptor binding in hippocampus

Adrenal steroid and stress effects were determined in hippocampus on levels of dynorphin (DYN) mRNA, expressed in dentate gyrus, and excitatory amino acid receptors, measured in Ammon's horn and dentate gyrus. Adrenalectomy (ADX) decreased DYN mRNA levels in dentate gyrus and replacement with aldosterone (ALDO), a specific type I adrenal steroid receptor agonist, prevented the decrease. Ru28362, a specific type II receptor agonist, had no effect. Likewise, kainate receptor binding to the stratum lucidum and hilus region of dorsal hippocampus was decreased after ADX and this decrease was prevented by ALDO but not by Ru28362 treatment. Similar though smaller effects were found for CNQX binding to AMPA receptors but only in the dentate gyrus molecular or infra- and supragranular layers. Although corticosterone (CORT) treatment of intact rats (40 mg/kg for 3 weeks) elevated DYN mRNA levels in dentate gyrus, up to 14 days of daily restraint stress (1 or 6 h/day) had no significant effect. Neither CORT treatment nor repeated restraint stress altered NMDA and non-NMDA glutamate receptors in hippocampus. The results of this study showing ADX-induced decreases of DYN mRNA and CNQX binding in dentate gyrus and decreased kainate binding in mossy fiber terminal regions are consistent with morphological evidence showing that adrenal steroids maintain normal integrity and structure of dentate gyrus neurons and do so via type I adrenal steroid receptors. These same parameters are apparently not sensitive to chronic restraint stress although the effects of other stressors must be examined.     

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.     

Selective excitotoxic pathology in the rat hippocampus

The pattern of cell loss and neuronal degeneration resulting from multiple microinjections of N -methyl-D-aspartate (NMDA), ibotenate (IBO), quisqualate (QUIS), and kainate (KA) into hippocampus was studied, together with the protection provided by the NMDA antagonist 3-(±)-2-carboxypiperazin-4-yl-propyl-l-phosphonate (CPP). Histological evaluation was carried out after 7 days of survival. NMDA and IBO resulted in an extensive loss of all cells in the hippocampus including dentate gyrus, hilar cells, and CA3-CA1 pyramidal cells, but there was an absence of damage to areas and structures outside hippocampus. After QUIS and KA injections the hippocampal damage was limited to hilar cells in the dentate gyrus, CA3 pyramidal cells, and partial loss of CA1 cells; there was extensive extrahippocampal damage including entorhinal cortex, amygdala, layers III, V, and VI of ventral neocortex, olfactory areas, and various thalamic nuclei. CPP provided almost complete protection from the effects of intrahippocampal injections of NMDA and IBO, but did not affect the hippocampal cell loss found after QUIS and KA (with the exception of minor protection of some CA1 cells). CPP protected most extrahippocampal sites from the damage resulting from QUIS and KA, indicating that such excitotoxic cell death is indirect and involves NMDA receptor activation by an endogenous agent. The use of multiple microinjections as opposed to single injections allows a clearer interpretation of selective excitotoxic vulnerability.     

AMPA,kainate, and NMDA receptor densities in the hippocampus of untreated male rats and females in estrus and diestrus

Steroid hormones systematically affect numerous neuronal targets, thus influencing, in a permanent or a transitory manner, the way the brain reacts to external and internal stimuli. The hippocampus is an important brain region for learning and memory and the glutamatergic intrahippocampal pathway plays a major role in performing such functions. We applied quantitative in vitro receptor autoradiography to examine how the in vivo hormone milieu affects the densities of AMPA, kainate, and NMDA receptors in the hippocampus of adult male rats and females in estrus and diestrus. All three examined receptor types presented significant gender-specific differences in their densities. The hippocampus of male rats contains significantly more AMPA, kainate, and NMDA receptors than that of female rats. Female rats in diestrus have significantly higher AMPA receptor densities than female rats in estrus. AMPA changes occurred to the same extent in CA1-3 and in the dentate gyrus. Significant differences in the densities of NMDA receptors were observed in the CA1-3 regions, whereas kainate receptor differences were restricted to the CA1 region. These results further support that steroid hormones, through their modulation of AMPA and NMDA receptors, may be involved in the control of synaptic efficacy and, therefore, influence learning and memory.     

Regulation of prodynorphin gene expression in the hippocampus by glucocorticoids.

The regulation of prodynorphin gene expression by glucocorticoids in the hippocampus was examined in rats that were adrenalectomized (ADX) either 7, 30, 60 and 90 days prior to sacrifice. Peptide levels in the hippocampus of ADX rats were determined by radioimmunoassay and immunocytochemistry. Prodynorphin (PDYN) mRNA was measured by Northern blot analysis and in situ hybridization. A time-dependent decrease in dynorphin A(1-8)(DYN) levels in the hippocampus (18% at 7 days; 44% at 30 days; 58% at 60 days) of ADX rats was found, which was accompanied by a comparable decrease in the abundance of PDYN mRNA. An in situ hybridization analysis revealed that both the number of positively hybridized cells and the number of silver grains per cell were decreased in the dentate gyrus after ADX. The administration of dexamethasone after surgery reversed the peptide and mRNA attenuation induced by ADX. ADX had no effect on the expression of proenkephalin mRNA or [Met5]-enkephalin immunoreactivity in the hippocampus. Examination of thionin-counterstained tissue showed that the dentate granule cell layer was intact. The decrement of DYN expression in this system is proposed to have resulted from the removal of glucocorticoid input and not dentate granule cell loss. This study provides the strong evidence for a differential susceptibility of these two opioid peptides in the hippocampus to the removal of glucocorticoids. In addition, these data provide support for a potentially selective, glucocorticoid-permissive component in PDYN gene expression.     

Cardiovascular changes induced by the local application of glutamate-related drugs in the rat nucleus tractus solitarii

The effects of the local application of drugs acting on glutamatergic receptors in the nucleus tractus solitarii (NTS) were investigated in anesthetized rats. Unilateral microinjection of agonists (L-glutamate, L-aspartate, N-methyl-D-aspartate (NMDA) and quisqualate) produced a dose-dependent hypotension and bradycardia. The effects of NMDA were prevented by low doses of the selective NMDA-receptor antagonist, 2-amino-5-phosphonovalerate (2-APV), or by the mixed NMDA/kainate antagonist, gamma-D-glutamylglycine. The response to all agonists and the bradycardia which was elicited in response to the intravenous administration of phenylephrine (vagal reflex response) could be prevented by the local microinjection of the glutamate antagonists kynurenic acid (3 nmol) and 2-APV (10 nmol) into the NTS. The present data suggest that in the NTS, NMDA and quisqualate receptors are implicated in blood pressure reflex regulation.     

N-methyl-D-aspartate, quisqualate and kainate receptors are all involved in transmission of photic stimulation in the suprachiasmatic nucleus in rats.

In order to clarify the neuronal transmission mechanism of photic stimulation in the suprachiasmatic nucleus (SCN), the effects of agonists and antagonists for excitatory amino acid receptors on N-acetyltransferase (NAT) activity in the pineal gland were observed following the microinjection of drugs into both sides of the nuclei. N-Methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate, and kainate (which are selective agonists for three different subtypes, i.e. NMDA, quisqualate and kainate receptors, respectively) significantly decreased NAT activity similarly to the suppressive effect of light. Moreover, compared with a control group, all the groups pretreated with a selective competitive antagonist for NMDA receptor (D-2-amino-5-phosphonovalerate or 3-((+-)-2-carboxypiperazine-4-yl)-propyl-1-phosphonate), or a selective non-competitive antagonist for non-NMDA receptors (Joro spider toxin-3 or 1-naphthylacetyl spermine) partially blocked the suppressive effect of photic stimulation on NAT activity. These results suggest that NMDA, quisqualate and kainate receptors are all involved in mediating photic stimulation in the SCN.     

Electrophysiological evidence for the presence of ionotropic and metabotropic excitatory amino acid receptors on dopaminergic neurons of the rat mesencephalon: an in vitro study.

The actions of the ionotropic and metabotropic excitatory amino acid agonists AMPA, quisqualate, kainate, NMDA and trans-ACDP were studied by means of intracellular electrophysiological recordings from dopaminergic neurons of rat mesencephalon in brain slices. It was observed that all these agents evoked an inward current in cells which were voltage-clamped near the resting potential (-50, -60 mV). The membrane responses produced by AMPA, kainate and quisqualate were associated with an increase of the apparent input conductance while the responses induced by NMDA and trans-ACDP were associated with a decrease in the apparent input conductance. Therefore, stimulation of ionotropic and metabotropic amino acid receptors activates inward currents in the dopaminergic cells by different mechanisms.     

NMDA receptor activation during epileptiform responses in the dentate gyrus of epileptic patients.

We previously showed that a low frequency (1 Hz) train of perforant path stimulation evokes burst discharges in the dentate gyrus of hippocampal slices obtained from patients surgically treated for intractable temporal lobe epilepsy. We report here that multiple population spikes that characterize the burst discharge are blocked reversibly by the specific NMDA receptor antagonist, D-(-)-2-amino-5-phosphonovaleric acid (D-APV). The epileptiform discharge evoked in human dentate gyrus by stimulation trains of 1 Hz could be reproduced in the rat dentate gyrus in vitro by the same stimulation protocol but required the presence of low concentrations (0.2-0.6 mM) of extracellular magnesium. We suggest that low frequency orthodromic stimulation of dentate granule cells through the perforant path progressively evokes an increase in the activation of NMDA receptors resulting in burst discharges in tissue from epileptic patients.     

Receptor types mediating the excitatory actions of exogenous L-aspartate and L-glutamate in rat olfactory cortex

Changes in potential between the pial and cut surfaces of rat olfactory cortex slices evoked by N-methyl-D-aspartate (NMDA), quisqualate, kainate, L-glutamate and L-aspartate and also by gamma-aminobutyric acid (GABA) have been monitored using extracellular electrodes. All agonists produced a pial-negative potential response when superfused onto the pial surface, GABA, L-aspartate and L-glutamate being less potent than the others. Repeated applications of NMDA, but not of the other agonists, led to a progressive reduction in response to approximately 30% of the initial depolarization. The responses to NMDA (100 microM) were selectively abolished by (+/-)2-amino-5-phosphonopentanoic acid (APP; 100 microM) while depolarizations evoked by L-glutamate and L-aspartate (both at 10 mM) were only antagonized by 21 +/- 2 (n = 12) and 36 +/- 3 (n = 12) percent respectively (means +/- S.E.M.). gamma-D-Glutamylglycine (gamma-DGG; 1 mM) and (+/-)cis-2,3-piperidine dicarboxylate (cis-PDA; 2 and 5 mM), in addition to antagonizing responses to NMDA, also partially blocked quisqualate- and kainate-evoked depolarizations. When a mixture of APP (100 microM), gamma-DGG (1 mM) and cis-PDA (5 mM) was applied to preparations, although NMDA receptors were completely blocked and responses to both quisqualate and kainate antagonized by approximately 80%, L-glutamate and L-aspartate evoked depolarizations were only reduced by 51 +/- 7 (n = 4) and 49 +/- 4 (n = 4) percent respectively (means +/- S.E.M.). The results are discussed in terms of the contributions made by NMDA, quisqualate and kainate receptors to the composite responses evoked by L-aspartate and L-glutamate.     

Seizure-related changes in the glutamate R2 and R5 receptor genes expression in the rat hippocampal formation

Summary The expression of mRNA coding for AMPA selective glutamate (Glu) R2 receptor and kainate selective GluR5 receptor was studied in the rat hippocampal formation in two animal models of limbic seizures evoked by systemic administration of pilocarpine (400 mg/kg ip) or kainate (15 mg/kg ip). As shown by an in situ hybridization study, pilocarpine decreased the GluR2 flip mRNA level in CA1 and CA3 areas of the hippocampus after 3h and kainate after 24 h, e.g. at the time preceding neuronal degeneration. No changes in the GluR2 flop or GluR5 mRNA level were found in those regions. In the dentate gyrus, resistant to neurodegeneration, pilocarpine and kainate differentialy affected the expression of GluR2 and GluR5 mRNAs. After 72 h pilocarpine, but not kainate, increased the GluR2 flop mRNA level and decreased the flip one, which suggests attenuation of the GluR2 sensitivity. On the other hand, kainate, elevated the GluR2 flip and GluR5 mRNA level in the dentate gyrus after 72 h. All in all the above data suggest that changes in the GluR2 gene expression may play some role in the neuronal damage to vulnerable areas (CA1, CA3). However, differences in the kainate- and pilocarpine-induced changes in the dentate gyrus at the late time points indicate that alterations in the stoichiometry of GluR2 forms or GluR5 gene expression in this brain region are not a common causal factor responsible for delayed neuronal hyperexcitability.     

Action of 3-((±)-2-car☐ypiperazin-4-yl)-propyl-1-phosphonic aci (CPP): a new and highly potent antagonist of N-methyl-d-aspartate receptors in the hippocampus

A new compound, 3-((±)-2-car☐ypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), has been evaluated as an excitatory amino acid receptor antagonist using electrophysiological assays and radioligand binding. In autoradiographic preparations, CPP reduces l-[³H]glutama binding in regions of the hippocampus rich in N-methyl-d-aspartate (NMDA) receptors, but not in regions richin kainate sites. In isolated membrane fraction preparations, CPP displaces l-[³H]glutamate binding to NMDA sites, but does not compete with the binding of selective kainate or quisqualate site ligands. CPP potently reduces depolarizations produced by application of NMDA but not depolarizations produced by quisqualate or kainate. Its order of potency against excitatory amino acid-induced responses in the hippocampus is NMDA > homocysteate > aspartate > glutamate > quisqualate. CPP has no efect on lateral perforant path responses or on inhibition of these responses by 2-amino-4-phosphonobutyrate. Finally, at doses that do not affect Schaffer collateral synpatic transmission, CPP reversibly blocks the induction of long-term potentiation of Schaffer synaptic responses. This new compounds is, therefore, a higly selective brain NMDA receptor blocker, and the most potent such by nearly an order of magnitude.     

Effects of adenosine and gamma-aminobutyric acid A receptor antagonists on N-methyl-D-aspartate induced neurotoxicity in the rat hippocampus

This study investigated the modulatory actions of adenosine and gamma-aminobutyric acid (GABA) on several aspects of N-methyl-D-aspartate (NMDA)-induced neurotoxicity, including neuronal loss, atrophy, necrosis, and calcium accumulation in the hippocampus. For this purpose, we combined unilateral intrahippocampal injections of NMDA (24 nmoles) with acute injections of the selective A1 adenosine receptor antagonist DPCPX (0.03 pmoles), the selective adenosine A2a receptor antagonist CSC (1.5 pmoles), a combination of these two antagonists, and injections of the selective GABA A receptor antagonist bicuculline (60 pmoles). Fifteen days after NMDA injection, neuronal loss with preservation of architecture was observed in stratum oriens, pyramidale, radiatum, lacunosum-moleculare, and stratum moleculare of Ammon's horn, and in radial and granular layers of the dentate gyrus. NMDA plus vehicle also produced a small degree of brain tissue necrosis (holes in the structure) in four of five brains. Acute injections of CSC, but not DPCPX or bicuculline, significantly increased the extent of neuronal loss produced by NMDA plus vehicle. CSC in combination with NMDA induced significantly more necrosis than NMDA plus vehicle. A significant degree of atrophy was observed in the hippocampus after treatment with NMDA plus vehicle, and bicuculline significantly increased the magnitude of this atrophy. NMDA-induced calcium deposits were detected within the radiatum and lacunosum-moleculare layers of the hippocampus and in the hilus of the dentate, but not in the stratum oriens, stratum pyramidale, or in the granular layer of the dentate gyrus. However, treatment with the different antagonists did not significantly modify the magnitude of the NMDA-induced calcium deposits. These results reveal a selective vulnerability of certain areas of the hippocampus to the accumulation of calcium deposits, and a selective interaction between adenosine receptors and NMDA-induced neurotoxicity in the hippocampus.