Stimulated Glutamate Release Research Articles

Up-regulation of neuropeptide Y-Y2 receptors in an animal model of temporal lobe epilepsy.

Receptor autoradiography with the Y2 receptor ligand 125I-peptide YY3-36 and in situ hybridization were applied to investigate changes in neuropeptide tyrosine-Y2 receptor expression after kainic acid-induced recurrent seizures in the rat hippocampus. In the strata oriens and radiatum of CA1 to CA3, which are densely innervated by Y2 receptor-bearing Schaffer collateral terminals, a transient 2-fold increase in Y2 receptor affinity was observed after 4-12 hr, with a later slow decline. No change was seen in Y2 mRNA expression in CA2/CA3 pyramidal cells, from which Schaffer collaterals originate. Conversely, in granule cells of the dentate gyrus, markedly elevated Y2 mRNA concentrations were observed (by 740% in the dorsal hippocampus) 24-48 hr after kainate injection. At the same time, a marked and lasting (up to 6 months) increase in the number of Y2 receptor sites (by 800%) was seen in the dentate hilus, which is innervated densely by mossy fibers. The early increase in Y2 receptor affinity in Schaffer collaterals was accompanied by a 60% decrease in the EC50 of peptide YY3-36 in inhibiting K(+)-stimulated glutamate release in hippocampal slices from kainic acid-treated rats. Our data indicate transient up-regulation of presynaptic Y2 receptors in Schaffer collaterals by a change in affinity and a permanent de novo synthesis of presynaptic Y2 receptors in granule cells/mossy fibers. These changes may cause augmented presynaptic inhibition of glutamate release from different hippocampal sites and, in conjunction with increased concentrations of neuropeptide tyrosine in mossy fibers, may represent an endogenous reactive anticonvulsant mechanism.

Cocaine and amphetamine preferentially stimulate glutamate release in the limbic system: studies on the involvement of dopamine.

The effects of cocaine and d-amphetamine on extracellular glutamate and aspartate levels in the nucleus accumbens, prefrontal cortex, and striatum were studied by in vivo microdialysis in awake, freely moving rats. In the nucleus accumbens, glutamate levels were stimulated by cocaine (15-30 mg/kg, i.p.), GBR 12909 (15 mg/kg, i.p.), and d-amphetamine (2 mg/kg, i.p.), while aspartate levels were not affected. The increase in nucleus accumbens glutamate levels following cocaine (30 mg/kg) was calcium-dependent and was blocked by pretreatment with dopamine antagonists; haloperidol (0.2 mg/kg, i.p.), SCH 23390 (0.02 mg/kg, i.p.), and raclopride (1 mg/kg, i.p.), as well as local 6-OHDA lesions of the nucleus accumbens. In the prefrontal cortex, glutamate levels were stimulated by both cocaine (15-30 mg/kg, i.p.) and d-amphetamine (2 mg/kg, i.p.), while aspartate levels were moderately stimulated by d-amphetamine only. The increase in prefrontal cortex glutamate levels following cocaine (30 mg/kg) was calcium-dependent and was blocked by pretreatment with SCH 23390 (0.02 mg/kg, i.p.), but not haloperidol (0.2 mg/kg, i.p.) or raclopride (1 mg/kg, i.p.). In the striatum, glutamate and aspartate levels were not affected by either cocaine (15-30 mg/kg, i.p.) or d-amphetamine (2 mg/kg, i.p.). These findings demonstrate that stimulants enhance glutamate release in limbic brain structures, nucleus accumbens, and prefrontal cortex, but not extrapyamidal brain structures, striatum. Furthermore, the increase in glutamate release in the nucleus accumbens may be mediated by dopamine.

Cocaine and amphetamine preferentially stimulate glutamate release in the limbic system: Studies on the involvement of dopamine

Cocaine and amphetamine preferentially stimulate glutamate release in the limbic system: Studies on the involvement of dopamine

Nociceptin induced inhibition of K+ evoked glutamate release from rat cerebrocortical slices.

Nociceptin, an endogenous ligand for the orphan receptor ORL1, has recently been described. In this study we have shown that nociception inhibits 46 mM K(+)-stimulated glutamate release from rat perfused cerebrocortical slices with an IC50 of 51 nM. At 100 nM the inhibition amounted to 68 +/- 14% and was naloxone (10 microM)-insensitive excluding an activation of mu, delta and kappa opioid receptors. These data demonstrate the functional coupling of ORL1 in glutamatergic neurones and implicates a role for nociceptin in glutamatergic neurotransmission.

Ageing is associated with changes in glutamate release, protein tyrosine kinase and [formula omitted] protein kinase II in rat hippocampus

We have used synaptosomes prepared from rat hippocampus to investigate the role of protein tyrosine kinase and Ca 2+ calmodulin-dependent protein kinase II in modulating glutamate release in young animals and to investigate possible parallel age-related changes in release and kinase activity. We report that depolarization of synaptosomes with 40 mM KCl, which stimulated glutamate release, also significantly increased activity of both kinases, while the protein tyrosine kinase inhibitor, genistein and the Ca 2+ calmodulin-dependent protein kinase II inhibitor, KN62 (1-( N,O-bis[5-isoquinolinesulfonyl]- N-methyl-tyrosyl)-4-phenylpiperaxine) decreased K +-stimulated, Ca 2+-dependent release of glutamate. K +-stimulated release of glutamate was significantly decreased in hippocampal synaptosomes prepared from aged, compared to young, animals. In parallel with these changes in release, we report an age-related decrease in activities of both protein tyrosine kinase and Ca 2+ calmodulin-dependent protein kinase II. We conclude that these kinases play a role in modulating release of glutamate in hippocampus and that the age-related decrease in glutamate release may be partly due to an age-related decrease in kinase activities.

Glutamate: a potential mediator of inorganic mercury neurotoxicity.

Exposure to mercury vapor (Hg0) produces neurotoxic effects which are for the most part subsequent to its biotransformation in brain to the mercuric cation (Hg2 +), which has an exceptionally strong affinity towards the SH groups in proteins. However, neurologic symptoms are often encountered in subjects in which Hg+ concentration in the brain remains in the submicromolar range, markedly below the anticipated threshold for direct inhibition of cerebral metabolism and function. In this report we review biochemical and morphological evidence obtained in this and other laboratories in tissue culture studies suggesting that in such instances mercury neurotoxicity may be mediated by excitotoxic activity of glutamate (GLU). Mercuric chloride (MC) at 1 microM concentration (or less) inhibits GLU uptake and stimulates GLU release in cultured astrocytes, which in vivo is likely to result in excessive GLU accumulation in the extracellular space of the CNS. Inhibition of GLU uptake and stimulation of GLU release by MC may be attenuated by addition to the cultures of a cell membrane-penetrating agent dithiothreitol (DTT) but not of glutathione (GSH), which is not transported to the inside of the cells. However, MC-stimulated release of GLU is suppressed when the intracellular GSH levels are increased by metabolic manipulation. The results indicate that the MC-vulnerable SH groups critical for GLU transport are located within the astrocytic membranes. Ultrastructural evidence for GLU-mediated MC neurotoxicity came from studies in an organotypic culture of rat cerebellum. We have shown that: 1) 1 microM MC lowers the threshold of GLU neurotoxicity, 2) the combined neurotoxic effect of GLU plus MC is attenuated by DTT but not by GSH, which is consistent with the involvement of impaired astrocytic GLU transport, and 3) neuronal damage induced by GLU plus MC becomes less accentuated in a medium with dizocilpine (MK-801), a noncompetitive NMDA receptor antagonist.

Rapid chelation of calcium entering isolated rat brain nerve terminals during stimulation inhibits neurotransmitter release

The intracellular actions of calcium chelators on the release of the neurotransmitter glutamate from isolated rat brain nerve terminals (synaptosomes) were examined. Preloading synaptosomes with the rapid calcium-binding chelator 1,2-bis(2-aminophenoxy)ethane- N,N,N′,N′- tetraacetic acid (BAPTA) resulted in a decrease in K +-stimulated glutamate release to nearly half that of controls, whereas preloading with the calcium chelator EGTA, whose action is less rapid than that of BAPTA, was without effect. Inhibition of glutamate release was also observed on preloading synaptosomes with dibromo-BAPTA, but not with dinitro-BAPTA. K +-stimulated, Ca 2+-dependent synaptosomal protein phosphorylation was not affected after preloading with BAPTA. The results suggest that the calcium-dependent intracellular component essential for triggering the secretory response in mammalian brain nerve terminals resides near the calcium channels, binding; calcium rapidly on its entry during stimulation.

Evidence for sensitization of cocaine-induced nucleus accumbens glutamate release.

Cocaine-stimulated glutamate release in the nucleus accumbens was studied following chronic cocaine or saline pretreatment in order to determine whether this effect was sensitized in rats showing augmented dopamine release, locomotor and stereotypy responses. Rats were pretreated with cocaine (30 mg kg-1) or saline for 5 consecutive days and were tested with cocaine (15 mg kg-1) after a 10-day withdrawal period. Cocaine-induced glutamate release, dopamine release, horizontal locomotor activity and stereotypy were monitored simultaneously in animals undergoing in vivo microdialysis in the nucleus accumbens. The basal levels of extracellular glutamate and dopamine, as well as locomotor activity, were not different between cocaine- and saline-pretreated groups. Following cocaine injection the increase in glutamate release, dopamine release, locomotor activity and stereotypy were greater in the cocaine-pretreated animals. These results show that cocaine-stimulated glutamate release in the nucleus accumbens is sensitized following chronic cocaine pretreatment.

Neuroligand-evoked calcium-dependent release of excitatory amino acids from cultured astrocytes.

The release of excitatory amino acids (EAAs) from neuron-free cultures of neocortical astrocytes was monitored using HPLC. The neuroligand bradykinin caused a dose-dependent receptor-mediated increase in release of the EAAs glutamate and aspartate from type 1 astrocyte cell cultures obtained from rat cerebral cortex. Removal of calcium from the extracellular fluid prevented the bradykinin-induced release of EAAs from astrocytes. The addition of the calcium ionophore ionomycin caused a calcium-dependent release of EAAs. Inhibitors of the glutamate transporters p-chloromercuriphenylsulfonic acid, L-trans-pyrrolidine-2,4-dicarboxylate, and dihydrokainate failed to impair the ability of bradykinin to stimulate glutamate release from astrocytes. alpha-Latrotoxin, an active compound of black widow spider venom, caused a significant increase of the release of glutamate in calcium-containing saline. In calcium-depleted saline, alpha-latrotoxin produced an initial increase in the concentration of glutamate followed by a decline in the concentration of glutamate indicating stimulation of exocytosis coupled with low calcium-induced inhibition of endocytosis. Taken together, these data suggest that astrocytes may release neurotransmitter through a mechanism that is similar to the neuronal secretory process. Given the important role of glutamate in the induction of long-term potentiation, learning, memory, and excitotoxicity, it will be important to determine external signals that control both the uptake and release of glutamate by astrocytes.

A novel action of lithium: Stimulation of glutamate release and inositol 1,4,5 trisphosphate accumulation via activation of the N-methyl D-aspartate receptor in monkey and mouse cerebral cortex slices

Beginning at therapeutic concentrations (1–1.5mM), the anti-manicdepressive drug, lithium, stimulated the release of the major excitatory central neurotransmitter, glutamate, in monkey cerebral cortex slices in a time- and concentration-dependent manner, and this was associated with increased inositol 1,4,5-trisphosphate [Ins(1,4,5)P 3] accumulation. (±)-3-(2-Carboxypiperazin-4-yl)-propyl-1-phosphoric acid (CPP), dizocilpine (MK-801), ketamine, and Mg 2+-antagonists to the N-methyl D-aspartate (NMDA) receptor/channel complex selectively inhibited lithium-stimulated Ins(1,4,5)P 3 accumulation. Antagonists to cholinergic-muscarinic, α 1-adrenergic, 5-HT 2-serotoninergic and H 1-histaminergic receptors had no effect. Antagonists to non-NMDA glutamate receptors had no effect on lithium-stimulated Ins(1,4,5)P 3 accumulation. Possible reasons for this are discussed. Similar results were obtained in mouse cerebral cortex slices. Carbetapentane, which inhibits glutamate release, inhibited lithium-induced Ins(1,4,5)P 3 accumulation in this model. It is concluded that the primary effect of lithium in the cerebral cortex slice model is stimulation of glutamate release, which, via activation of the NMDA receptor, leads to Ca 2+ entry. Ca 2+ entry, in turn, activates phospholipase C. These effects may have relevance to the therapeutic action of lithium in the treatment of manic-depression, as well as its toxic effects, especially at lithium blood levels above 1.5mM. A general conclusion which can be drawn from these studies and earlier studies in our laboratory is that lithium potentiates the action of phospholipase C, whether this enzyme is activated by lithium-induced presynaptic release of a neurotransmitter, such as glutamate, or by the addition of an exogenous neurotransmitter, such as acetylcholine. However, this does not appear to be due to a direct activation of phospholipase C.

Mediators of injury in neurotrauma: intracellular signal transduction and gene expression.

Membrane lipid-derived second messengers are generated by phospholipase A2 (PLA2) during synaptic activity. Overstimulation of this enzyme during neurotrauma results in the accumulation of bioactive metabolites such as arachidonic acid, oxygenated derivatives of arachidonic acid, and platelet-activating factor (PAF). Several of these bioactive lipids participate in cell damage, cell death, or repair-regenerative neural plasticity. Neurotransmitters may activate PLA2 directly when linked to receptors coupled to G proteins and/or indirectly as calcium influx or mobilization from intracellular stores is stimulated. The release of arachidonic acid and its subsequent metabolism to prostaglandins are early responses linked to neuronal signal transduction. Free arachidonic acid may interact with membrane proteins, i.e., receptors, ion channels, and enzymes, modifying their activity. It can also be acted upon by prostaglandin synthase isoenzymes (the constitutive prostaglandin synthase PGS-1 or the inducible PGS-2) and by lipoxygenases, with the resulting formation of different prostaglandins and leukotrienes. Glutamatergic synaptic activity and activation of postsynaptic NMDA receptors are examples of neuronal activity, linked to memory and learning processes, which activate PLA2 with the consequent release of arachidonic acid and platelet-activating factor (PAF), another lipid mediator. Both mediators may exert presynaptic and postsynaptic effects contributing to long-lasting changes in glutamate synaptic efficacy or long-term potentiation (LTP), PAF, a potential retrograde messenger in LTP, stimulates glutamate release. The PAF antagonist BN 52021 competes for receptors in presynaptic membranes and blocks this effect. PAF may also be involved in plasticity responses because PAF leads to the expression of early response genes and subsequent gene cascades. The PAF antagonist BN 50730, selective for PAF intracellular binding, blocks PAF-mediated induction of gene expression. A consequence of neural injury induced by ischemia, trauma, or seizures is an increased release of neurotransmitters, that in turn generates an overproduction of second messengers. Glutamate, a key player in excitotoxic neuronal damage, triggers increased permeation of calcium mediated by NMDA receptors and activation of PLA2 in postsynaptic neurons. NMDA receptor antagonists reduce the accumulation of free fatty acids and elicit neuroprotection in ischemic damage. Increased production of free arachidonic acid and PAF converges to exacerbate glutamate-mediated neurotransmission. These neurotoxic actions may be brought about by arachidonic acid-induced potentiation of NMDA receptor activity and decreased glutamate reuptake. On the other hand, PAF stimulates the further release of glutamate at presynaptic endings. The neuroprotective effects of the PAF antagonist BN 52021 in ischemia-reperfusion are due, at least in part, to an inhibition of presynaptic glutamate release. PAF also induces expression of the inducible prostaglandin synthase gene, and PAF antagonists selective for the intracellular sites inhibit this effect. The PAF antagonist also inhibits the enhanced abundance, due to vasogenic cerebral edema and ischemia-reperfusion damage, of inducible prostaglandin synthase mRNA in vivo. Therefore, PAF, an injury-generated mediator, may favor the formation of other cell injury and inflammation mediators by turning on the expression of the gene that encodes prostaglandin synthase.

The involvement of multiple calcium channel sub-types in glutamate release from cerebellar granule cells and its modulation by GABAB receptor activation.

In this study, we have examined both the ability of various Ca2+ channel sub-types to support the release of [3H]glutamate from cerebellar granule neurons and the mechanism of action involved in the modulation of glutamate release by the GABAB receptor agonist, (-)-baclofen. Cerebellar granule neurons were stimulated to release newly synthesized [3H]glutamate by K(+)-evoked depolarization. Stimulated release was entirely calcium-dependent and abolished by the presence of 200 microM cadmium. Release of glutamate was not affected by either tetrodotoxin or 5-aminophosphonovaleric acid but was potentiated by dihydrokainate and inhibited by 6-cyano-7-nitroquinoxaline-2,3-dione. Stimulated glutamate release was partially inhibited by both the L-type calcium channel blocker, nicardipine, and the N-type calcium channel blocker, omega-conotoxin GVIA; however, the P/Q-type calcium channel blocker omega-agatoxin IVA inhibited release of glutamate only after pre-incubation of cells with omega-conotoxin GVIA. K(+)-stimulated release of glutamate was observed when stimulated either in the presence of Ca2+ or of Ba2+ and similar inhibition of release by (-)-baclofen was seen under both conditions. In contrast to these results, ionomycin-evoked glutamate release was greatly reduced as compared to K(+)-evoked release and was not modulated by (-)-baclofen. In the presence of omega-conotoxin GVIA alone, inhibition of release by (-)-baclofen was attenuated but not abolished. Following block of nicardipine-sensitive channels, inhibition of release by (-)-baclofen was still present, and after prior block of omega-conotoxin GVIA-sensitive channels the presence of nicardipine restored the ability of (-)-baclofen to inhibit residual release of glutamate. Modulation of glutamate release by (-)-baclofen was unaffected by the presence of omega-agatoxin IVA alone; however, after block of both omega-conotoxin GVIA- and omega-agatoxin IVA-sensitive channels, inhibition of release by (-)-baclofen was completely abolished. These results indicate that multiple sub-types of voltage-dependent calcium channels are present on the presynaptic terminals of cerebellar granule neurons and support K(+)-stimulated release of [3H]glutamate. Modulation of release by GABAB receptor activation appears to be dependent upon interaction of this receptor with a number of voltage-sensitive calcium channels, including omega-conotoxin GVIA-sensitive and omega-agatoxin IVA-sensitive channels.

Prostaglandin E 2 stimulates glutamate release from synaptosomes of rat spinal cord

We recently reported that intrathecal administration of prostaglandin E 2 induced hyperalgesic and allodynic effects through the glutamate receptor system. Here we examined whether prostaglandin E 2 could evoke amino acid release from nerve terminals using rat spinal cord synaptosomes. Exposure in superfusion to prostaglandin E 2 significantly increased endogenous glutamate and aspartate release and dose dependencies showed bell-shaped patterns with a peak at 1 nM. Both releases were almost absolutely Ca 2+-dependent. These results demonstrate that prostaglandin E2 may stimulate the release of excitatory amino acids presynaptically in the spinal cord.

Studies of four novel diphenylbutylpiperazinepyridyl derivatives on release and inhibition of reuptake of dopamine, serotonin and noradrenaline by rat brain in vitro

Four novel diphenylbutylpiperazinepyridyl derivatives (FG5865 (2-[4-[4,4-bis(4-fluorophenyl)butyl]-1-piperazinyl]-3-pyridinecarboxamide), FG5891 (2-[4-[4,4-bis(4-fluorophenyl)butyl]-1-piperazinyl]- N-methyl-3-pyridinecarboxamide), FG5893 (2-[4-[4,4-bis(4-fluorophenyl)butyl]-1-piperazinyl]-3-pyridinecarboxylic acid methyl ester) and FG5909 (2-[4-[4,4-bis(4-fluorophenyl)butyl]-1-piperazinyl]-3-hydroxypyridine) were tested concerning their effects on in vitro release and reuptake of neurotransmitters. Serotonin, noradrenaline and dopamine were those considered, with rat synaptosomes prepared respectively from frontal cortex, cortex and striatum. FG5865, FG5891, FG5893 and FG5909 were found to potently inhibit the uptake of all three neurotransmitters. In addition, FG5865, FG5891 and FG5893 increased the release of serotonin and dopamine from perfused frontal cortical and striatal tissue; FG5865 was most potent in this regard. The release induced by the FG compounds was, however, much less than that induced by e.g. fenfluramine or amphetamine. All four FG compounds were also found to inhibit glutamate-stimulated release of dopamine from striatal tissue; for FG5893 this inhibition occurred at nanomolar concentrations.

Modulation of glutamate release by a κ-opioid receptor agonist in rodent and primate striatum

The influence of the κ-opioid receptor agonist, enadoline, on endogenous glutamate release was investigated in rat and marmoset striatal synaptosomes. Enadoline decreased 4-aminopyridine (2 mM)-stimulated glutamate release (rat: IC 50 ∼ 8.7 μM, marmoset: IC 50 ∼ 2.9 μM). The effect of enadoline was reversed by nor-binaltorphimine (5 μM). These data indicate that, in the striatum of the rat and marmoset, κ-opioid receptor agonists can modulate glutamate release. These findings may have implications for the treatment of Parkinson's disease.

D-serine, an endogenous synaptic modulator: localization to astrocytes and glutamate-stimulated release.

Using an antibody highly specific for D-serine conjugated to glutaraldehyde, we have localized endogenous D-serine in rat brain. Highest levels of D-serine immunoreactivity occur in the gray matter of the cerebral cortex, hippocampus, anterior olfactory nucleus, olfactory tubercle, and amygdala. Localizations of D-serine immunoreactivity correlate closely with those of D-serine binding to the glycine modulatory site of the N-methyl-D-aspartate (NMDA) receptor as visualized by autoradiography and are inversely correlated to the presence of D-amino acid oxidase. D-Serine is enriched in process-bearing glial cells in neuropil with the morphology of protoplasmic astrocytes. In glial cultures of rat cerebral cortex, D-serine is enriched in type 2 astrocytes. The release of D-serine from these cultures is stimulated by agonists of non-NMDA glutamate receptors, suggesting a mechanism by which astrocyte-derived D-serine could modulate neurotransmission. D-Serine appears to be the endogenous ligand for the glycine site of NMDA receptors.

α-Latrotoxin stimulates glutamate release from cortical astrocytes in cell culture

The mechanism responsible for the ability of bradykinin to cause calcium-dependent release of glutamate from astrocytes in vitro was investigated. The glutamate transport inhibitor, dihydrokainate, did not block bradykinin-induced glutamate release, and bradykinin did not cause cell swelling. These data exclude the involvement of glutamate transporters or swelling mechanisms as mediating glutamate release in response to bradykinin. α-Latrotoxin (3 nM), a component of black widow spider venom, stimulated calcium-independent glutamate release from astrocytes. Since α-latrotoxin induces vesicle fusion and calcium-independent neuronal neurotransmitter release, our data suggest that astrocytes may release neurotransmitter using a mechanism similar to the neuronal secretory process.

The role of adenosine A1 receptor activation in ethanol-induced inhibition of stimulated glutamate release in the hippocampus of the fetal and adult guinea pig

The role of adenosine A 1 receptor activation in ethanol-induced inhibition of stimulated l-glutamate (Glu) release was determined in transverse hippocampal slices of the near-term fetal guinea pig and the adult guinea pig. Exposure of the slices to 48 mM ethanol inhibited K +-stimulated Glu efflux. Pretreatment with 8-cyclopentyltheophylline (CPT), a selective adenosine A 1 receptor antagonist, blocked the ethanol-induced inhibition of K +-stimulated Glu efflux in the near-term fetal and adult hippocampus. In the near-term fetus, 2-chloro-N 6-cyclopentyladenosine (CCPA), a selective adenosine A 1 agonist, and exogenous adenosine each blocked K +-stimulated Glu efflux similar to that produced by 48 mM ethanol. In the adult, although K + increased Glu efflux in the presence of CCPA or adenosine, the magnitude of increase was less than that of the K +-stimulated Glu efflux for the control condition. Exposure to ethanol alone or ethanol plus CPT produced a transient increase in endogenous adenosine efflux in the near-term fetal and adult hippocampus, which was not temporally related to the ethanol-induced inhibition of K +-stimulated Glu efflux. Overall, the data indicate that adenosine A 1 receptor activation mediates ethanol-induced inhibition of stimulated Glu release in the hippocampus of the near-term fetal and adult guinea pig.

Glutamate efflux from rat brain slices and cultures: A comparison of the depolarizing agents potassium, 4-aminopyridine, and veratrine

The major excitatory amino acid neurotransmitter in the mammalian brain is glutamate (GLU). GLU release from nerve terminals is both calcium-dependent and -independent, yet these mechanisms of release are not fully understood. Potassium, 4-aminopyridine (4-AP) and veratrine are commonly used depolarizing agents that were studied for their ability to stimulate GLU efflux from brain slices. These agents produced significant regional variations in GLU efflux from rat brain slices. Potassium was the most potent of the three secretogogues tested. 4-AP produced a significant GLU efflux only in the cerebellum. Veratrine produced consistent stimulation of GLU efflux from all brain regions tested. Potassium was the only depolarizing agent tested that stimulated GLU release from primary astroglial cultures of rat cerebral cortex. All three agents also demonstrated an ability to inhibit GLU reuptake in brain slice preparations. This data suggest that both GLU release and uptake are modulated in a regionally selective manner, and that commonly used depolarizing agents affect not only calcium-dependent neuronal release, but also uptake and glial responses.

The release of glutamate and aspartate from rat brain synaptosomes in response to domoic acid (amnesic shellfish toxin) and kainic acid.

Kainic acid is known to stimulate the release of glutamate (GLU) and aspartate (ASP) from presynaptic neurons. It has been suggested that the enhanced release of these endogenous EAA's plays a significant role in the excitotoxic effects of KA. Domoic acid (DOM), a shellfish toxin, is structurally similar to KA, and has been shown to be 3-8 times more toxic than KA. In this study, effects of KA and DOM on the release of GLU and ASP from rat brain synaptosomes were investigated. Amino acid analysis was performed by the reversed phase HPLC, following derivatization with 9-fluorenylmethyl chloroformate (FMOC). Potassium chloride (40 mM) was used as a positive control, and stimulated GLU release from rat brain synaptosomes in presence or absence of Ca2+. DOM enhanced the release of GLU, whereas KA stimulated the release of both GLU and ASP from synaptosomes in the presence of Ca2+. However, their potency to stimulate GLU and ASP release was enhanced in absence of Ca2+. These results indicate that different mechanisms may be involved in the release of GLU and ASP in response to DOM and KA, and that neurotransmitter release appeared to be highly specific for these agonists. It would appear that DOM and KA may interact with different receptors on the presynaptic nerve terminal, and/or activate different subtypes of voltage-dependent Ca2+ channels to promote influx of Ca2+ which is targeted for different pools neurotransmitters.