The effect of methamphetamine on the release of glutamate from striatal slices

Summary One postulated role of dopamine in the striatum is to reduce neuronal activity in cortico-striatal glutamergic terminals. We investigated the effects of methamphetamine, which displaces dopamine, on glutamate release from rat striatal slices. Methamphetamine significantly reduced K⁺-stimulated (45 mM) glutamate release. In slices prepared from rats treated 8 days previously with methamphetamine there was enhanced (approximately 200%) release of glutamate. This study demonstrates that dopamine has a modulatory effect on glutamate release in the striatum.     

Endogenous GABA release from slices of rat cerebral cortex and hippocampus in vitro

Using a rapid, simple and sensitive radioreceptor assay, a Ca²⁺-dependent K⁺-evoked release of endogenous GABA was demonstrated from rat cortical and hippocampal slices in vitro. This evoked-release of endogenous GABA was similar to tha of [³H]GABA release (in its Ca²⁺ dependency) but differed from the latter in having a higher signal to noise level. Neither 5-HT nor a stable enkephalin analogue had any effect on endogenous GABA release from hippocampus slices.     

The effect of isoflurane on the release of [H]-acetylcholine from rat brain cortical slices

Volatile general anaesthetics are believed to affect synaptic transmission, but their actions in the central nervous system (CNS) remains unclear. Acetylcholine (ACh) is one of the most important neurotransmitter in the CNS and thus, it is possible that its release could be one of the targets for volatile anaesthetic action. However, the effects of these agents on the release of ACh are not yet fully understood. Rat brain cortical slices were loaded with [³H]-choline in order to study the effect of isoflurane on the release of [³H]-ACh from this preparation. Isoflurane (28, 43, 54, 95 and 182 nM) significantly increased the basal release of [³H]-ACh. This effect was independent of the extracellular sodium and calcium concentration but was decreased by tetracaine and dantrolene, inhibitors of Ca^2+-release from intracellular stores. These findings indicate that isoflurane may cause a Ca^2+-release from internal stores that increases [³H]-ACh release in rat brain cortical slices.     

Immediate and delayed effects of in vitro ischemia on glutamate efflux from guinea-pig cerebral cortex slices

Immediate and delayed effects of glucose deprivation, oxygen deprivation (hypoxia) and both oxygen and glucose deprivation (in vitro ischemia) on glutamate efflux from guinea pig cerebral cortex slices were studied. Immediate effects were evaluated by measuring changes of glutamate efflux during the metabolic insults. Delayed effects were evaluated by measuring the response of the tissue to a 50 mM KCl pulse applied 60 minafter the metabolic insults. Deprivation of glucose in the medium did not induce either immediate or delayed effects, while hypoxic condition produced an immediate slight stimulation of glutamate efflux without any delayed effect. Conversely, in vitro ischemia produced both immediate and delayed effects on glutamate efflux. During in vitro ischemia glutamate efflux dramatically increased in a calcium-independent and tetrodotoxin-sensitive manner; this effect was potentiated by a low sodium containing medium. The blockade of the sodium/potassium ATP_ase exchanger by ouabain caused a glutamate outflow similar to that induced by in vitro ischemia. On the whole, these data demonstrate the central role played by the sodium electrochemical gradient and by the membrane glutamate uptake system in the glutamate overflow induced by in vitro ischemia. Moreover, in slices previously exposed to both oxygen and glucose deprivation the effect of KCl on glutamate efflux was potentiated. This in vitro ischemia-induced delayed potentiation of neurotransmitter efflux, until now unreported in the literature, was found to be selectively restricted to glutamatergic structures and to be mainly due to an enhancement of the exocytotic component of glutamate release. © 1997 Elsevier Science B.V. All rights reserved.     

Facilitation of glutamate release from rat cerebral cortex nerve terminal by subanesthetic concentration propofol

Propofol is now the most commonly used intravenous anesthetic‐for general anesthesia and sedation because of its rapid onset and recovery. Besides the well‐known adverse effects of cardiovascular and respiratory depression, recent studies indicate that propofol may cause excitatory phenomena such as myoclonus, opisthotonus, and even seizure. However, the mechanisms of these excitatory effects of propofol have not been elucidated. Considering glutamate as the principle excitatory neurotransmitter in the central nervous system and excessive glutamatergic synaptic transmission can cause seizure, we examined the effect of propofol on the release of glutamate from rat cerebral cortex nerve terminals (synaptosomes). Results showed that subanesthetic concentration propofol facilitated 4‐aminopyridine (4‐AP), but not KCl‐ or ionomycin‐evoked glutamate release from nerve terminals. The facilitation of 4‐AP‐evoked glutamate release by propofol also occurred in the calcium chelation and significantly attenuated by glutamate transporter inhibitors, DL ‐threo‐β‐benzyloxyaspartic acid (DL ‐TBOA) and L ‐trans‐pyrrolidine‐2,4‐dicarboxylic acid (L ‐trans‐PDC). In addition, propofol increased 4‐AP‐evoked depolarization of the plasma membrane potential. Furthermore, protein kinase C (PKC) inhibition suppressed propofol‐mediated facilitation of glutamate release. These results suggest that subanesthetic concentration propofol facilitates glutamate release from rat cerebrocortical glutamatergic terminals by increasing nerve terminal excitability, likely through the activation of PKC pathway. This finding may provide an explanation for propofol‐induced excitatory phenomena. Synapse 63:773–781, 2009. © 2009 Wiley‐Liss, Inc.     

Mechanisms underlying the honokiol inhibition of evoked glutamate release from glutamatergic nerve terminals of the rat cerebral cortex

The effect of honokiol, an active component of Magnolia officinalis, on glutamate release from isolated nerve terminals (synaptosomes) was examined. Honokiol potently inhibited 4-aminopyridine (4-AP)-evoked glutamate release in a concentration-dependent manner, and this effect resulted from a reduction of vesicular exocytosis and not from an inhibition of Ca(2+)-independent efflux via glutamate transporter. The inhibitory action of honokiol was not due to decreasing synaptosomal excitability or directly interfering with the release process at some point subsequent to Ca(2+) influx, because honokiol did not alter the 4-AP-evoked depolarization of the synaptosomal plasma membrane potential or Ca(2+) ionophore ionomycin-induced glutamate release. Rather, examination of the effect of honokiol on cytosolic [Ca(2+)] revealed that the diminution of glutamate release could be attributed to a reduction in voltage-dependent Ca(2+) influx. Consistent with this, the honokiol-mediated inhibition of 4-AP-evoked glutamate release was completely prevented in synaptosomes pretreated with a wide-spectrum blocker of N-, P-, and Q-type Ca(2+) channels, omega-conotoxin MVIIC. In addition, honokiol modulation of 4-AP-evoked glutamate release appeared to involve a protein kinase C (PKC) signaling cascade, in so far as pretreatment of synaptosomes with the PKC inhibitors Ro318220 or GF109203X all effectively occluded the inhibitory effect of honokiol. Furthermore, honokiol attenuated 4-AP-induced phosphorylation of PKC. Together, these results suggest that honokiol effects a decrease in PKC activation, which subsequently attenuates the Ca(2+) entry through voltage-dependent N- and P/Q-type Ca(2+) channels to cause a decrease in evoked glutamate exocytosis. These actions of honokiol may contribute to its neuroprotective effect in excitotoxic injury.     

Somatostatin facilitates the serotonin release from rat cerebral cortex, hippocampus and hypothalamus slices

The effect of somatostatin on the release of serotonin (5-HT) from the superfused slices of rat cerebral cortex, hippocampus and hypothalamus preloaded with [³H]5-HT was examined. Somatostatin (0.38–1.53 μM) was found to facilitate dose-dependently the electrically- or high K⁺-stimulated release of [³H]5-HT. The order of the stimulatory potency of somatostatin on the evoked release in the 3 areas of the brain was cortex,hippocampus>hypothalamus.     

The effects of convulsant and anticonvulsant drugs on the release of radiolabeled GABA, glutamate, noradrenaline, serotonin and acetylcholine from rat cortical slices

A possible presynaptic site of action of convulsant and anticonvulsant drugs has been evaluated by studying their effect on depolarization-induced transmitter release using radiolabeled GABA, glutamate, noradrenaline, serotonin and acetylcholine. The antiepileptic diphenylhydantoin (DPH) inhibited the release of noradrenaline and serotonin at concentrations that are antiepileptic in vivo. The release of the other transmitters was affected only with higher concentrations. Phenobarbital (PhB) reduced the release of all transmitters studied at concentrations much above the levels that are considered antiepileptic in vivo. Comparison with the anesthetic barbiturate pentobarbital (PB) further indicated that the presynaptic effects of PhB were related to its sedative rather than antiepileptic properties. Diazepam and sodium valproate had little effect; only GABA release was slightly reduced with diazepam at the highest concentration studied. The convulsants penicillin and pentylenetetrazole both increased the release of glutamate at concentrations that induce epileptiform activity in vivo or in vitro. Other transmitter systems were differentially affected by the two convulsants. A small increase of noradrenaline and serotonin release was observed with penicillin, but not with pentylenetetrazole. A presynaptic site of action for some, but not all, epileptogenic and antiepileptic drugs probably exists in addition to other, postsynaptic mechanisms. Glutamate is probably a major excitatory neurotransmitter in the brain and many physiological studies have suggested a role of excitatory pathways in the generation of epileptiform activity. The observed increase of glutamate release in the presence of two epileptogenic drugs observed in this in vitro study is in agreement with such a hypothesis.     

Patterns of endogenous amino acid release from slices of rat and guinea-pig olfactory cortex

A study has been made of the effects of depolarizing stimuli on the release of endogenous amino acid neurotransmitter candidates (aspartate, glutamate, GABA and taurine) from in vitro preparations of rat and guinea pig olfactory cortex. Exposure of small cubes of olfactory cortex tissue from either species to potassium chloride (50 mM) was accompanied by a calcium-dependent release of aspartate, glutamate and GABA. A similar release pattern was evoked by protoveratrine A (100 μM) although the release was largely calcium-independent. Neither agent led to increased release of taurine. Electrical stimulation of the excitatory input (lateral olfactory tract) of freshly prepared, synaptically intact olfactory cortex slices of both species induced significant release of aspartate and GABA from the uncut pial surface and of aspartate, GABA and glutamate from the cut surface. Evoked taurine release occurred from both surfaces of rat olfactory cortex slices but no release was detected from guinea pig olfactory cortex slices. These patterns of release were unaffected by changes in stimulus frequency and were mimicked by protoveratrine A (100 μM) applied to one or other surface. Preincubation of slices from rats for 2 h led to loss of tissue amino acids and to changes in their release patterns; the presence of glutamine (5 mM) during preincubation prevented the loss of amino acids but did not alter their pattern of release. Because of the close similarities between both the electrophysiological properties and the patterns of amino acid release it is concluded that there is probably an identity of amino acid neurotransmitters (aspartate, glutamate and GABA) in rat and guinea pig olfactory cortex. The role of taurine in the rat olfactory cortex is unknown but would seem unlikely to be that of a neurotransmitter. The results are discussed: (i) in terms of the cellular origins of the released amino acids; and (ii) with respect to apparent experimental discrepancies which have appeared in the literature^3,10.     

Pre- and post-synaptic functions of kainate receptors at glutamate and GABA synapses in the rat entorhinal cortex

Oscillatory network activity in cortical areas is seen as vital to physiological processes of cognition, learning, and memory, and fundamental to disorders such as epilepsy. Increasing attention is being paid to the role of kainate receptors (KAr) in the generation of network oscillations and synchrony. The entorhinal cortex (EC) plays a key role in learning and memory, and is a major site of dysfunction in temporal lobe epilepsy. KAr have been implicated in oscillogenesis in the EC, but limited information is available concerning the physiological roles of KAr in synaptic transmission in this area. Here, we make a detailed analysis of KAr function in Layer III of the EC, a site known to be highly susceptible to oscillogenesis, using whole-cell patch clamp recording of evoked and spontaneous synaptic currents in rat brain slices. We demonstrate that KAr containing the GluK1-subunit act as facilitatory autoreceptors at glutamatergic synapses on pyramidal neurones in Layer III. In addition, GluK1-containing KAr mediate an excitatory drive at glutamatergic synapses on GABAergic interneurones. In contrast, a different KAr, which is likely to contain the GluK2-subunit mediates a slow postsynaptic excitation at glutamatergic synapses on principal neurones, and may also act as a heteroreceptor, facilitating GABA release at inhibitory terminals on principal neurones. Reducing [Mg(2+) ](o) , which we have previously shown can generate KAr-dependent slow network oscillations in Layer III, enhances both glutamate and GABA release. Both effects are partly sustained by increased activation of GluK1-containing KAr. Increased activation of the GluK1-containing autoreceptor also results in an enhancement of the postsynaptic response mediated by GluK2-containing receptors. Finally, spontaneous release of both transmitters shows a rhythmic periodicity in low-Mg, and, again, this is dependent on GluK1-containing KAr. The results show that KAr contribute a facilitatory function at multiple levels in the networks of the EC, and provide a basis for dissecting the role of these receptors in oscillogenesis in this area.     

The effect of ACPD on the responses to NMDA and AMPA varies with layer in slices of rat visual cortex

The effect of 1S,3R-aminocyclopentane dicarboxylic acid (ACPD) was measured on cells from various layers in slices of the rat visual cortex using whole-cell recording techniques. The position of the recorded cell was estimated by distance from pia to the layer VI/white matter boundary, and verified in 34/97 cells by staining with biocytin. Potentiation or depression of the responses to NMDA and AMPA by the metabotropic glutamate agonist ACPD was examined by iontophoresis of the drugs close to the cell body. Iontophoresis of ACPD had different effects in different layers. In layer VI, ACPD produced a substantial depolarization, which augmented the responses to NMDA and AMPA. In layer V, ACPD did not produce a significant depolarization, but potentiated the response to NMDA and AMPA. In layer IV, ACPD produced a small hyperpolarization, and depressed the response to NMDA. In layers II and III, the results were small and variable. Most recordings from stained cells were from pyramidal cells. Where recordings from non-pyramidal cells were obtained (3/34), results were the same as from pyramidal cells in the same layer. The same results were obtained when tetrodotoxin was in the bath solution. We conclude that the potentiation or depression of the response to NMDA and AMPA by ACPD varies with layer in rat visual cortex.     

Cation effects on taurine release from brain slices: comparison to GABA

The effects of cations on the spontaneous and potassium-stimulated taurine release from mouse cerebral cortex slices were assessed with an emphasis on the as yet unestablished calcium dependence of the stimulated release. Spontaneous and stimulated GABA release was analyzed for comparison. A depolarizing concentration (50 mM) of potassium ions caused an approximately 3.5-fold increase in taurine release and the omission of sodium a 6-fold enhancement. GABA release was increased by the same stimuli about 20- and 34-fold, respectively. Omission of calcium ions greatly enhanced basal taurine and GABA release when the medium was supplemented with the calcium chelator ethylenediaminetetraacetate. The potassium stimulation was then abolished, however, with taurine even more readily than with GABA. Magnesium and calcium ions had antagonistic effects on the stimulated release, more clearly with taurine than with GABA. Verapamil abolished the potassium stimulation of both taurine and GABA release, the latter being more sensitive. Although the stimulated taurine release was less in magnitude and had a slower time course than the GABA release, the results are not at variance with the possible neurotransmitter role of taurine.     

Postnatal development of presynaptic alpha-adrenoceptors in rat cerebral cortex: studies with brain slices and synaptosomes

The development of α-adrenoceptor mediated presynapic modulation of noradrenaline (NA) release in the rat cerebral cortex was studied during the first postnatal month by examining the effects of exogenous NA and phentolamine on K⁺-induced [³H]NA release from superfused slices or synaptosomes.Exogenous NA strongly inhibited [³H]NA release from cortex slices of newborn rats, indicating that presynaptic α-receptors exist already on the day of birth. In fact, at least up to and including the 17th day after birth, NA caused a stronger inhibition of [³H]NA release from neonatal than from adult cortex slices. However, with synaptosomal preparations the inhibitory effect of NA was found to be similar in neonatal and adult rats. Further analysis of the data obtained with brain slices suggested that the differences between neonatal and adult rats observed with regard to the presynaptic modulation of [³H]NA release could be ascribed to differences in the concentrations of released endogenous NA in the extracellular spaces of brain slices. This hypothesis was strengthened by the data obtained in experiments using cortex slices of adult rats in which the endogenous NA level was lowered by pretreatment with α-methylparatyrosine.     

Effects of morphine and naloxone on the K-stimulated release of methionine-enkephalin from slices of rat corpus striatum

Methionine-enkephalin (ME) released from superfused slices of rat corpus striatum was estimated by radioimmunoassay (RIA). The basal release of2.5 ± 0.2pmol/g/min (0.15% of content per min) was increased approximately 3-fold upon exposure of tissue to 30 mM K⁺ for 5 min. This increase in release was not observed in the absence of Ca²⁺. Both morphine (10⁻⁵ M) and (−)-naloxone (10−⁵ and 10−⁶ M) significantly depressed the release of ME evoked by 30 mM K⁺ but did not alter basal release. The +-isomer of naloxone, which lacks opiate antagonist activity, did not affect basal or evoked release. A consistent depression of release was not observed when 47 mM K⁺ was used to evoke the release of ME. The issue of whether a feedback mechanism controls the release of ME from the striatum cannot be resolved until it is known whether the effect of morphine and naloxone on ME release are mediated by opiate or non-opiate mechanisms.     

Continuous measurement of net potassium movements in rat brain cortex suspensions: Effects of glutamate, veratridine, creatine and other substances

Net K fluxes in in vitro suspensions of sliced rat brain cortex were studied by means of a K-sensitive electrode. When incubation was in 3 mM K, a net K efflux occurred. It could be resolved into two first-order rate constants: k1 = 0.486 min-1, and k2 = 0.0102 min-1, that originated from compartments that contained 18% and 82% of tissue K, respectively. k1 Was suppressed by tetrodotoxin (TTX), and k2 was increased 38-fold by veratridine. The latter effect was blocked by TTX, methylphenidate (1 mM), creatine (25 mM), apamin (50 nM), quinine (100 microM), verapamil (22 microM) or D-600 (38 microM). Net K loss was greatly increased by 1 mM ouabain, and enhanced by sodium azide plus iodoacetamide, but not by 0.1 M ethanol. Glutamate (5 mM) induced a considerable and rapid net uptake of K, while aspartate or N-methylaspartate increased K efflux.     

Release of endogenous dopamine from electrically stimulated slices of rat striatum

An experimental system is described for measuring the release of endogenous dopamine from electrically stimulated slices of rat striatum. Striatal slices were field-stimulated by two high frequency trains (S1 and S2) applied 10, 30 or 60 min apart. The quantities of dopamine released by the two stimuli were compared from slices incubated with and without dopamine's precursor, L-tyrosine. Sustained release of dopamine evoked by the two stimuli was shown to require the inclusion of tyrosine (50 microM) in the superfusate.     

Noradrenergic modulation of glutamate release in the cerebellum

The effect of alpha- and beta-adrenergic agonists has been studied on the release of newly synthesized [3H]glutamate and [3H]GABA from slices of rat cerebellum. The beta 2-adrenergic agonist salbutamol, and also noradrenaline in the presence of the alpha-adrenergic antagonist phentolamine, both potentiated the K+-evoked release of [3H]glutamate. This potentiation appears to be mediated by adenylate cyclase activation. No effects of beta-adrenergic stimulation were observed on [3H]GABA release. The alpha-adrenergic agonist clonidine inhibited both the [3H]glutamate and the [3H]GABA release evoked by K+. The results suggest that noradrenergic modulation of cerebellar activity may have a presynaptic as well as postsynaptic origin.     

Demonstration of hyperaccumulation of [18F]2-fluoro-2-deoxy-D-glucose under oxygen deprivation in living brain slices using bioradiography

To clarify the mechanism of hyperaccumulation of glucose in acute brain ischemia by PET, changes of glucose metabolism and mitochondrial electron transfer function were examined in living brain slices in vitro during control, hypoxic, and anoxic conditions by positron autoradiography using [(18)F]2-fluoro-2-deoxy-D-glucose ([(18)F]FDG) and [(15)O]oxygen. [(15)O]Oxygen fixation reflecting mitochondrial electron transfer function was reduced and [(18)F]FDG uptake reflecting glucose metabolism was increased in proportion to the strength of oxygen deprivation during anoxia and hypoxia. Mitochondrial electron transfer function decreased with no regional differences, whereas the glucose metabolism was the most enhanced in the hippocampus and thalamus. The enhanced glucose metabolism was associated with an increased glutamate efflux after hypoxia and anoxia. Glucose metabolism was also increased by the addition of glutamate and was attenuated by the N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 in the hippocampus and thalamus. The hyperaccumulation of glucose in acute brain ischemia was demonstrated in living brain slices using bioradiography with reduced mitochondrial electron transfer. The activation of NMDA receptors by glutamate during acute brain ischemia might be responsible for hyperutilization of glucose in the hippocampus and thalamus.     

Desentization to glutamate and aspartate in rat olfactory (prepyriform) cortex slice

Olfactory cortex brain slices were subject to multiple bath applications of either glutamate or aspartate. The effectiveness of these amino acids (measured by quantitating the amplitude of lateral olfactory tract-stimulated field potentials) was progressively reduced with each successive perfusion of the agonist. However, the effectiveness of the endogenous neurotransmitter recovered to control in each intervening wash period. Thus, repeated applications of glutamate or aspartate desensitized olfactory cortex receptors to these amino acids but did not desensitize the receptors to the endogenous transmitter. These data support the hypothesis that neither glutamate nor aspartate is the neurotransmitter released from the lateral olfactory tract onto pyramidal cells of the olfactory cortex.