Presynaptic Glutamate Receptors Regulate Noradrenaline Release from Isolated Nerve Terminals

The wide-ranging neuronal actions of excitatory amino acids, such as glutamate, are thought to be mediated mainly by postsynaptic N-methyl-D-aspartate (NMDA) and non-NMDA receptors. We now report the existence of presynaptic glutamate receptors in isolated nerve terminals (synaptosomes) prepared from hippocampus, olfactory bulb, and cerebral cortex. Activation of these receptors by NMDA or non-NMDA agonists, in a concentration-dependent manner, resulted in Ca(2+)-dependent release of noradrenaline from vesicular transmitter stores. The NMDA-stimulated release was potentiated by glycine and was blocked by Mg2+ and selective NMDA antagonists. In contrast, release stimulated by selective non-NMDA agonists was blocked by 6-cyano-7-nitroquinoxaline-2,3- dione, but not by Mg2+ or NMDA antagonists. Our data suggest that the presynaptic glutamate receptors can be classified pharmacologically as both the NMDA and non-NMDA types. These receptors, localized on nerve terminals of the locus ceruleus noradrenergic neurons, may play an important role in interactions between noradrenaline and glutamate.     

Presynaptic k-Opioid and Muscarinic Receptors Inhibit the Calcium-Dependent Component of Evoked Glutamate Release from Striatal Synaptosomes

In addition to cytosolic efflux, reversal of excitatory amino acid (EAA) transporters evokes glutamate exocytosis from the striatum in vivo. Both kappa-opioid and muscarinic receptor agonists suppress this calcium-dependent response. These data led to the hypothesis that the calcium-independent efflux of striatal glutamate evoked by transporter reversal may activate a transsynaptic feedback loop that promotes glutamate exocytosis from thalamo- and/or corticostriatal terminals in vivo and that this activation is inhibited by presynaptic kappa and muscarinic receptors. Corollaries to this hypothesis are the predictions that agonists for these putative presynaptic receptors will selectively inhibit the calcium-dependent component of glutamate released from striatal synaptosomes, whereas the calcium-independent efflux evoked by an EAA transporter blocker, L-trans-pyrrolidine-2,4-dicarboxylic acid (L-trans-PDC), will be insensitive to such receptor ligands. Here we report that a muscarinic agonist, oxotremorine (0.01-10 microM), and a kappa-opioid agonist, U-69593 (0.1-100 microM), suppressed the calcium-dependent release of glutamate that was evoked by exposing striatal synaptosomes to the potassium channel blocker 4-aminopyridine. The presynaptic inhibition produced by these ligands was concentration dependent, blocked by appropriate receptor antagonists, and not mimicked by the delta-opioid agonist [D-Pen2,5]-enkephalin. The finding that glutamate efflux evoked by L-trans-PDC from isolated striatal nerve endings was entirely calcium independent supports the notion that intact basal ganglia circuitry mediates the calcium-dependent effects of this agent on glutamate efflux in vivo. Furthermore, because muscarinic or kappa-opioid receptor activation inhibits calcium-dependent striatal glutamate release in vitro as it does in vivo, it is likely that both muscarinic and kappa receptors are inhibitory presynaptic heteroceptors expressed by striatal glutamatergic terminals.     

Interacting Presynaptic k-Opioid and GABAA Receptors Modulate Dopamine Release from Rat Striatal Synaptosomes

Abstract— The presynaptic regulation of stimulated dopa-mine release from superfused rat striatal synaptosomes by opioids and γ-aminobutyric acid (GABA) was studied. It was found that in addition to dopamine D₂ autoreceptors, calcium-dependent K⁺-stimulated [³H]dopamine release was inhibited through activation of a homogeneous population of k -opioid receptors in view of the potent inhibitory effect of the k -selective agonist U69.593 (EC₅₀ 0.2 nM) and its antagonism by norbinaltorphimine. Neither μ-nor δ-selective receptor agonists affected release of [³H]-dopamine. In addition, GABA potently inhibited the evoked [³H]dopamine release (EC₅₀ 0.4 nM) through activation of GABA_A receptors in view of the GABA-mimicking effect of muscimol, the sensitivity of its inhibitory effect to picro-toxin and bicuculline, and the absence of an effect of the GABA_B receptor agonist baclofen. In the presence of a maximally effective concentration of GABA, U69,593 did not induce an additional release-inhibitory effect, indicating that these receptors and the presynaptic D₂ receptor are colocalized on the striatal dopaminergic nerve terminals. The excitatory amino acid agonists N-methyl-d -aspartate and kainate, as well as the cholinergic agonist carbachol, stimulated [³H]dopamine release, which was subject to k -opioid receptor-mediated inhibition. In conclusion, striatal dopamine release is under regulatory control of multiple excitatory and inhibitory neurotransmitter by activation of colocalized presynaptic receptors for excitatory amino acids, acetylcholine, dopamine, dynorphins, and GABA within the dopaminergic nerve terminals. Together, these receptors locally control ongoing dopamine neurotransmission.     

Presynaptic Adenosine A2 and N-Methyl-D-Aspartate Receptors Regulate Dopamine Synthesis in Rat Striatal Synaptosomes

Dopamine synthesis rate and cyclic AMP concentration were measured in synaptosomes prepared from rat striatum. Dopamine synthesis rate was decreased by the addition of either adenosine deaminase or 8-phenyltheophylline, an adenosine receptor blocker, and was increased by the addition of 2-chloroadenosine. The addition of L-glutamate in the absence of adenosine deaminase decreased both dopamine synthesis rate and cyclic AMP concentration; in the presence of adenosine deaminase, glutamate had no effect on basal dopamine synthesis, but enhanced K(+)-stimulated synthesis. Both these effects of glutamate were abolished in Ca2(+)-free medium or in the presence of 2-amino-5-phosphonovalerate, an N-methyl-D-aspartate (NMDA) receptor blocker. In Mg2(+)-free medium with adenosine deaminase, glutamate enhanced both basal and K(+)-stimulated synthesis. These results suggest that dopaminergic terminals have A2 adenosine receptors, whose activation can stimulate dopamine synthesis by a cyclic AMP-dependent mechanism, and NMDA receptors, which modulate dopamine synthesis by a Ca2(+)-dependent mechanism.     

Nicotinic Modulation of [3H]Dopamine Release from Striatal Synaptosomes: Pharmacological Characterisation

Presynaptic nicotinic acetylcholine receptors on striatal nerve terminals modulate the release of dopamine. We have compared the effects of a number of nicotinic agonists and antagonists on a perfused synaptosome preparation preloaded with [3H]dopamine. (-)-Nicotine, acetylcholine, and the nicotinic agonists cytisine and 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), at micromolar concentrations, stimulated the release of [3H]dopamine from striatal nerve terminals. Carbamylcholine was a much weaker agonist. The actions of (-)-nicotine, cytisine, and DMPP were inhibited by low concentrations of the nicotinic antagonists dihydro-beta-erythroidine, mecamylamine, pempidine, and neosurugatoxin; alpha-bungarotoxin was without effect, and extending the time of exposure to this toxin resulted in only very modest inhibition. This pharmacology points to a specific nicotinic receptor mechanism that is clearly distinct from that at the neuromuscular junction. Atropine failed to antagonise the effects of acetylcholine and carbamylcholine, suggesting that no muscarinic component is involved. The nicotinic receptor ligands (-)-[3H]nicotine and 125I-alpha-bungarotoxin bound to specific sites enriched in the synaptosome preparation. Drugs tested on the perfused synaptosomes were examined for their ability to interact with these two ligand binding sites in brain membranes. The differential sensitivity to the neurotoxins alpha-bungarotoxin and neosurugatoxin of the 125I-alpha-bungarotoxin and (-)-[3H]nicotine binding sites, respectively, leads to a tentative correlation of the (-)-[3H]nicotine site with the presynaptic nicotinic receptor on striatal nerve terminals.     

Stereoselective Nicotine-Induced Release of Dopamine from Striatal Synaptosomes: Concentration Dependence and Repetitive Stimulation

Using a sensitive perfusion system we have studied the nicotine-induced release of [3H]dopamine ([( 3H]DA) from striatal synaptosomes. Nicotine-evoked release was concentration dependent with an EC50 of 3.8 microM. The response to 1 microM nicotine was comparable to that to 16 mM K+; 10 microM veratridine evoked a larger response. All three stimuli were Ca2+ dependent but only the response to veratridine was blocked by tetrodotoxin. Repetitive stimulations by 1 microM (-)-nicotine (100 microliters) at 30-min intervals resulted in similar levels of [3H]DA release; higher concentrations of (-)-nicotine resulted in an attenuation of the response particularly following the third stimulation. This may reflect desensitisation or tachyphylaxis of the presynaptic nicotinic receptor. The action of nicotine was markedly stereoselective: a 100-fold higher concentration of (+)-nicotine was necessary to evoke the same level of response as 1 microM (-)-nicotine. It is proposed that these presynaptic nicotinic receptors on striatal terminals are equivalent to high-affinity nicotine binding sites described in mammalian brain.     

Ionic Basis of Inhibitory Presynaptic Modulation in Rat Cortical Synaptosomes

We have investigated the possibility that, regardless of the involvement of a second messenger system, the ultimate effect of presynaptic, receptor-activated inhibitory modulation is the opening of a K channel. With the consequent hyperpolarization of the terminal, less Ca2+ would enter and this would result in the observed diminished release of a neurotransmitter. This possibility was explored utilizing rat cortical synaptosomes that were prelabeled with either 86Rb or [3H]acetylcholine, depolarizing with either K+ or veratridine, and measuring either efflux of 86Rb or release of [3H]acetylcholine in the presence or absence of inhibitory presynaptic modulators. The modulating agents used were 2-chloroadenosine, carbamylcholine, clonidine, and morphine. In all instances, these agents promoted an increased efflux of 86Rb, indicating hyperpolarization, and decreased release of acetylcholine. These results are compatible with our suggestion that an increase in K conductance may be responsible for presynaptic inhibition of the release of neurotransmitters.     

Glutamatergic Control of Dopamine Release in the Rat Striatum: Evidence for Presynaptic N-Methyl-D-Aspartate Receptors on Dopaminergic Nerve Terminals

The N-methyl-D-aspartate (NMDA) receptor-mediated regulation of the release of newly synthesized [3H]dopamine [( 3H]DA) was studied in vitro, both on rat striatal slices using a new microsuperfusion device and on rat striatal synaptosomes. Under Mg2(+)-free medium conditions, the NMDA (5 X 10(-5) M)-evoked release of [3H]DA from slices was found to be partly insensitive to tetrodotoxin (TTX). This TTX-resistant stimulatory effect of NMDA was blocked by either Mg2+ (10(-3) M) or the noncompetitive antagonist MK-801 (10(-6) M). In addition, the TTX-resistant NMDA-evoked response could be potentiated by glycine (10(-6) M) in the presence of strychnine (10(-6) M). The coapplication of NMDA (5 X 10(-5) M) and glycine (10(-6) M) stimulated the release of [3H]DA from striatal synaptosomes. This effect was blocked by Mg2+ (10(-3) M) or MK-801 (10(-5) M). These results indicate that some of the NMDA receptors involved in the facilitation of DA release are located on DA nerve terminals. These presynaptic receptors exhibit pharmacological properties similar to those described in electrophysiological studies for postsynaptic NMDA receptors.     

Serotonin Inhibits Acetylcholine Release from Rat Striatum Slices: Evidence for a Presynaptic Receptor-Mediated Effect

Rat brain striatum slices were incubated with [3H]choline, perfused with a physiological buffer, and stimulated by perfusion with a K+-enriched buffer for 2 min. The tritium overflow evoked by K+ was decreased by 5-hydroxytryptamine (serotonin, 5-HT) (maximal inhibition 10(-6) M). This effect of 5-HT was mimicked by several agonists (5-methoxytryptamine, N,N-dimethyl-tryptamine, bufotenin) and blocked by serotonergic antagonists (methiothepin, methysergide, cinanserin) but not by haloperidol; methiothepin and methysergide alone slightly increased the K+-evoked overflow of tritium (3H). Inhibition of the tritium release by 5-HT was not suppressed in the presence of tetrodotoxin (TTX) (10(-6) M). These results suggest that 5-HT tonically inhibits acetylcholine (ACh) release from striatal cholinergic neurons by acting on a presynaptic receptor localized on cholinergic terminals.     

Diacylglycerol-Induced Stimulation of Neurotransmitter Release from Rat Brain Striatal Synaptosomes

These studies were undertaken to test the hypothesis that alterations in phosphatidylinositol metabolism can modulate neurotransmitter release in the central nervous system. The effects of 1,2-diacylglycerols (DAGs) on dopamine release in the rat central nervous system were determined by measuring dopamine release from rat striatal synaptosomes in response to two DAGs (sn-1,2-dioctanoylglycerol and 1-oleoyl-2-acetylglycerol) that can activate protein kinase C and one DAG (deoxydioctanoylglycerol) that does not activate this kinase. Dioctanoylglycerol and 1-oleoyl-2-acetylglycerol, at a concentration of 50 micrograms/ml, stimulated the release of labeled dopamine from striatal synaptosomes by 35-50 and 17%, respectively. Dioctanoylglycerol-induced release was also demonstrated for endogenous dopamine. In contrast, deoxydioctanoylglycerol (50 micrograms/ml) did not stimulate dopamine release. Dioctanoylglycerol-induced dopamine release was independent of external calcium concentration, indicating a utilization of internal calcium stores. Dioctanoylglycerol (50 micrograms/ml) also produced a 38% increase in labeled serotonin release from striatal synaptosomes. The addition of dioctanoylglycerol to the striatal supernatant fraction increased protein kinase C activity. These results are consistent with the concept that an increase in phosphatidylinositol metabolism can stimulate neurotransmitter release in the central nervous system via an increase in DAG concentration. The data suggest an involvement of protein kinase C in the DAG-induced release, but other sites for DAG action are also possible.     

Characterisation of Inhibitory 5-Hydroxytryptamine Receptors That Modulate Dopamine Release in the Striatum

Abstract: The effect of a series of indoleamines on the potassium-evoked tritium release of previously accumulated [³H]dopamine from rat striatal slices has been investigated. The indoleamines 5-hydroxytryptamine, 5-methoxy-tryptamine, 5-methoxy- N, N' -dimethyltryptamine and tryptamine (10⁻⁷ to 10⁻³ M) all reduced potassium-evoked release of tritium, to a maximum of 50%. The uptake of [³H]dopamine was unaffected by these compounds. A series of 5-hydroxytryptamine antagonists were examined for their ability to reduce the inhibition of potassium-evoked tritium release induced by 5-methoxytryptamine. The relative order of antagonist potency obtained was methysergide > metergoline > methiothepin > cinanserin > cyproheptadine > mianserin, and was consistent with an action on 5-hydroxytryptamine receptors. It is concluded that there are inhibitory 5-hydroxytryptamine receptors located on the terminals of dopaminergic neurones in the striatum.     

Rapid Release of [3H]Dopamine from Median Eminence and Striatal Synaptosomes

Release of preaccumulated, tritium-labeled dopamine ([3H]DA) from preparations of isolated nerve terminals (synaptosomes) of rat median eminence (ME) and corpus striatum (CS) was examined over short time intervals (1-20 s). In both preparations, basal efflux of [3H]DA was linear with time. Depolarization with high K+ resulted in an initial rapid release of [3H]DA which stabilized by 20 s, whereas veratridine elicited an increased rate of release over basal levels that was linear over the first 20 s. The calculated rate constants of release for both the initial phase of K+- and the veratridine-stimulated release were approximately threefold greater in CS than in ME synaptosomes. The major component of the high K+-induced release of [3H]DA from both synaptosome preparations increased as a graded function of [Ca2+]o. However, a smaller component, independent of external Ca2+, existed in both ME and CS synaptosomes. Increasing the [Mg2+] in the external solution resulted in a right shift of both the [K+]o and the [Ca2+]o dose-response curves, consistent with actions of Mg2+ on screening surface membrane charges and blocking voltage-dependent Ca2+ channels. In all studies, steady-state uptake of the [3H]DA was about twofold greater into CS than into ME synaptosomes. Moreover, the fraction of incorporated [3H]DA released by stimulation from the CS was much greater than that released from ME synaptosomes. These data are consistent with differences between these two types of dopaminergic terminals with respect to packaging and/or distribution of the accumulated neurotransmitter in intraneuronal pools, as well as marked differences in the apparent kinetics of DA release.     

Metabotropic Glutamate Receptors Modulate Ischemia-Induced GABA Release in Mouse Hippocampal Slices

The involvement of glutamate receptors in GABA release in ischemia was investigated in hippocampal slices from adult (3-month-old) and developing (7-day-old) mice. For in vitro ischemia, the slices were superfused in glucose-free media under nitrogen. Ionotropic glutamate receptor agonists failed to affect the ischemia-induced basal GABA release at either age. The K(+)-stimulated release in the immature hippocampus was potentiated by N-methyl-D-aspartate receptors, whereas in adults this release was reduced by both kainate and 2-amino-3-hydroxy-5-methyl-4-isoxazoleproprionate receptor activation. The group I metabotropic receptor agonist (1+/-)-1-aminocyclopentane-trans-1,3-dicarboxylate enhanced the basal ischemic GABA release in a receptor-mediated manner in adults, this being concordant with the positive modulation of GABAergic neurotransmission by group I metabotropic glutamate receptors. (1 +/-)-1-Aminocyclopentane-trans-1,3-dicarboxylate and (S)-3,5-dihydroxyphenylglycine also enhanced the K(+)-stimulated release in the developing hippocampus in a receptor-mediated manner. Because group I receptors generally increase neuronal excitability, the enhanced GABA release may attenuate hyperexcitation or strengthen inhibition, being thus neuroprotective, particularly under ischemic conditions. Group III metabotropic glutamate receptors were not at all involved in ischemic GABA release in the immature mice, but in adults their activation by O-phospho-L-serine potentiated the basal release and reduced the K(+)-stimulated release. These opposite effects were abolished by the antagonist (RS)-2-cyclopropyl-4-phosphonophenylglycine. Metabotropic glutamate receptors, namely group I and III receptors, are able to modify the release of GABA from hippocampal slices under ischemic conditions, both positive and negative effects being discernible, depending on the age and type of receptor activated.     

Effects of Glutamate Antagonists on Methamphetamine and 3,4-Methylenedioxymethamphetamine-Induced Striatal Dopamine Release In Vivo

Abstract: Several amphetamine analogues are reported to increase striatal glutamate efflux in vivo, whereas other data indicate that glutamate is capable of stimulating the efflux of dopamine (DA) in the striatum via a glutamate receptor-dependent mechanism. Based on these findings, it has been proposed that the ability of glutamate receptor-blocking drugs to antagonize the effects of amphetamine may be explained by their capacity to inhibit DA release induced by glutamate. To examine this possibility further, we investigated in vivo the ability of glutamate antagonists to inhibit DA release induced by either methamphetamine (METH) or 3,4-methylenedioxymethamphetamine (MDMA). Both METH and MDMA increased DA efflux in the rat striatum and, in animals killed 1 week later, induced persistent depletions of DA and serotonin in tissue. Pretreatment with MK-801 or CGS 19755 blocked the neurotoxic effects of METH and MDMA but, did not significantly alter striatal DA efflux induced by either stimulant. Infusion of 6-cyano-7-nitroquinoxaline-2,3-dione into the striatum likewise did not alter METH-induced DA overflow, and none of the glutamatergic antagonists affected the basal release of DA when given alone. The findings suggest that the neuroprotective effects of NMDA antagonists do not involve an inhibition of DA release, nor do the data support the proposal that glutamate tonically stimulates striatal DA efflux in vivo. Whether phasic increases in glutamate content might stimulate DA release, however, remains to be determined.     

In vivo Effects of the Anatoxin-a on Striatal Dopamine Release

Anatoxin-a is an important neurotoxin that acts a potent nicotinic acetylcholine receptor agonist. This characteristic makes anatoxin-a an important tool for the study of nicotinic receptors. Anatoxin-a has been used extensively in vitro experiments, however anatoxin-a has never been studied by in vivo microdialysis studies. This study test the effect of anatoxin-a on striatal in vivo dopamine release by microdialysis.The results of this work show that anatoxin-a evoked dopamine release in a concentration-dependent way. Atropine had not any effect on dopamine release evoked by 3.5 mM anatoxin-a. However, perfusion of nicotinic antagonists mecamylamine and α-bungarotoxin induced a total inhibition of the striatal dopamine release. Perfusion of α7*-receptors antagonists, metillycaconitine or α-bungarotoxin, partially inhibits the release of dopamine stimulated by anatoxin-a. These results show that anatoxin-a can be used as an important nicotinic agonist in the study of nicotinic receptor by in vivo microdialysis technique and also support further in vivo evidences that α7*nicotinic AChRs are implicated in the regulation of striatal dopamine release.     

Adenosine and Glutamate Modulate Each Other's Release from Rat Hippocampal Synaptosomes

Abstract: In rat hippocampal synaptosomes, adenosine decreased the K⁺ (15 mM) or the kainate (1 mM) evoked release of glutamate and aspartate. An even more pronounced effect was observed in the presence of the stable adenosine analogue, R-phenylisopropyladenosine. All these effects were reversed by the selective adenosine A₁ receptor antagonist 8-cyclo-pentyltheophylline. In the same synaptosomal preparation, K⁺ (30 mM) strongly stimulated the release of the preloaded [³H]adenosine in a partially Ca²⁺-dependent and tetrodotoxin (TTX)-sensitive manner. Moreover, in the same experimental conditions, both l -glutamate and l -aspartate enhanced the release of [³H]adenosine derivatives ([³H]ADD). The gluta-mate-evoked release was dose dependent and appeared to be Ca²⁺ independent and tetrodotoxin insensitive. This effect was not due to metabolism because even the nonmetabolizable isomers d -glutamate and d -aspartate were able to stimulate [³H]ADD release. In contrast, the specific glutamate agonists N-methyl-d -aspartate, kainate, and quisqualate failed to stimulate [³H]ADD release, suggesting that glutamate and aspartate effects were not mediated by known excitatory amino acid receptors. Moreover, NMDA was also ineffective in the absence of Mg²⁺ and l -glutamate-evoked release was not inhibited by adding the specific antagonists 2-amino-5-phosphonovaleric acid or 6–7-dinitroquinoxaline-2, 3-dione. The stimulatory effect did not appear specific for only excitatory amino acids, as γ-anunobutyric acid stimulated [³H]ADD release in a dose-related manner. These results suggest that, at least in synaptosomal preparations from rat hippocampus, adenosine and glutamate modulate each other's release. The exact mechanism of such interplay, although still, unknown, could help in the understanding of excitatory amino acid neurotoxicity.     

Heterogeneity of N-Methyl-D-Aspartate Receptors Regulating the Release of Dopamine and Acetylcholine from Striatal Slices

In an attempt to examine some functional characteristics of the N-methyl-D-aspartate (NMDA) receptor complex, the NMDA-evoked effluxes of endogenous dopamine (DA) and [³H]acetylcholine ([³H]ACh) were simultaneously examined in a rat Striatal slice preparation. NMDA induced release of both DA and ACh in a concentration-dependent, Ca²⁺-, Mg²⁺-, and tetrodotoxin-sensitive manner. These release responses were remarkably reduced by long-term pre-treatment with a low concentration of NMDA. an indication of the desensitization of the NMDA receptor. Glycine was potent in reversing the desensitization-related reduction of DA release but failed to reverse the diminution of ACh release in the same slices. Our results indicate that the NMDA receptors regulating the release of DA and ACh are different with respect to their glycine modulatory site. This finding is consistent with a functional heterogeneity of the NMDA receptor complex in the rat striatum.     

Presynaptic α2 Adrenoceptors Inhibit Glutamate Release from Rat Spinal Cord Synaptosomes

Abstract: The presynaptic regulation of amino acid release from nerve terminals was investigated using synaptosomes prepared from the rat spinal cord. The basal releases of endogenous glutamate (Glu), aspartate (Asp), and γ-amino-butyric acid (GABA) were 34.6, 21.5, and 10.0 pmol/min/mg of protein, respectively. Exposure to a depolarizing concentration of KCl (30 m M ) evoked 2.7-, 1.5-, and 2.9-fold increases in Glu, Asp, and GABA release, respectively. Clonidine reduced the K⁺-evoked overflow of Glu to 56% of the control overflow with a potency (IC₅₀) of 17 n M , but it did not affect K⁺-evoked overflow of Asp, GABA, and their basal releases. Similarly, noradrenaline inhibited the K⁺-evoked overflow of Glu, although phenylephrine and isoproterenol showed no effect. The inhibitory effect of clonidine was counteracted by α₂-adrenoceptor antagonists, rauwolscine, yohimbine, and idazoxan, regardless of the imidazoline structures. Because Glu is considered a neurotransmitter of primary afferents that transmit both nociceptive and nonnociceptive stimuli in the spinal cord, these data suggest that part of Glu release may be regulated by the noradrenergic system through α₂ adrenoceptors localized on the primary afferent terminals.     

NMDA and Non-NMDA Ionotropic Glutamate Receptors Modulate Striatal Acetylcholine Release via Pre- and Postsynaptic Mechanisms

Abstract: The effects of NMDA and α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) on endogenous acetylcholine release from rat striatal slices and synaptosomes were investigated. Both agonists (1–300 µ M ) facilitated acetylcholine release from slices in a dose-dependent manner. NMDA (100–300 µ M ) and AMPA (30–300 µ M ), however, subsequently inhibited acetylcholine release. NMDA (100 µ M )-induced facilitation was antagonized by 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid (CPP) and dizocilpine (both 1–10 µ M ), whereas the 10 µ M AMPA effect was antagonized by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 1–30 µ M ). NMDA (100 µ M )-induced inhibition was counteracted by CPP, but not dizocilpine, and by the nitric oxide synthase inhibitor l -nitroarginine (1–100 µ M ). Tetrodotoxin (0.5 µ M ) prevented the facilitatory effect of 3 µ M NMDA and AMPA, but left unchanged that of 30 µ M NMDA and 100 µ M AMPA. Acetylcholine release from synaptosomes was stimulated by KCI (7.5–100 m M ) in a dose-dependent manner. NMDA and AMPA maximally potentiated the 20 m M KCl effect at 1 µ M and 0.01 µ M , but were ineffective at 100 µ M and 10 µ M , respectively. Inhibition of acetylcholine release was never found in synaptosomes. The effects of 1 µ M NMDA and 0.01 µ M AMPA were antagonized by CPP (0.0001–1 µ M ) or dizocilpine (0.0001–10 µ M ) and by CNQX (0.001–1 µ M ), respectively. These data suggest that glutamatergic control of striatal acetylcholine release is mediated via both pre- and post-synaptic NMDA and non-NMDA ionotropic receptors.     

Presynaptic Control of the Synthesis and Release of Dopamine from Striatal Synaptosomes: A Comparison Between the Effects of 5-Hydroxytryptamine, Acetylcholine, and Glutamate

The effects of 5-HT and glutamate on dopamine synthesis and release by striatal synaptosomes were investigated and compared with the action of acetylcholine, which acts presynaptically on this system. 5-HT inhibited (28%) synthesis of [¹⁴C]dopamine from L-[U-¹⁴C]tyrosine, at 10-⁵M and above. This contrasts with the action of acetylcholine, which stimulated [¹⁴C]-dopamine synthesis by 24% at 10-⁴ M. Tissue levels of GABA were unaffected by either 5-HT or acetylcholine up to concentrations of 10-⁴ M. The inhibitory action of 5-HT (5 × 10^?5 M and 2 × 10^?4 M) on [¹⁹C]dopamine synthesis was completely abolished by methysergide (2 × 10^?6 M). Higher concentrations of methysergide (10^?4 M) or cyproheptadine (10^?5 M) inhibited [¹⁴C]dopamine synthesis by 28% and 25%, respectively, when added alone to synaptosomes. However, only methysergide prevented the further inhibition of synthesis caused by 5-HT. At concentrations of 2 × 10^?5 M and above, 5-HT stimulated [¹⁴C]dopamine release. This releasing action differed from that of acetylcholine, which occurred at lower concentrations (e.g., 10^?6 M). Methysergide (up to 10^?4 M) or cyproheptadine (2 × 10^?4 M) did not reduce the 5-HT (5 × 10^?5 M)-induced release of [¹⁴C]dopamine, but methysergide (10^?4 M) showed a potentiation (49%) of this increased release. The stimulatory effects of 5-HT (2 × 10^?5 M) and K⁺ (56 mM) on [¹⁴C]dopamine release were additive, indicating that two separate mechanisms were involved. However, when both agents were present the stimulatory effect of K⁺ (56 mM) on [¹⁴C]dopamine synthesis was not seen above the inhibitory effect of 5-HT. Glutamate (0.1-5 mM) did not affect [⁴C]dopamine release or its synthesis from L-[U-¹⁴C]tyrosine. It is concluded that 5-HT modulates the synthesis of dopamine in striatal nerve terminals through a presynaptic receptor mechanism, an action antagonised by methysergide. The releasing action of 5-HT apparently occurs through a separate mechanism which is also distinct from that involved in the response to K⁺ depolarisation.