Inhibitory effects of cis- and trans-resveratrol on noradrenaline and 5-hydroxytryptamine uptake and on monoamine oxidase activity

This study investigated for the first time the potential effects of cis- and trans-resveratrol (c-RESV and t-RESV) on noradrenaline (NA) and 5-hydroxytryptamine (5-HT) uptake by synaptosomes from rat brain, on 5-HT uptake by human platelets, and on monoamine oxidase (MAO) isoform activity. Both c-RESV and t-RESV (5-200 microM) concentration-dependently inhibited the uptake of [3H]NA and [3H]5-HT by synaptosomes from rat brain and the uptake of [3H]5-HT by human platelets. In both experimental models, t-RESV was slightly more efficient than c-RESV. Furthermore, in synaptosomes from rat brain, the RESV isomers were less selective against [3H]5-HT uptake than the reference drug fluoxetine (0.1-30 microM). On the other hand, both c-RESV and t-RESV (5-200 microM) concentration-dependently inhibited the enzymatic activity of commercial (human recombinant) MAO isoform (MAO-A and MAO-B) activity, c-RESV being slightly less effective than t-RESV. In addition, both RESV isomers were slight but significantly more selective against MAO-A than against MAO-B. Since the principal groups of drugs used in the treatment of depressive disorders are NA/5-HT uptake or MAO inhibitors, under the assumption that the RESV isomers exhibit a similar behaviour in humans in vivo, our results suggest that these natural polyphenols may be of value as structural templates for the design and development of new antidepressant drugs with two important biochemical activities combined in the same chemical structure: NA/5-HT uptake and MAO inhibitory activity.     

Effects of Antidepressant Drug Treatments on α2-Adrenoceptor Control of [3H]Noradrenaline Release from Hypothalamic Synaptosomes

KCl (16 mM) stimulated the release of [3H]noradrenaline ([3H]NA) from rat hypothalamic synaptosomes in a Ca2+-dependent manner; this release was attenuated by clonidine (0.01-100 microM). Changes in the release of [3H]NA and the functional status of alpha 2-adrenoceptors in the medial hypothalamus of rats treated acutely and chronically with clorgyline (1 mg/kg/day) or desipramine (DMI, 10 mg/kg/day) were assessed using superfused synaptosomes in which the attenuating effects of clonidine (1 microM) or the potentiating effects of yohimbine (1 microM) on K+-evoked release of [3H]NA were measured. After acute administration of DMI, significantly less [3H]NA was accumulated into synaptosomes. Although total (spontaneous + K+-evoked) [3H]NA release from these synaptosomes was unchanged, a significant reduction was apparent in the K+-evoked release from the DMI-treated tissue. Attenuation of K+-evoked release by clonidine was abolished in both these acute treatment groups. Following the chronic antidepressant drug regimens, [3H]NA uptake into DMI-treated tissue remained significantly reduced although total percent and K+-evoked [3H]NA release were unchanged. The K+-evoked release of [3H]NA in S1 was significantly enhanced (by 22%) in the clorgyline treatment group. Attenuation of K+-evoked [3H]NA release by clonidine in both chronic antidepressant-treated tissues was not significantly changed. It is concluded that the functional sensitivity of alpha 2-adrenoceptors on nerve endings in the medial hypothalamus is unchanged by these chronic antidepressant drug regimens. In synaptosomes from untreated tissue, yohimbine significantly potentiated K+-evoked release of [3H]NA; this effect was unchanged after acute regimens and reduced after chronic administration of both the antidepressants.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of psychotropic substances on dopamine uptake by synaptosomes and glial cells]

Dophamine-H3 uptake by synaptosomes and glial cells of the cerebral cortex in rabbits and by the synaptosomes of the cerebral cortex in rats was studied. KM value of the dophamine-H3 uptake was found to be the same (0.075 +/- +/- 0.01 micrometer) in case of synaptosomes and glial cells. The rate of dophamine-H3 uptake by the rabbit cerebral cortex synaptosomes was twice that of the gial cells (as compared by protein). Among the psychotropic agents studied the most active symaptosome uptake inhibitors were phenamine and cocaine; they were less active in respect to glial uptake. Both types of uptake were inhibited by neuroleptics and antidepressants to the same degree. Results of this work conformed to the view on the participation of the strial dophaminergic link in the action mechanism of psychostimulants.     

Uptake of L-glutamine into synaptosomes. Is the γ-glutamyl cycle involved?

Transport of L-glutamine into rat cortical synaptosomes has been investigated by (14C)L-glutamine uptake experiments. This amino acid enters synaptosomes both by an active carrier mediated system, which may be the result of gamma-glutamyl cycle activity and by a Na+-dependent transport system. This view is supported by the following observations: a) as demonstrated previously (10), glutamine inside synaptosomes reaches concentrations higher than those of the incubation medium, and initial rates of uptake approach saturation kinetics; b) the uptake of glutamine is inhibited by uncouplers; c) the uptake is inhibited by methionine sulfoximine, a suicide-inhibitor of an enzyme of the gamma-glutamyl cycle; d) the initial rate of uptake is lowered by decreasing the Na+-level of the incubation medium or by adding ouabain. The validity of this hypothesis is discussed.     

Inhibition of serotonin uptake in synaptosomes and glia cells by some pharmacological substances

The uptake of serotonin -14 C by glial cells and synaptosomes of the rabbit brain cortex was studied. The Km value of the uptake of serotonin -14 C proved to be equal (0.83 + 0.02 microM) both for synaptosomes and glial cells. Synaptosomes of the rabbit brain cortex take up serotonin -14 C twice as fast as glial cells (uptake rates were compared from protein). Among psychotropic drugs studied the tricyclic antidepressant imipramine and psychostimulant cocaine turned out the most active inhibitors of both synaptosomal and glial uptake of serotonin -14 C. The drugs in 50 microM concentration inhibit the uptake of serotonin -14 C in synaptosomes and glial cells by 90 and 75-80%, respectively.     

Plant-Derived Neurotoxic Amino Acids (β-N-Oxalylamino-l-Alanine and β-N-Methylamino-l-Alanine): Effects on Central Monoamine Neurons

In the present study the subacute effects of beta-N-oxalylamino-L-alanine (BOAA) and beta-N-methylamino-L-alanine (BMAA) on CNS monoamine neurons in rats were investigated following intracisternal injections or local intracerebral administration into substantia nigra. In vitro effects of BOAA and BMAA on high-affinity synaptosomal uptake of dopamine (DA), noradrenaline (NA), and serotonin (5-HT) were also examined. Intracisternal administration of BMAA decreased NA levels in hypothalamus, whereas no effects were seen on DA or 5-HT levels. Following intranigral injections of BOAA, NA levels tended to decrease in several regions, whereas the DA levels and the levels of DA metabolites were unaffected in all regions analyzed. Loss of tyrosine hydroxylase (TH) immunoreactivity in the intranigral injection sites and the presence of TH-immunoreactive pyknotic neurons near the borders of the injection sites were observed following both BOAA and BMAA treatments. Furthermore, substance P-immunoreactive terminals in substantia nigra pars reticulata were also found to have disappeared within the lesioned area following either BOAA or BMAA injections. Incubations with both BOAA and BMAA (10(-5) M) reduced high-affinity [3H]NA uptake in cortical synaptosomes to 69% and 41% of controls, respectively, whereas the striatal high-affinity [3H]DA uptake and the cortical high-affinity [3H]5-HT uptake were unaffected by BOAA or BMAA. The results demonstrate that both BOAA and BMAA can affect central monoamine neurons, although the potency and specificity of these substances on monoamine neurons when administered acutely into cerebral tissue or liquor cerebri seem to be low. However, the in vitro studies indicate selective effects of both compounds on NA neurons in synaptosomal preparations.     

Effect of Lithium and of Other Drugs Used in the Treatment of Manic Illness on the Cation-Transporting Properties of Na+, K+-ATPase in Mouse Brain Synaptosomes

We have developed and used a novel technique to investigate the effects of lithium and other psychotropic drugs on the cation-transporting properties of the sodium- and potassium-activated ATPase enzyme (Na+,K+-ATPase) in intact synaptosomes. Rubidium-86 uptake into intact synaptosomes is an active process and is inhibited by approximately 75% in the presence of the Na+,K+-ATPase inhibitor acetylstrophanthidin. In vitro addition of lithium to synaptosomes prepared from untreated mice causes a progressive inhibition of acetylstrophanthidin-sensitive 86Rb uptake, but only at concentrations higher than the clinical therapeutic range. However, pretreatment of mice for 14 days in vivo with lithium, carbamazepine, and haloperidol, but not phenytoin, causes a significant stimulation of 86Rb uptake into synaptosomes via Na+,K+-ATPase.     

DEVELOPMENTAL AND OTHER ASPECTS OF [35S]CYSTEINE TRANSPORT BY RAT BRAIN SYNAPTOSOMES

Abstract— Cysteine uptake by rat brain synaptosomes occurs by active transport. The uptake by synaptosomes isolated from newborn brain is slower and the concentration gradient achieved is lower than that observed in adult tissue. Synaptosomal fractions from both adult and newborn rat brains accumulate cysteine by two saturable systems. The calculated parameters show that the maximum rates of cysteine uptake in adult synaptosomes are approximately twice that observed in newborn synaptosomes for both the high and low affinity systems. The uptake by the high affinity system is sodium dependent and is inhibited by glycine and dibasic amino acids. Uptake by synaptosomes from 14-day-old animals is close to that observed in adult tissue. The uptake of cysteine differs greatly from that of cystine since the oxidized form, cystine, is taken up more slowly by systems with low affinities which are sodium independent, do not interact with dibasic amino acids and are independent of age.     

Chronic nicotine causes functional upregulation of ionotropic glutamate receptors mediating hippocampal noradrenaline and striatal dopamine release

It has been proposed that (-)-nicotine can activate release-stimulating presynaptic nicotinic acetylcholine receptors (nAChRs) on glutamatergic nerve terminals to release glutamate, which in turn stimulates the release of noradrenaline (NA) and dopamine (DA) via presynaptic ionotropic glutamate receptors on catecholaminergic terminals. The objective of this study was to compare the function of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methylisoxazide-4-propionic acid (AMPA) glutamate receptors in synaptosomes of rat hippocampus and striatum following acute and chronic (-)-nicotine administration. In hippocampal synaptosomes, prelabeled with [3H]NA, both the NMDA- and AMPA-evoked releases were higher in (-)-nicotine-treated (10 days) than in (-)-nicotine-treated (1 day) or vehicle-treated (1 or 10 days) rats. In striatal synaptosomes prelabeled with [3H]DA, the NMDA-evoked, but not the AMPA-evoked, release of [3H]DA was higher in (-)-nicotine-treated (10 days) than in nicotine-treated (1 day) or vehicle-treated (1 or 10 days) animals. Chronic (-)-nicotine did not affect catecholamine uptake, basal release and release evoked by high-K+ depolarization. Thus, chronic exposure to nicotine enhances the function of ionotropic glutamate receptors mediating noradrenaline release in the hippocampus and dopamine release in the striatum.     

Studies on the Role of α2-Adrenoceptors in the Control of Synaptosomal [3H]5-Hydroxytryptamine Release: Effects of Antidepressant Drugs

The sensitivity of alpha 2-adrenoceptors on 5-hydroxytryptamine (5-HT) nerve endings obtained from rat cerebral cortex was investigated following treatment with the antidepressant drugs desipramine (10 mg/kg/day for 21-28 days) or clorgyline (1 mg/kg/day for 21-28 days). [3H]5-HT (100 nM) was used to load cortical synaptosomes (P2) after experiments with uptake inhibitors confirmed that this concentration of amine ensured exclusive uptake into 5-HT nerve terminals. The sensitivity of K+-stimulated release of [3H]5-HT to alpha 2-adrenoceptor occupancy was assessed in a superfusion system by means of the dose-dependent inhibition of [3H]5-HT release by clonidine. This is blocked by yohimbine (1 microM), which, when administered alone, enhances release, suggesting that endogenous catecholamines released from other synaptosomes act on these alpha 2-heteroreceptors. The effect of addition of citalopram (1 microM) to superfusates suggests that some reuptake of [3H]5-HT occurs during superfusion. Of the tritium released into superfusates during "background" and K+-stimulated release, 17 and 90%, respectively is [3H]5-HT. The attenuation of K+-stimulated release by clonidine is apparently diminished by the chronic clorgyline regimen but not by desipramine. However, clorgyline elevates catecholamine levels, and this might increase endogenous noradrenaline (NA) efflux, which by competition with clonidine could appear to alter alpha 2-adrenoceptor sensitivity. This possibility was investigated by depleting NA with the neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP4). These studies showed that the apparent effect of chronic clorgyline on alpha 2-adrenoceptor sensitivity to clonidine was due to competition with increased levels of endogenous NA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Assessment of Noradrenaline and 5-Hydroxytryptamine Uptake in Intact Rat Brain

A method for the measurement of 3H-monoamine [noradrenaline (NA) and 5-hydroxytryptamine (5-HT)] uptake into intact brain following intracerebroventricular injection is described. Most of the accumulated monoamine was associated with nerve terminals (synaptosomes). Radioactivity in the synaptosomal fraction was retained by 0.45-micron filters and was osmotically sensitive, features indicative of accumulation within particles rather than binding to membranes. [3H]5-HT associated with synaptosomes was reduced in animals pretreated with the neurotoxin 5,7-dihydroxytryptamine, in parallel with a reduction in endogenous 5-HT levels. Oral administration of tricyclic antidepressants including clomipramine, desipramine, and imipramine inhibited the synaptosomal accumulation of 3H-monoamines. Overall, the features of the accumulation of 3H-monoamines, after intracerebroventricular administration, displayed many of the characteristics expected of a physiological monoamine reuptake system.     

3H]adrenaline release from hypothalamic synaptosomes and its modulation by clonidine: effects of chronic antidepressant drug regimens

[3H]Adrenaline ([3H]ADR, 40 nM) was accumulated by rat hypothalamic synaptosomes (P2) more rapidly and in significantly greater amounts than by similar preparations from cerebral cortex. There was no significant difference between these two tissues in the rate or amount of [3H]noradrenaline ([3H]NA, 40 nM) accumulation. Talusupram (10 microM), maximally inhibited the uptake of [3H]ADR into hypothalamic synaptosomes by 60%. Nomifensine further inhibited uptake by 14%. From these observations it was concluded that some [3H]ADR was accumulated into non adrenergic neuronal terminals. The effects of desipramine (DMI, 10 mg/kg/day and clorgyline (1 mg/kg/day) administration for 28 days on K+-evoked release of [3H]ADR was investigated using superfused hypothalamic synaptosomes. After both chronic antidepressant drug regimens, total [3H]ADR release (spontaneous + evoked) was significantly reduced. Evoked release of [3H]ADR (by KCl, 16 mM) was significantly reduced after the DMI but not the clorgyline regimens. Presynaptic alpha 2-adrenoceptor function in the hypothalamus was assessed during superfusion by measuring the reduction in K+-evoked release of [3H]ADR caused by clonidine (1 microM). The attenuating effects of clonidine on [3H]ADR release (42% in untreated controls and 36% after chronic clorgyline) was diminished (to 4%) after chronic DMI administration. Alpha 2 adrenoceptor numbers in the rat hypothalamus were not significantly changed after clorgyline or DMI administration, suggesting that the functional subsensitivity seen in synaptosomes after DMI, may not be related to alpha 2 adrenoceptor down regulation.     

ON THE UPTAKE OF INOSITOL BY RAT BRAIN SYNAPTOSOMES

The uptake of inositol by rat brain synaptosomes occurs via an unsaturable process that even at substrate concentrations as low as 1 μM is unable to achieve a concentration gradient indicative of active transport. Dinitrophenol, ouabain and cytochalasin B did not affect uptake of the cyclitol. The data indicate that inositol uptake by rat synaptosomes occurs by diffusion or by a system with an affinity so low it is difficult to discern. The low capacity, saturable inositol uptake system observed in rabbit brain slices may reflect a species difference or uptake by elements of the slice other than neuronal membranes.     

Effect of leu-enkephalin and the delta-sleep-inducing peptide, M (DSIP), on endogenous noradrenaline release by brain synaptosomes in the rat

Fluorometry was employed to measure the noradrenaline (NA) content in rat brain synaptosomes depending on the duration of incubation, depolarization effects (40 mM KCl or 1.5 mM ouabain), composition of the synaptosomal fraction and concentration of the peptides. The 10-minute incubation in a potassium medium of a suspension of light synaptosomes was used as an optimal test-system for studying the peptide action. Leu-enkephalin inhibited the depolarization-induced NA release. The effect was abolished by naloxone. The delta-sleep-inducing peptide (DSIP) did not influence the neurotransmitter release at concentrations of 10(-8)-10(-5) M. A mixture of amino acids imitating the amino acid composition of the DSIP influenced spontaneous release of NA. This effect is discussed in connection with the physiological action of the peptide on its intraventricular injection.     

Chronic nicotine differentially affects the function of nicotinic receptor subtypes regulating neurotransmitter release

It is known that nicotine can activate several subtypes of release-regulating presynaptic nicotinic receptors (nAChRs) including those situated on central noradrenergic, dopaminergic, cholinergic and glutamatergic axon terminals. The objective of this study was to investigate the effects of chronic administration of (-)nicotine on the function of the above autoreceptors and heteroreceptors using rat superfused synaptosomes. In hippocampal synaptosomes prelabelled with [3H]noradrenaline (NA) the nicotine-evoked overflow of [3H]NA was higher in rats treated with nicotine for 10 days (via osmotic mini-pumps) than in vehicle-treated rats. In striatal synaptosomes, prelabelled with [3H]dopamine (DA), chronic nicotine did not modify the releasing effect of nicotine. No significant change was observed in experiments with synaptosomes from nucleus accumbens prelabelled with [3H]DA. Exposure of hippocampal synaptosomes prelabelled with [3H]choline to nicotine elicited release of [3H]acetylcholine; this effect was almost abolished in synaptosomes from animals administered nicotine for 10 days, suggesting down-regulation of nicotinic autoreceptors. In hippocampal synaptosomes prelabelled with [3H]D-aspartate, the releasing effect of epibatidine following chronic nicotine treatment did not differ from that in controls. The K+-evoked exocytotic release of the neurotransmitters tested was not modified by long-term nicotine administration. The results show that chronic nicotine differentially affects the function of release-regulating nAChR subtypes.     

Similarities Between A.P.U.D. Cells and Monoaminergic Neurons: Evidence for a Dopamine Uptake Mechanism in C Cells of Chicken Ultimobranchial Gland

Abstract: The uptake of [³H]dopamine was studied in the C cells of chicken ultimobranchial gland (UBG). DA uptake appears to be an active and saturable mechanism (K_m= 8.6 μM). The C cells' affinity for DA is intermediate between dopaminergic synaptosomes and such extraneural tissues as platelets and heart. UBG incubation with DA leads to the synthesis of a noticeable quantity of DOPAC, indirectly showing the presence of MAO activity. Thus UBG C cells, besides having both an embryological derivation from the neural crest and some morphological aspects similar to synaptosomes, also possess an active uptake mechanism for a central neurotransmitter.     

Membrane-bound histamine N-methyltransferase in mouse brain: possible role in the synaptic inactivation of neuronal histamine

In the CNS, histamine is a neurotransmitter that is inactivated by histamine N-methyltransferase (HNMT), a soluble enzyme localized to the cytosol of neurons and endothelial cells. However, it has not been established how extracellular histamine, a charged molecule at physiological pH, reaches intracellular HNMT. Present studies investigated two potential routes of histamine inactivation in mouse brain nerve terminal fractions (synaptosomes): (i) histamine uptake and (ii) histamine metabolism by HNMT. Intact synaptosomes demonstrated a weak temperature-dependent histamine uptake (0.098 pmol/min-mg protein), but contained a much greater capacity to metabolize histamine by HNMT (1.4 pmol/min-mg protein). Determination of the distribution of HNMT within synaptosomes revealed that synaptosomal membranes (devoid of soluble HNMT) contribute HNMT activity equivalent to intact synaptosomes (14.3 +/- 2.2 and 18.2 +/- 4.3 pmol/min-tube, respectively) and suggested that histamine-methylating activity is associated with the membrane fraction. Additional experimental findings that support this hypothesis include: (i) the histamine metabolite tele-methylhistamine (tMH) was found exclusively in the supernatant fraction following an HNMT assay with intact synaptosomes; (ii) the membrane-bound HNMT activity was shown to increase 6.5-fold upon the solubilization of the membranes with 0.1% Triton X-100; and (iii) HNMT activity from the S2 fraction, ruptured synaptosomes, and synaptosomal membranes displayed different stability profiles when stored over 23 days at - 20 degrees C. Taken together, these studies demonstrate functional evidence for the existence of membrane-bound HNMT. Although molecular studies have not yet identified the nature of this activity, the present work suggests that levels of biologically active histamine may be controlled by an extracellular process.     

Effects of long-term treatment of rats with antidepressants on adrenergic-receptor sensitivity in cerebral cortex: Structure activity study

The effects of 20 tricyclic and 12 chemically unrelated ‘atypical’ antidepressant drugs on the noradrenaline (NA) receptor coupled adenylate cyclase system were investigated in slices of the rat cerebral cortex. While no changes occurred after a single dose, 14 tricyclic compounds down-regulated the receptor system when administered for 9 days. Six tricyclic antidepressants (trimipramine, butriptyline, noxiptyline, doxepine, dosulepine, propizepine) failed to desensitize the NA sensitive adenylate cyclase although some were potent inhibitors of the neuronal uptake of NA. Using the two optically active enantiomers of oxaprotiline inhibition of NA uptake was observed with the (+)-enantiomer while the (?)-enantiomer had only weak inhibitory potency. However, in contrast to published data, both enantiomers and the racemate administered at 30 mg/kg for 9 days did down-regulate the NA receptor coupled adenylate cyclase. Therefore, the experiments were repeated with Sprague-Dawley rats from a different supplier. Now, data published earlier were reproducible, only the racemate and the (+)-enantiomer of oxaprotiline being significantly active on the desensitization of the NA sensitive adenylate cyclase. Using F-344, Long Evans and Wistar rats significant differences were found in the response of the adrenoceptor coupled adenylate cyclase to a 9 day treatment with 30 mg/kg imipramine. Although some of the atypical antidepressants are potent inhibitors of the biogenic amine uptake systems none of these compounds lead to statistically significant changes of the NA stimulated cAMP formation after a 9-day treatment period. Only with the NA uptake inhibitor tandamine and with the serotonin uptake inhibitors zimelidine and fluoxetine a trend toward adrenergic down-regulation was recognized. Using enantiomers of mianserin only the (?)-isomer which is a poor NA uptake inhibitor, was slightly active. It thus appears that the therapeutic action of antidepressant drugs cannot generally be related to postsynaptic adaptive changes in the sensitivity of the noradrenergic adenylate cyclase receptor system. Variabilities in pharmacokinetics and in NA sensitivity of the cAMP generating system in various rat strains and possibly in different animal species may be important factors determining whether β-receptor down-regulation will occur during chronic treatment with antidepressant drugs.     

An inhibitor of dopamine uptake,LR5182, CIS-3-(3,4-dichlorophenyl)-2-N,N-dimethylaminomethyl-bicyclo-[2,2,2]-octane,hydrochloride

LR5182 inhibited the uptake of dopamine in rat striatal synaptosomes and the uptake of norepinephrine in cortical synaptosomes with inhibitor constants, Ki values, of 3nM and 58nM, respectively. It was only a week inhibitor of serotonin uptake in cortical synaptosomes with a Ki value of 1.7μM. The uptake of dopamine and norepinephrine were significantly lowered within an hour after an intraperitoneal injection of LR5182. Among known inhibitors of dopamine uptake in synaptosomes of rat brain, LR5182 is most effective and selective. The rigid structure of LR5182 (Figure 1) suggested a gauche conformation of dopamine to be favored by the striatal uptake of dopamine.     

The Effects of Scorpion Venom Toxin on the Release of Amino Acid Neurotransmitters from Cerebral Cortex In Vivo and In Vitro

Tityustoxin, the active component of the venom of the Brazilian yellow scorpion Tityus serrulatus, caused specific release of the neurotransmitter amino acids glutamate, aspartate and GABA in vivo from the superfused sensori-motor cortex of conscious unanesthetised rats and in vitro from rat cortical synaptosomes. The effects on synaptosomes appear to be due to a depolarising action. Synaptosomal potassium levels were depleted by the toxin. The action was also blocked both in vivo and in vitro by tetrodotoxin and was Ca2+-dependent. The uptake of [U-14C]GABA was inhibited by tityustoxin but this action was prevented by tetrodotoxin (1 microM). Since the release of [U-14C]GABA from synaptosomes due to the tityustoxin was also prevented by tetrodotoxin under identical circumstances, it is concluded that the tityustoxin has a primary action on release of neurotransmitters rather than on uptake.