Effect of Hypophysectomy and Adrenalectomy on 5-Hydroxytryptamine Uptake by Rat Hypothalamic Synaptosomes and Blood Platelets

Abstract: The effect of adrenalectomy and hypophysectomy on 5-HT uptake by rat platelets and hypothalamic synaptosomes taken from the same animal was studied. The experiments were performed 1–2 weeks after the operations or the shamoperations. After adrenalectomy a significant decrease of 25% in 5-HT uptake by platelets was noted. In synaptosomes there was a similar trend, but the decrease was not significant. After hypophysectomy a significant 25% decrease in 5-HT uptake was observed both in platelets and in synaptosomes. These results suggest that corticosteroid hormones might mediate adaptive changes of 5-HT uptake both in synaptosomes and in blood platelets. Blood platelets have been used as a peripheral model of serotoninergic nerve terminal in studies concerning physiology of monoamines systems, but the present results suggest that also in some pathological conditions changes in platelet biochemistry could reflect those in the CNS neurones.     

Unusual binding of [3H] MDL 72222 in guinea pig hippocampus

[³H] MDL 72222 labeled a non-homogeneous population of sites in guinea pig hippocampal membranes (K_d1 = 1 nM; K_d2 = 60 nM). The binding was not sodium dependent. Competition studies with a variety of characterizing agents showed displacement of [³H] MDL 72222 binding by 5-HT uptake inhibitors. [³H] MDL 72222 binding was not effectively displaced by established 5-HT₃ antagonists. MDL 72222, fluoxetine, fluvoxamine and citalopram competitively inhibited the uptake of [³H] 5-HT into guinea pig hippocampal synaptosomes with K_i values of 1.97, 0.02, 0.023, 0.049 μM, respectively. The results demonstrate that [³H] MDL 72222 labels a non-homogeneous population of sites in guinea pig brain, as well as inhibiting 5-HT uptake into synaptosomal preparations.     

Effect of a specific 5-HT uptake inhibitor,citalopram (Lu 10-171), on 3H-5-HT uptake in rat brain synaptosomes in vitro

The uptake of ³H-5-HT in synaptosomes from rat brains was investigated. Addition of DA or NA had only a slight or no effect on the uptake. When the uptake into NA and DA neurons was inhibited by the addition of high concentrations of NA and DA, the uptake of ³H-5-HT was unchanged. This was also found after destruction of NA and DA neurons by 6-hydroxydopamine treatment. Furthermore, the uptake of ³H-5-HT was almost equal in different brain parts containing NA and DA in very different amounts. These observations show that the uptake measured with ³H-5-HT is specific for 5-HT neurons.The present study revealed that citalopram and chlorimipramine inhibited uptake competitively, and in this respect the two drugs were equipotent. Compared with a series of tricyclic thymoleptics, the two drugs were the most potent inhibitors of 5-HT uptake, about 20 to 35 times more active than imipramine and amitriptyline. The metabolites of citalopram were also rather potent. The results obtained in the present study correlate closely with those obtained using inhibition of ¹⁴C-5-HT uptake in blood platelets, or using the inhibition of H 75/12-induced 5-HT depletion in rat brain.When rats were treated orally with citalopram or chlorimipramine, the inhibition of ³H-5-HT uptake in synaptosomes derived from these rats was two times greater after citalopram than after chlorimipramine.     

Comparative effects of trazodone and tricyclic antidepressants on uptake of selected neurotransmitters by isolated rat brain synaptosomes

The effect of trazodone, a new antidepressant agent, on uptake of serotonin (5-HT), norepinephrine (NE), and dopamine (DA) by crude synaptosome preparations from rat hypothalamus was compared with imipramine, desipramine, and clomipramine. Trazodone was determined to be a very selective inhibitor of the 5-HT uptake mechanism with IC₅₀ values of 5.67×10^-7, 3,54×10^-5, and 5.25×10^-5 M, for 5-HT, NE, and DA uptake, respectively. Clomipramine, the only other selective inhibitor of 5-HT uptake studied, had IC₅₀ values of 7.59×10^-9, 1.12×10^-7, and 2.51×10^-7 M, for 5-HT, NE, and DA, respectively. Although less potent, trazodone was 4±0.6 times more selective than clomipramine in its ability to inhibit synaptosomal uptake of 5-HT with respect to NE. This selectivity for the 5-HT uptake mechanism is consistent with the clinical antidepressant efficacy of trazodone.     

Unaltered 5-HT- and desipramine-sensitive [3H]imipramine binding and [3H]5-HT uptake in rat brain after chronic imipramine and norzimeldine treatment

Several reports have shown heterogeneity of [³H]imipramine binding to brain membranes. Recently, a high affinity and 5-HT sensitive [³H]imipramine binding site of protein nature, that was suggested to be identical to the substrate recognition site for 5-HT uptake, was demonstrated. Since most studies on the regulation of the [³H]imipramine binding sites by antidepressants have used desipramine displaceable binding, which is heterogenous in nature and contains binding not related to 5-HT uptake sites, the present report studies the possible effects of chronic (3 weeks) administration of imipramine or norzimeldine (10 mg/kg intraperitoneally twice daily) on 5-HT sensitive [³H]imipramine binding sites. For comparison, desipramine sensitive binding was also studied, as well as the physiological correlate 5-HT uptake. There were no changes in either [³H]imipramine binding or 5-HT uptake after the antidepressant treatment.Supported by the Swedish Medical Research Council Offprint requests to: J. Marcusson at Dept. of Geriatric Medicine     

Inhibition of rat platelet 5-hydroxytryptamine uptake by chlordimeform

Chlordimeform, a formamidine insecticide and acaricide, and two of its toxic N-demethyl metabolites inhibited rat blood platelet 5-hydroxytryptamine (5-HT) uptake. A direct relationship existed between formamidine uptake inhibitory potency and N-demethylation. Didemethylchlordimeform, the most potent formamidine inhibitor of 5-HT uptake examined, had a pI₅₀ of 4.2. However, it was appreciably less active than the classical inhibitor imipramine, which yielded 86.4% inhibition of 5-HT uptake at a concentration of 1 · 10^?5 M.     

Evidence for various tryptamines and related compounds acting as substrates of the platelet 5-hydroxytryptamine transporter

Summary The aim of the present study was to answer the question whether amines other than 5-hydroxytryptamine (5-HT) and tryptamine act as substrates of the platelet 5-HT transporter. To this end, a large number of tryptamines, 5-HT receptor agonists and phenethylamines (which had IC₅₀ values for ³H-5-HT uptake inhibition of 145–24500 nmol l^–1) was examined in rabbit platelets in order to determine their ability to induce an outward transport of ³H-5-HT Platelets (the MAO of which was blocked) from reserpine-pretreated animals were loaded with ³H-5-HT and then exposed for 5 min to various concentrations (ranging from 0.25 to 40 times the IC₅₀) of each compound. The concentration-effect curves for the drug-induced increase in ³H-5-HT efflux served to determine values of E_max (maximum increase in efflux expressed in % of the ³H-5-HT content of cells) and EC₅₀ (drug concentration producing E_max/2).For the 24 compounds studied here (which included the 5-HT uptake inhibitors imipramine, citalopram, fluoxetine and cocaine) a linear correlation between EC₅₀ and IC₅₀ (r = 0.975) and a mean ratio of EC₅₀/IC₅₀ of 2.4 was found. Most of the compounds [e.g., (±)8-hy-ydroxy-2-(N,N-dipropylamino)tetralin, S(+)-methyl-5-HT, 5-carboxamidotryptamine and 5-methoxytryptamine] gave rise to E_max values (15.8–32.5%) that exceeded that brought about by imipramine (6.6%), indicating that they act as substrates of the 5-HT transporter; the ³H-5-HT outward transport observed in response to these substances was abolished in the presence of imipramine. Others (e.g., 2-methyl-5-HT and 5-methylurapidil) produced Emax values (3.4–14.3%) not significantly different from that of imipramine and, therefore, can be classified either as poor substrates or as inhibitors of the 5-HT transporter.Hence, many tryptamines and 5-HT receptor agonists are substrates of the platelet 5-HT transporter. The property of being substrates gives them the latent capacity to bring about release of endogenous 5-HT and, as a result, to cause indirect 5-HT receptor-mediated effects.Abbreviations MAO monoamine oxidase - 5-HT 5-hydroxytryptamine - 2-M-5-HT 2-methyl-5-HT - N-M-5-HT N-methyl-5-HT - N,N-DM-5-HT N,N-dimethyl-5-HT - S(+)-M-5-HT S(+)-methyl-5-HT - 5-CT 5-carboxamidotryptamine - 5-M-tryptamine 5-methyltryptamine - 5-MO-tryptamine 5-methoxytryptamine - 7-M-tryptamine 7-methyltryptamine - N-M-tryptamine N-methyltryptamine - N,N-DM-tryptamine N,N-dimethyltryptamine - N,N-DM-5-MO-tryptamine N,N-dimethyl-5-methoxytryptamine - (±)8-OH-DPAT (±)8-hydroxy-2-2-(N,N-dipropylamino)tetralin - 5-M-urapidil 5-methyl-urapidil Send offprint requests to R. Wölfel at the above address     

Tricyclic Antidepressant Agents. I. Comparison of the Inhibition of the Uptake of 3H-Noradrenaline and 14C-5-Hydroxytryptamine in Slices and Crude Synaptosome Preparations of the Midbrain-Hypothalamus Region of the Rat Brain

Abstract: The simultaneous uptake of ³H-I-noradrenaline (NA) and ¹⁴C-5-hydroxytryptamine (5-HT) in slices from the midbrain-hypothalamus region of the rat brain was compared with the corresponding uptake in crude synaptosome preparations of the same brain region. In both preparations the uptake of the two amines was selective at the concentration used (1 x 10^-7 M or lower). The K_M values for the amines (NA: 2 x 10^-7 M in synaptosomes and 5 x 10^-7 M in slices; 5-HT: 8 x 10^-8 M in synaptosomes and 6 x 10^-7 M in slices) and the inhibitory concentrations (IC₅₀) of the antidepressant agents were lower in the synaptosome experiments than in the slices experiments. Moreover the order of the inhibitory activities differed between the two preparations. In the slices experiments the NA uptake was inhibited most markedly by desipramine followed by imipramine > chlorimipramine = nortriptyline ≥ amitriptyline ≥ chlordesipramine whereas in the synaptosome experiments the order was desipramine > nortriptyline ≥ chlordesipramine ≥ imipramine > amitriptyline ≥ chlorimipramine. For the 5-HT uptake in slices the order of activity was: chlorimipramine > imipramine ≥ amitriptyline ≥ chlordesipramine = desipramine ≥ nortriptyline whereas in the synaptosome preparations the order was: chlorimipramine > imipramine ≥ amitriptyline ≥ chlordesipramine > nortriptyline = desipramine. The role of protein binding and diffusion barriers in the causation of the difference in the results obtained with the two preparations is discussed.     

Binding of some antidepressants to the 5-hydroxytryptamine transporter in brain and platelets

Antidepressant agents with properties to inhibit 5-hydroxytryptamine (5-HT, serotonin) uptake in brain tissue and platelets bind with high affinities to neuronal and platelet membranes. [³H]Imipramine, [³H]paroxetine and [³H]citalopram label specific binding sites related to the 5-HT transporter. [³H]Paroxetine and [³H]citalopram appear to be better ligands than [³H]imipramine. The former label a homogenous population of binding sites, whereas the displaceable binding of [³H]imipramine is heterogenous. Recent observations in several laboratories, which have taken the heterogeneity of [³H]imipramine binding into account, indicate that the binding of antidepressants to the 5-HT transporter probably occurs to the same site that binds 5-HT for transport and not to a separate site as previously suggested. Additional bonds to subsites in close vicinity to the 5-HT recognition site may contribute to the binding. No convincing evidence has been presented of the existence of an endogenous ligand other than 5-HT itself that binds to the [³H]imipramine binding site. Recent studies also suggest that repeated treatment of rats with antidepressant agents does not produce any alterations of the binding of [³H]imipramine or [³H]paroxetine to membranes of cerebral cortex. It is also doubtful whether the density of the 5-HT uptake site in platelets measured with these ligands is decreased in affective disorders as first reported.     

Biochemical characteristics of a potential antidepressant, 2-(7-indenyloxymethyl)morpholine hydrochloride (YM-08054-1)

The effects of potential antidepressant, 2-(7-indenyloxymethyl)morpholine hydrochloride (YM-08054-1), on uptake, release and oxidative deamination of catecholamines and serotonin [5-hydroxytryptamine(5-HT)] were studied in vitro with rat brain, and compared with the effects of several tricyclic antidepressants and viloxazine a structural analogue of YM-08054-1. YM-08054-1 at concentrations of less than 1 μM. considerably inhibited 5-HT uptake by synaptosomes from rat whole brain as well as noradrenaline (NA) uptake by synaptosomes from rat hippocampus, similar to amitriptyline. Viloxazine and iprindole were weak inhibitors in both uptake reactions. In the release of either [¹⁴C] -5-HT from [¹⁴C]-5-HT-prelabeled hypothalamic slices or [¹⁴C]NA from [¹⁴C] NA-prelabeled hippocampal slices, YM-08054-1 and amitriptyline were much less potent than such typical potent releasers as methamphetamine and tyramine. YM-08054-1 was also found to be a weak inhibitor of both type A and type B monoamine oxidases, whereas the tricyclic antidepressants preferentially inhibited the type B enzyme. These biochemical results indicate that YM-08054-1 may be an amitriptyline-like antidepressant, but distinct from viloxazine.     

Examination of the relationship between the uptake site for 5-hydroxytryptamine and the high affinity binding site for [3H]imipramine. II. The role of sodium ions

Exogenous sodium ions stimulated both the high affinity binding of [3H]5-imipramine to membranes from the cortex of the rat and the high affinity accumulation of [3H]5-hydroxytryptamine (5-HT) into synaptosomes from the cortex of the rat with similar potencies. Imipramine and zimelidine inhibited synaptosomal uptake of [3H]5-HT potently in standard Tris-Krebs medium, but in a low-sodium medium their inhibitory potencies were significantly attenuated. The inhibitory potencies of panuramine and exogenous 5-HT on the uptake of [3H]5-HT were not significantly affected whether the uptake was measured in a normal or low-sodium Tris-Krebs. Imipramine and zimelidine were potent blockers of high affinity binding of [3H]imipramine whereas panuramine and 5-HT only inhibited the binding of [3H]imipramine at concentrations in excess of those required to inhibit the uptake of [3H]5-HT. It is suggested that imipramine inhibits the uptake of 5-HT by a sodium-dependent action probably at the high affinity binding site for [3H]imipramine, whereas panuramine and 5-HT inhibit the uptake of 5-HT by a sodium-independent mechanism at a site other than the binding site for [3H]imipramine.     

Inhibition of 5-hydroxytryptamine accumulation and deamination by substituted phenylalkylamines in hypothalamic synaptosomes from normal and reserpine-pretreated rats

Summary 1. In the present study the abilities of different compounds to inhibit MAO inside and outside the serotonergic neurons, to inhibit the accumulation of 5-HT and to release 5-HT were separated by using different in vitro techniques. With these methods a number of substituted phenylalkylamines, which are reversible inhibitors of monoamine oxidase (MAO) type A, were characterized. 2. The compounds were examined regarding their ability to inhibit the accumulation of 5-HT and to inhibit MAO in the same synaptosomal preparation of hypothalamus from normal and reserpine-pretreated rats. The difference in the uptake of ¹⁴C-5-HT (0.1 mol/l) in the absence and presence of citalopram (0.25 mol/l) was taken as a measure of the accumulation into the serotonergic synaptosomes. The deamination of ¹⁴C-5-HT (0.1 mol/l) in the presence of citalopram (0.25 mol/l) was considered as that brought about outside the serotonergic synaptosomes, whereas the difference between the deamination in the absence and presence of citalopram was taken as the MAO activity inside the serotonergic synaptosomes. 3. Most of the phenylalkylamines were slightly more potent as MAO inhibitors outside serotonergic synaptosomes than as inhibitors of 5-HT accumulation in normal rats. The most potent MAO inhibitors, both in absolute terms and in comparison with uptake inhibitory potency, were the 2,6-dichloro-(FLA 365) and the phenylpropylene-(FLA 417) derivatives. 4. A difference in potency on the accumulation in synaptosomes from normal and reserpine-pretreated rats was found for many of the phenylalkylamines with the exception of FLA 365, FLA 417 and the 2,5-dimethyl derivative RAN 113. The compounds shared this difference with the 5-HT releasers p-chloroamphetamine (pCA) and H75/12, but not with the uptake inhibitors citalopram, cocaine, alaproclate, norzimeldine and fluoxetine. 5. For most of the phenylalkylamines, the MAO inhibition obtained within serotonergic synaptosomes was not higher than that obtained outside these. This was in contrast to previously shown in vivo results were a preference for inhibiting MAO inside the serotonergic neurons, due to a transport into the neurons by the amine carrier, was found. For the uptake inhibitors, the inhibition of 5-HT deamination intrasynaptosomally was a more sensitive measure of inhibition of accumulation than to determine this accumulation directly. 6. It is concluded that a classification of uptake inhibitors, releasing compounds, and MAO inhibitors that are and that are not transported by the 5-HT carrier may be performed by measuring the inhibition of the uptake and the deamination in synaptosomes from normal and reserpine-pretreated rats.Parts of this work were presented at the 14th CINP Congress, Florence, Italy, June 19–23, 1984 and the VIIIth International Symposium on Medicinal Chemistry, Uppsala, Sweden, August 27–31, 1984 Send offprint requests to A.-L. Ask     

Effect of acute and prolonged tianeptine administration on the 5-HT transporter: electrophysiological,biochemical and radioligand binding studies in the rat brain

In the present study, in vivo extracellular unitary recordings, in vitro [³H]5-HT uptake and [³H]cyanoimipramine binding assays were used to assess the effect of acute and prolonged administration of the putative antidepressant tianeptine, on the 5-hydroxytryptamine (5-HT) transporter. Microiontophoretic application of tianeptine onto dorsal hippocampus CA₃ pyramidal neurons, as well as its intravenous administration (2 mg/kg), increased their firing frequency. Following intracerebroventricular administration of 5,7-dihydroxytryptamine, the activation induced by the microiontophoretic application of tianeptine remained unchanged, thus suggesting that the 5-HT carrier is not involved in this effect. Furthermore, in spite of its activating effect on CA₃ pyramidal neuron firing frequency, the intravenous administration of tianeptine did not alter the time of recovery of these neurons from microiontophoretic applications of 5-HT, an index of 5-HT uptake activity. In keeping with this observation, the acute administration of tianeptine did not change the effectiveness of the 5-HT reuptake blocker paroxetine (1 mg/kg, i.v.) in prolonging the suppressant effect of microiontophoretically-applied 5-HT. However, in rats that had received tianeptine for 14 days (20 mg/kg/day, s.c.), the recovery time from the suppressant effect of microiontophoretic applications of 5-HT was reduced by 40% and the effectiveness of paroxetine (1 mg/kg, i.v.) was decreased. These effects were no longer observed following a 48 h washout period. In a second series of experiments, the ability of tianeptine to interfere with the uptake blocking capacity of paroxetine was assessed in vitro, using hippocampal slices obtained from rats that had been treated with tianeptine for 14 days (20 mg/kg/day, s.c.; by minipump). The effectiveness of paroxetine to block [³H]5-HT uptake was unchanged in slices obtained from rats still bearing the osmotic minipump at the time of the sacrifice, as well as from those which had undergone a 48 h washout period. To assess whether prolonged administration of tianeptine would induce adaptive changes on 5-HT uptake sites, [³H]cyanoimipramine-binding parameters were measured following a 48 h washout period. Affinity values remained unchanged while density values were significantly increased in cortex (+22%) but not in hippocampus (+12%). It is concluded that, i) the activation of CA₃ pyramidal neurons observed following acute tianeptine administration cannot be attributed to its 5-HT uptake enhancing properties and ii) the prolonged administration of tianeptine induces adaptive changes on cortical but not on hippocampal 5-HT transporters.Deceased 10 May 1994     

The nature of [3H]imipramine binding to synaptosomes

In contrast to serotonin “uptake”, the saturable binding of imipramine to rat brain synaptosomes does not involve an active transport process, since it is independent of time, temperature and Na⁺ K⁺-ATPase. It is moreover unaffected by a high concentration of serotonin. confirming that the “uptake” site for this amine is not implicated. Analysis of the saturation curve for binding of imipramine to intact synaptosomes gives a binding constant of 3.5 ± 1.0 × 10^?5 M. Identical binding characteristics are found with a synaptosomal membrane “ghost” fraction. When the membrane fraction is solubilized in Triton X-100, saturable binding is still observed but the affinity for imipramine decreases while the number of binding sites increases. Diverse psychoactive compounds effectively compete with imipramine for binding to synaptosomal membranes and the order of their activity is correlated with their lipophilicity. It is suggested that these compounds interact with a lipophilic pocket of low affinity and low specificity. If a more specific interaction exists and is associated with inhibition of transmitter amine “uptake” by tricyclic antidepressants, it could be masked by this process.     

Tricyclic Antidepressant Agents. II. Effect of Oral Administration on the Uptake of 3H-Noradrenaline and 14C-5-Hydroxytryptamine in Slices of the Midbrain-Hypothalamus Region of the Rat

Abstract: The inhibition of the simultaneous uptake of ³H-l-noradrenaline (NA) and ¹⁴C-5-hydroxytryptamine (5-HT) in slices of the midbrain-hypothalamus region of the rat brain after oral administration of desipramine, imipramine, nortriptyline, amitriptyline, chlordesipramine and chlorimipramine was determined. All compounds were more active in inhibiting the NA uptake than the 5-HT uptake. This difference was very marked for desipramine, imipramine, nortriptyline and chlordesipramine. Chlorimipramine was almost as active on the 5-HT uptake (ED₅₀ = 35 mg/kg orally) as on the NA uptake (ED₆₀ = 20 mg/kg orally) and amitriptyline had low activity on both uptake mechanisms (ED₅₀ > 50 mg/kg orally). Desipramine and imipramine were the most active compounds on the NA uptake (ED₅₀ = 8 mg/kg orally for both compounds) and the duration of the action was very long. The ED₅₀ values for nortriptyline and chlordesipramine in inhibiting the NA uptake were about 20 mg/kg orally for both compounds. The inhibition of the 5-HT uptake was less than 50% at 50 mg/kg orally for all compounds except for imipramine (ED₅₀ = 50 mg/kg orally) and for chlorimipramine. The role of the biotransformation for the inhibitory activities of imipramine, chlorimipramine and amitriptyline was investigated in animals pre-treated with SKF 525 A. The inhibitory potency of imipramine was increased by the same factor for both uptake mechanisms probably due to the large increase in the concentration of imipramine in the rat brain, which was demonstrated after the administration of ¹⁴C-imipramine. The inhibitory activity of chlorimipramine was somewhat more increased for the 5-HT uptake than for the NA uptake. The low activity of amitriptyline seems to be mainly due to poor resorption, since pretreatment of the animals with SKF 525 A only slightly increased the potency whereas intraperitoneal injection of amitriptyline had a rather marked effect on the NA uptake (ED₅₀ = 11 mg/kg intraperitoneally).     

Effect of lofepramine and other antidepressants on the uptake of 5-hydroxytryptamine and noradrenaline into rat brain monoaminergic neurons

Lofepramine, (N-methyl-N-[4-chlorobenzoylmethyl]-3-[10, 11-dihydro-5H-dibenz(b,f)-azepin-5-yl]-propylamine hydrochloride), is a new antidepressant with low toxicity and no peripheral anticholinergic activity. Its effect on 5-hydroxytryptamine (5-HT) and noradrenaline uptake into rat brain monoaminergic neurons was studied and compared with that of other antidepressants, particularly with that of imipramine and desipramine. Lofepramine inhibited both 5-HT and noradrenaline uptake into synaptosomal fractions in vitro but was 4 times more potent in inhibiting noradrenaline than 5-HT uptake, indicating the effect resembles that of desipramine. Noradrenaline uptake was also preferentially inhibited in synaptosomes from brain of rats treated previously with lofepramine or desipramine (i.p.). Pretreatment with SKF 525A (i.p.) did not diminish the effect of lofepramine but rather potentiated it. Therefore it is suggested that the formation of desipramine is not necessary for lofepramine to exhibit the effect on amine uptake in vivo. Both lofepramine and desipramine inhibited intraventricular noradrenaline uptake into synaptosomes without any effect on 5-HT uptake. These results suggest that lofepramine is qualitatively similar to desipramine with respect to preferential inhibition of noradrenaline uptake into central noradrenergic neurons.     

Enhancement of 5-Hydroxytryptamine Uptake in Rabbit Hypothalamic Synaptosomes but not in Blood Platelets by Zinc and Lead ex Vivo

Abstract: Lead (0.2%), zinc (0.5%) or their combination was given to rabbits as acetate salts in drinking water for 2 or 4 weeks. Cerebral Pb increased within exposure time but the increase of Zn was marginal when given alone. After combination of Pb+Zn, blood Pb increased but Zn decreased as compared to the administration of the single metal alone. Concurrently brain Pb increased but remained at a lower level as compared with sole Pb administration while brain Zn did not differ from controls. The uptake of 5-hydroxytryptamine (5-HT) into hypothalamic synaptosomes increased 70–85% with Pb, Zn and Pb+Zn after 2 weeks and 93% with Zn and 76% with Pb+Zn after 4 weeks. Dopamine uptake into striatal synaptosomes did not change. Zn decreased endogenous noradrenaline concentrations by 21% and Pb+Zn that of 5-HT by 36% in the striatum. Pb+Zn decreased cortical dopamine concentrations by 26%. Concurrently, 5-HT uptake in blood platelets, platelet numbers and their content of endogenous 5-HT did not change significantly. These results indicate that high Zn and Pb have parallel effects on 5-HT uptake in hypothalamic synaptosomes and their combination may be neurochemically more toxic than the individual metals. The increase of 5-HT uptake in hypothalamic synaptosomes but not in blood platelets requires further methodological work to explain the differences between synaptosomes and platelets.     

Effect of acute and chronic tramadol on [3H]-5-HT uptake in rat cortical synaptosomes

1. Tramadol hydrochloride is a centrally acting opioid analgesic, the efficacy and potency of which is only five to ten times lower than that of morphine. Opioid, as well as non-opioid mechanisms, may participate in the analgesic activity of tramadol.
2. [³H]-5-hydroxytryptamine (5-HT) uptake in rat isolated cortical synaptosomes was studied in the presence of tramadol, desipramine, fluoxetine, methadone and morphine. Methadone and tramadol inhibited synaptosomal [³H]-5-HT uptake with apparent K_is of 0.27±0.04 and 0.76±0.04 μM, respectively. Morphine essentially failed to inhibit [³H]-5-HT uptake (K_i 0.50±0.30 M).
3. Methadone, morphine and tramadol were active in the hot plate test with ED₅₀s of 3.5, 4.3 and 31 mg kg⁻¹, respectively. At the highest tested dose (80 mg kg⁻¹) tramadol produced only 77±5.3% of the maximal possible effect.
4. When [³H]-5-HT uptake was examined in synaptosomes prepared from rats 30 min after a single dose of morphine, methadone or tramadol, only tramadol (31 mg kg⁻¹, s.c., equal to the ED₅₀ in the hot plate test) and methadone (35 mg kg⁻¹, s.c., equal to the ED₉₀ in the hot plate test) decreased uptake.
5. Animals were chronically treated for 15 days with increasing doses of tramadol or methadone (5 to 40 mg kg⁻¹ and 15 to 120 mg kg⁻¹, s.c., respectively). Twenty-four hours after the last drug injection, a challenge dose of methadone (35 mg kg⁻¹, s.c.) or tramadol (31 mg kg⁻¹, s.c.) was administered. [³H]-5-HT uptake was not affected in synaptosomes prepared from rats chronically-treated with methadone, whereas chronic tramadol was still able to reduce this parameter by 42%.
6. Rats chronically-treated with methadone showed a significant increase in [³H]-5-HT uptake (190%) 72 h after drug withdrawal. In contrast, [³H]-5-HT uptake in rats chronically-treated with tramadol (110%) did not differ significantly from control animals.
7. These results further support the hypothesis that [³H]-5-HT uptake inhibition may contribute to the antinociceptive effects of tramadol. The lack of tolerance development of [³H]-5-HT uptake, together with the absence of behavioural alterations after chronic tramadol treatment, suggest that tramadol has an advantage over classical opioids in the treatment of pain disorders.

     

Binding and uptake studies with [3H]CP-93, 129, a radiolabeled selective 5-HT1B receptor ligand

3-(1,2,5,6-Tetrahydro-4-pyridyl)-5-pyrrolo[3,2-b]pyridone, CP-93, 129, is a selective agonist ligand for 5-HT_1B receptors. High affinity binding sites of [³H]CP-93, 129 were found in rat whole brain membranes, which showed K_D and B_max values similar to those for 5-HT_1B sites labeled by [³H]5-HT. Uptake of [³H]CP-93, 129 in crude rat synaptosomes was also observed, which was potently inhibited by 5-HT uptake blockers and 5-HT but not by desipramine (NE uptake blocker) or tametraline (NE and DA uptake blocker). Because of this sensitivity to 5-HT uptake inhibitors and the structural similarity of CP-93, 129 to serotonin, [³H]CP-93, 129 uptake probably occurred in 5-HT neurons.     

Increase in Dopamine Uptake in Rat Striatal Synaptosomes after an Acute in Vivo Administration of Organic and Inorganic Lead

Abstract: Adult male Wistar rats were administered acute toxic doses of lead (Pb), triethyl lead (TriEL) or tetraethyl lead (TEL) by gavage. The ability of striatal, hypothalamic and cortical synaptosomes to take up tritiated monoamines was assayed 24 hours later. Pb, TriEL as well as TEL increased dopamine (DA) uptake into striatal synaptosomes at least at some dose level. Pb increased, but TEL decreased 5-hydroxytryptamine(5-HT) uptake into hypothalamic synaptosomes, while TEL increased noradrenaline (NA) uptake into cortical synaptosomes. After a 3 week administration to initially 4 week old rats of even toxic doses of Pb in drinking water, monoamine uptake was not significantly affected. On the contrary, the neurotoxicity of TEL was cumulative in that a much lower dose decreased 5-HT uptake when divided over a 3 week period than acutely. In vitro TriEL inhibited DA uptake (IC50; 0.8 μM) into striatal and 5-HT uptake (5.0 μM) into hypothalamic synaptosomes but TEL and delta-aminolevulinic acid did not. The results suggest that dopaminergic and serotonergic neurones differ in their response to alkyl lead in vivo. The differences could be due to basic differences in the neurochemical behaviour of these two types of nerve endings.