Effects of a selective 5-HT reuptake blocker,citalopram, on the sensitivity of 5-HT autoreceptors: Electrophysiological studies in the rat brain

Summary Citalopram (CIT), is a selective serotonin (5-HT) reuptake blocker and a clinically effective antidepressant. The present electrophysiological studies were undertaken to investigate in vivo the acute and long-term effects of CIT administration on 5-HT neurotransmission. In a first series of experiments, a single dose of CIT (0.05–0.5 mg/kg) was administered intravenously to naive rats while recording the activity of a 5-HT-containing neuron in the nucleus raphe dorsalis. A dose-response relationship of the inhibitory effect of CIT on the firing activity of 5-HT neurons was obtained with an ED₅₀ of 0.23±0.03 mg/kg. In a second series of experiments, rats were treated with CIT (20 mg/kg/day, i.p.) for 2, 7 and 14 days. In rats treated for 2 days, there was a marked reduction in the firing activity of 5-HT neurons in the nucleus raphe dorsalis; there was a partial recovery after 7 days and a complete recovery after 14 days of treatment. The response of 5-HT neurons to intravenously administered LSD was decreased in rats treated for 14 days with CIT, indicating a desensitization of the somatodendritic 5-HT autoreceptor. In a third series of experiments, carried out in rats treated with CIT (20 mg/kg/day, i.p.) for 14 days, the suppression of firing activity of CA₃ hippocampal pyramidal neurons produced by microiontophoretically-applied 5-HT and by the electrical activation of the ascending 5-HT pathway was measured. Long-term treatment with CIT did not modify the responsiveness of these neurons to microiontophoretically-applied 5-HT; however, the effect of the electrical activation of the ascending 5-HT pathway on these same neurons was enhanced. To determine if 5-HT reuptake blockade could be responsible for this enhancement, CIT (1 mg/kg) was injected intravenously in naive rats while stimulating the ascending 5-HT pathway; it failed to modify the effectiveness of the stimulation. To assess the involvement of the 5-HT terminal autoreceptor, methiothepin, a 5-HT autoreceptor antagonist, was injected intravenously (1 mg/kg) in naive rats and in rats treated for 14 days with CIT while stimulating the ascending 5-HT pathway. Methiothepin enhanced the effect of the stimulation in naive rats but failed to do so in the CIT-treated rats. It is concluded that long-term CIT treatment enhances 5-HT neurotransmission by desensitizing both the somatodendritic and terminal 5-HT autoreceptors.     

Effect of prolonged administration of tianeptine on 5-HT neurotransmission: an electrophysiological study in the rat hippocampus and dorsal raphe

Extracellular unitary recordings of dorsal hippocampus CA₃ pyramidal neurons and of dorsal raphe 5-hydroxytryptamine (5-HT) neurons were used to assess the effect of tianeptine, a putative antidepressant, on the efficacy of 5-HT neurotransmission. Sustained tianeptine administration (20 mg/kg/day, s.c. × 14 days) did not modify the firing activity of 5-HT neurons in the dorsal raphe. Their responsiveness to the intravenous injection of LSD, an agonist of the somatodendritic 5-HT autoreceptor, and of 8-OH-DPAT, a selective 5-HT_1A agonist, was also unaffected by this treatment. The responsiveness of CA₃ pyramidal neurons to microiontophoretic application of 5-HT remained unchanged after sustained tianeptine administration, but it was markedly enhanced in rats treated with repeated electroconvulsive shocks. Finally, the duration of suppression of firing activity of CA₃ pyramidal neurons produced by electrical stimulation of the ascending 5-HT pathway, delivered at 1 Hz and 5 Hz, was not modified in rats treated with tianeptine. Methiothepin, an antagonist of the terminal autoreceptor enhanced the effectiveness of 5-HT pathway stimulation to the same extent in control and tianeptinetreated rats. The present results indicate that, administered at a dose known to stimulate 5-HT reuptake (20 mg/kg/day, s.c.; by minipump), and for a period of time (14 days) for which other antidepressant treatments have been shown to enhance 5-HT function, tianeptine does not modify the efficacy of 5-HT synaptic transmission in the rat hippocampus.     

Species differences in presynaptic serotonin autoreceptors: mainly 5-HT1B but possibly in addition 5-HT1D in the rat, 5-HT1D in the rabbit and guinea-pig brain cortex

Summary The pharmacological properties of presynaptic serotonin autoreceptors were compared in slices of rat, rabbit, and guinea-pig brain cortex. The slices were preincubated with ³H-serotonin and then superfused with medium containing fluvoxamine 3 mol/l and stimulated four times by trains of four pulses delivered at 100 Hz. Cumulative concentration-response curves were determined and used for the calculation of agonist EC₅₀ values and maximal effects and antagonist K _B values.Unlabelled serotonin itself and the serotonin receptor agonists 5-carboxamidotryptamine (5-CT), 5-methoxy-3-(1,2,3,6-tetrahydro-4-pyridinyl)-1H-indole (RU 24969) and (±)-8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) reduced the stimulation-evoked overflow of tritium with a rank order of potency 5-CT = RU 24969 > serotonin > 8-OH-DPAT in the rat and 5-CT > serotonin > RU 24969 > 8-OH-DPAT in the rabbit and guinea-pig. Ipsapirone caused no change. Metitepine and metergoline antagonized the effect of 5-CT; the K _B values were lower in the rabbit and guinea-pig than in the rat. Yohimbine at up to 1 mol/1 did not reduce the evoked overflow of tritium and did not antagonize the inhibitory effect of 5-CT in the rat but reduced the evoked overflow in the rabbit and counteracted the effect of 5-CT in the guinea-pig. (–)-Propranolol, conversely, reduced the evoked overflow of tritium in the rat but neither reduced the evoked overflow nor antagonized the effect of 5-CT in the rabbit and guinea-pig. Isamoltane did not significantly change the effect of 5-CT in any species. In the rat, it also failed to antagonize the inhibitory effect of 8-OH-DPAT but did antagonize the effect of RU 24969. The inhibition caused by 8-OH-DPAT persisted in the presence of idazoxan but was attenuated by metitepine in all species.The experimental conditions used permit the determination of the constants of agonist and antagonist action undistorted by autoinhibition. The results confirm the view that the serotonin axons of rat brain possess 5-HT_1B autoreceptors. They show by direct comparison under identical conditions that the autoreceptors in rabbit and guinea-pig are very similar to each other but differ markedly from those in the rat. The results give additional credence to previous suggestions that, in the rabbit and guinea-pig, the autoreceptors are 5-HT_1D. The serotonin axons of rat brain cortex may possess 5-_1D in addition to 5-HT_1B autoreceptors. In many previous studies agonist potencies at, and antagonist affinities for, presynaptic serotonin autoreceptors have been underestimated due to the use of too intense stimuli to elicit serotonin release. Send offprint requests to N. Limberger at the above address     

5-HT autoreceptors in the regulation of 5-HT release from guinea pig raphe nucleus and hypothalamus

5-HT autoreceptors involved in the regulation of 5-HT release in the guinea pig dorsal raphe nucleus have been studied in comparison with those in the hypothalamus. In vitro release was measured in slices of raphe and hypothalamus prelabelled with [³H]5-HT, superfused with Krebs solution and depolarized electrically. The non-selective 5-HT receptor agonist, 5-carboxamidotryptamine (5-CT) (0.1–10 nM for raphe; 1–100 nM for hypothalamus) and antagonist, methiothepin (10–1000 nM), decreased and increased, respectively, the release of [³H]5-HT evoked by electrical stimulation in either of these regions when given alone. The selective 5-HT_1B/D receptor antagonist, GR127935 (100–1000 nM), and the 5-HT_1D receptor antagonist, ketanserin (300–1000 nM), had no significant effect on this release in either of these regions. Methiothepin and GR127935 (100–1000 nM) shifted to the right the concentration-effect curve of 5-CT in both the raphe and the hypothalamus. At 300 nM, ketanserin shifted to the right the concentration-effect curve of 5-CT in the raphe but did not modify the 5-CT curve in the hypothalamus. In microdialysis experiments ketanserin, applied locally at 10 μM, increased the extracellular levels of 5-HT in the dorsal raphe nucleus of the freely moving guinea pig, whereas 5-HT levels were unchanged in the hypothalamus. Ketanserin at 1 μM did not affect the decrease in 5-HT output induced by the selective 5-HT_1B/D receptor agonist, naratriptan (used at 10 μM in raphe and 0.1 μM in hypothalamus), in the raphe or the hypothalamus. In the raphe, WAY100635, a 5-HT_1A receptor antagonist, at 1 μM, did not prevent naratriptan (10 μM) from reducing the extracellular levels of 5-HT. These results suggest that, in the conditions used in this study, the release of 5-HT in the dorsal raphe nucleus is possibly modulated in part by 5-HT_1B receptors but essentially the control is through 5-HT receptors whose subtype is still to be determined. In the hypothalamus, however, it is clear that only 5-HT_1B receptors are involved in the modulation of 5-HT neurotransmission.     

In vivo control of 5-hydroxytryptamine release by terminal autoreceptors in rat brain areas differentially innervated by the dorsal and median raphe nuclei

Selective serotonin reuptake inhibitors (SSRIs) reduce the 5-HT release in vivo. This effect is due to the activation of somatodendritic 5-HT_1A receptors and it displays a regional pattern comparable to that of selective 5-HT_1A agonists, i.e., preferentially in forebrain areas innervated by the dorsal raphe nucleus (DRN). However, despite a comparatively lower 5-HT_1A-mediated inhibition of 5-HT release and a greater density of serotonergic uptake sites in hippocampus, the net elevation produced by the systemic administration of SSRIs is similar in various forebrain areas, regardless of the origin of serotonergic fibres. As terminal autoreceptors may also limit the SSRI-induced elevations of 5-HT in the extracellular brain space, we reasoned that a differential control of 5-HT release by terminal autoreceptors in DRN- and median raphe-innervated areas might be accountable. To examine this possibility, we have conducted a regional microdialysis study in the DRN, MRN and four forebrain regions preferentially innervated either by the DRN (frontal cortex, striatum) or the median raphe nucleus (MRN; dorsal and ventral hippocampus) using freely moving rats. Dialysis probes were perfused with 1 μM of the SSRI citalopram to augment the endogenous tone on terminal 5-HT autoreceptors. The non-selective 5-HT₁ antagonist methiothepin (10 and 100 μM, dissolved in the dialysis fluid) increased extracellular 5-HT in frontal cortex and dorsal hippocampus in a concentration-dependent manner. The 5-HT_1B/1D antagonist GR 127935 was ineffective at 10 μM and tended to reduce 5-HT in dorsal hippocampus at 100 μM. The local infusion of 100 μM methiothepin significantly elevated the extracellular 5-HT concentration to 142–173% of baseline (mean values of 260 min post-administration) in the DRN, MRN, frontal cortex, striatum and hippocampus (dorsal and ventral). Comparable elevations were noted in the four forebrain regions examined. As observed in frontal cortex and dorsal hippocampus, the perfusion of 10 μM GR 127935 did not elevate 5-HT in DRN, MRN, striatum or ventral hippocampus. Because the stimulated 5-HT release in the DRN has been suggested to be under control of 5-HT_1B/1D receptors, we examined the possible contribution of these receptor subtypes to the effects of methiothepin in the DRN. The perfusion of sumatriptan (0.01–10 μM) or GR 127935 (0.01–10 μM) did not significantly modify the 5-HT concentration in dialysates from the DRN. Thus, the present data suggest that the comparable effects of SSRIs in DRN- and MRN-innervated forebrain regions are not explained by a preferential attenuation of 5-HT release by terminal 5-HT_1B autoreceptors in hippocampus, an area with a low inhibitory influence of somatodendritic 5-HT_1A receptors. Methiothepin-sensitive autoreceptors (possibly 5-HT_1B) appear to play an important role not only in the projection areas but also with respect to the control of 5-HT release in the DRN and MRN. In addition, our findings indicate that GR 127935 is not an effective antagonist of the actions of 5-HT at rat terminal autoreceptors. Received: 27 February 1998 / Accepted: 12 June 1998     

Identification of presynaptic 5-HT1 autoreceptors in pig brain cortex synaptosomes and slices

Summary Pig brain cortex synaptosomes and slices preincubated with ³H-5-hydroxytryptamine (³H-5-HT) were superfused with physiological salt solution containing citalopram (an inhibitor of 5-HT uptake), and the effects of indolethylamines and 5-HT receptor antagonists on the potassium- or electrically evoked ³H overflow were determine. The potassium (25 mmol/l)-evoked tritium overflow from cortex synaptosomes was inhibited by 5-HT; the inhibitory effect of 5-HT was counteracted by metitepine, which, by itself, did not affect the evoked overflow. 5-Methoxytryptamine (examined in the absence of citalopram) also produced an inhibition of the evoked overflow. In cortex slices, the electrically (3 Hz) evoked overflow was inhibited by 5-HT and 5-carboxamidotryptamine. The inhibitory effect of 5-HT was antagonized by metitepine, which, given alone, increased the evoked overflow, but was not attenuated by ketanserin and ICS 205-930 ([³-tropanyl]-1H-indole-3-carboxylic acid ester), which, by themselves, did not influence the evoked overflow. The present results suggest that the serotoninergic nerve fibres of the pig brain cortex are endowed with presynaptic 5-HT₁ receptors, which can be activated by endogenous and exogenous 5-HT.Send offprint requests to E. Schlicker at the above address     

Serotonin autoreceptor in rat hippocampus: Pharmacological characterization as a subtype of the 5-HT1 receptor

Summary The 5-hydroxytryptamine (5-HT) autoreceptors mediating inhibition of [³H]5-HT release in rat hippocampus have been characterized pharmacologically in terms of 5-HT receptor subtype by using superfused synaptosomes depolarized with 15 mM KCl. Exogenous 5-HT inhibited in a concentration-dependent way (pEC₃₀=8.74) the K⁺-evoked release of [³H]5-HT. Methiothepin shifted the concentration-response curve of 5-HT to the right (pA₂=8.62). The 5-HT₂ receptor antagonists, ketanserin, methysergide or spiperone were ineffective against 5-HT. The 5-HT₁ receptor agonist, 5-methoxy-3-[1,2,3,6-tetra-hydropyridin-4-yl]-1H-indole (RU 24969) mimicked 5-HT and was equipotent as an inhibitor of the release of [³H]5-HT. In contrast, the putative 5-HT_1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) was almost ineffective at 1 M. Finally, (–)propranolol, used as a non-selective 5-HT_1A/5-HT_1B receptor antagonist, shifted to the right (pA₂=7.91) the concentration-response curve of 5-HT whereas the 5-HT_1C receptor antagonist mesulergine was ineffective. In conclusion, 5-HT nerve terminals of rat hippocampus possess autoreceptors which appear to belong to the 5-HT_1B subtype.     

Inhibition of 5-HT cell firing in the DRN by non-selective 5-HT reuptake inhibitors: studies on the role of 5-HT1A autoreceptors and noradrenergic mechanisms

Improved clinical antidepressant efficacy may result if the acute inhibition of 5-HT cell firing induced by antidepressants is prevented. Here we examined whether inhibition of 5-HT cell firing by non-selective 5-HT uptake inhibiting antidepressant drugs is reversed by a selective 5-HT_1A receptor antagonist. In addition, we examined whether concomitant blockade of NA uptake offsets the inhibition of 5-HT cell firing resulting from 5-HT uptake blockade. Antidepressants which block 5-HT uptake (paroxetine, clomipramine, amitriptyline, venlafaxine), all caused dose-dependent and complete inhibition of 5-HT cell firing. Desipramine, a selective NA uptake blocker, caused a slight reduction in firing. The selective 5-HT_1A receptor antagonist, WAY 100635, reversed the inhibition of 5-HT cell firing induced by clomipramine, amitriptyline, venlafaxine, and paroxetine, but not that induced by the α₁ adrenoceptor antagonist, prazosin. Desipramine, at a dose which increased extracellular NA in the DRN, reversed the effect of prazosin but did not alter the ability of paroxetine to inhibit 5-HT cell firing. Our data indicate that antidepressant drugs with 5-HT uptake blocking properties inhibit 5-HT cell firing via activation of 5-HT_1A autoreceptors, and do so irrespective of their effects on NA uptake. These data are discussed in relation to the application of 5-HT_1A receptor antagonists to enhance the clinical efficacy of antidepressant drugs. Received: 15 July 1996 / Final version: 11 November 1996     

Evidence for presynaptic location of inhibitory 5-HT1D\-like autoreceptors in the guinea-pig brain cortex

The effects of 5-hydroxytryptamine (5-HT) receptor agonists and antagonists on tritium overflow evoked by high K⁺ were determined in superfused synaptosomes and slices, preincubated with [³H]5-HT, from guinea-pig brain cortex. In addition, we estimated the potencies of 5-HT receptor ligands in inhibiting specific [³H]5-HT binding (in the presence of 8-hydroxy-2(di-n-propylamino)tetralin and mesulergine to prevent binding to 5-HT_1A and 5-HT_2C sites) to guinea-pig cortical synaptosomes and membranes.5-HT receptor agonists inhibited the K⁺-evoked tritium overflow from synaptosomes and slices. In synaptosomes the rank order of potencies was 2-[5-[3-(4-methylsulphonylamino)benzyl-1,2,4-oxadiazol-5-yl]-1H-indole-3-yl] ethylamine (L-694,247) >5-carboxamidotryptamine (5-CT) > oxymetazoline (in the presence of idazoxan) 5-HT > sumatriptan 5-methoxy-3(1,2,3,6-tetrahydropyridin-4-yl)-1H-indole (RU 24969). The potencies of the agonists in inhibiting tritium overflow from slices correlated with those in synaptosomes, suggesting that the same site of action is involved in both preparations. In synaptosomes the nonselective antagonist at cloned human 5-HT_1D, and 5-HT_1D receptors, methiothepin, shifted the concentration-response curve for 5-CT to the right (apparent pA₂: 7.87). In contrast, ketanserin at a concentration which should block the 5-HT_1D, but not the 5-HT_1D\, receptor did not alter the inhibitory effect of 5-CT on tritium overflow. In cortical synaptosomes and membranes, [³H]5-HT bound to a single site with high affinity. In competition experiments, 5-HT receptor agonists and antagonists inhibited specific [³H]5-HT binding. In synaptosomes the rank order was L-694,247 > methiothepin >5-CT >5-methoxytryptamine >5-HT sumatriptan oxymetazoline > RU 24969 > ketanserin > ritanserin. A very similar rank order was obtained in cerebral cortical membranes. The potencies of the 5-HT receptor agonists in inhibiting tritium overflow from synaptosomes and slices correlated with their potencies in inhibiting [³H]5-HT binding to synaptosomes and membranes.In conclusion, the 5-HT receptors mediating inhibition of 5-HT release in the guinea-pig cortex are located on the serotoninergic axon terminals and, hence, represent presynaptic inhibitory autoreceptors. The [³H]5-HT binding sites in cerebral cortical synaptosomes and membranes exhibit the pharmacological properties of 5-HT_1D receptors. The correlation between the functional responses and the binding data confirms the 5-HT_1D character of the presynaptic 5-HT autoreceptors. According to the results of the interaction experiment of ketanserin and methiothepin with 5-CT on 5-HT release, the presynaptic 5-HT autoreceptors can be subclassified as 5-HT_1D\-like.     

The 5-HT1A receptor agonist, 8-OH-DPAT,preferentially activates cell body 5-HT autoreceptors in rat brain in vivo

Summary The present study was undertaken in an attempt to assess whether the effects of the potent and selective 5HT_1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin, 8-OH-DPAT, on cerebral 5-hydroxytryptamine (5-HT) neurochemistry in vivo are mediated via 5-HT autoreceptors on the cell bodies or on the terminals, and/or via postsynaptic 5-HT receptors. To this end we determined in vivo indices of 5-HT synthesis and release/turnover rates in a number of prominent 5-HT neuronal projection areas in the CNS i) after systemic administration of 8-OH-DPAT to rats with an acute unilateral axotomy of the ascending mesencephalic monoamine neurones, or ii) after local infusion of the compound into the dorsal raphé (DRN) 5-HT cell body region of intact rats. Transection did not alter 5-HT synthesis per se, but prevented the synthesis-inhibitory effect of 8-OH-DPAT. Thus, the 5-HT synthesis-inhibiting action of 8-OH-DPAT is highly dependent upon intact impulse flow in the central 5-HT neurones. On the other hand, local DRN application of the compound (1 g) resulted in a clearcut reduction of the 5-HT synthesis and release indices measured in 5-HT terminals in, e. g., the striatum. These findings provide direct neurochemical evidence that by preferentially stimulating somatodendritic 5-HT_1A receptors, 8-OH-DPAT inhibits the 5-HT neuronal impulse flow, thereby effectuating decreased terminal 5-HT synthesis and release. Taken together, the data are consistent with the suggestion that 8-OH-DPAT acts as an agonist preferentially at cell body vs. terminal 5-HT autoreceptors, therefore also emphasizing the distinction between terminal and cell body 5-HT autoreceptors. The results obtained may have important implications for the understanding of mechanisms involved in regulating the activity of central serotoninergic neurones.Part of these data were presented at the 6th European Winter Conference on Brain Research, Avoriaz, France, March 9–15, 1985, and at the 18th Annual Meeting, Society for Neuroscience, Washington (DC), USA, Nov. 9–15, 1986 (Hjorth et al. 1986, 1987). Send offprint requests to S. Hjorth at the above address     

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     

5-HT1A receptor-mediated autoinhibition does not function at physiological firing rates: evidence from in vitro electrophysiological studies in the rat dorsal raphe nucleus

5-HT(1A)-mediated autoinhibition of neurones in the dorsal raphe nucleus (DRN) is considered to be the principal inhibitory regulator of 5-HT neuronal activity. The activation of this receptor by endogenous 5-HT was investigated using electrophysiological recordings from the rat DRN in vitro. At a concentration which blocked the inhibitory effect of exogenous 5-HT, the 5-HT(1A) antagonist WAY 100635 did not alter basal firing rate or modulate the excitatory response to the alpha(1)-agonist phenylephrine. Blockade of 5-HT reuptake by a concentration of fluoxetine, which enhanced the inhibitory effect of exogenous 5-HT, lowered phenylephrine-induced basal firing presumably due to potentiation of the effect of endogenous 5-HT. However, this effect was not firing rate dependent and neither the proportional increase nor the time-course of the response to a higher concentration of phenylephrine were altered in the presence of fluoxetine. These data suggest that the inhibitory 5-HT(1A) receptor on raphe neurones is neither tonically activated nor plays any role in modulating the response to excitatory transmitters. Thus, at physiological firing rates this receptor does not appear to function as an autoreceptor of serotonergic neurones of the DRN.     

Effects of co-administration of a selective serotonin reuptake inhibitor and monoamine oxidase inhibitors on 5-HT-related behavior in rats

5-hydroxytryptamine (5-HT) syndrome is a dangerous condition of 5-HT excess that can occur in the case of co-administration of a monoamine oxidase (MAO) inhibitor and a serotonin reuptake inhibitor (SSRI). The goal of the present study was to investigate the effects of acute administration of MAO inhibitors and subchronic administration of fluvoxamine on 5-HT-related behaviors (head shaking and 5-HT syndrome) in rats treated with 5-hydroxytryptophan (5-HTP). Administration of the non-selective MAO inhibitor, pargyline, and the selective MAO-A inhibitor, clorgyline, resulted in 5-HT syndrome in 5-HTP-treated rats, and subchronic co-administration of fluvoxamine intensified the syndrome. However, administration of the selective MAO-B inhibitor, selegiline, did not induce 5-HT syndrome with or without subchronic fluvoxamine co-administration. These data suggest that non-selective MAO and selective MAO-A inhibitors can induce 5-HT syndrome in humans when co-administered with SSRI. Further, the risk of 5-HT syndrome may be lower with the selective MAO-B inhibitor, selegiline.     

Suppression of the hypo- and hyperthermic responses to 5-HT agonists following the repeated administration of monoamine oxidase inhibitors

Hypo- and hyperthermic responses resulting from the activation of putative 5-HT_1A and 5-HT₂ receptors, respectively, were examined after the chronic treatment of rats with monoamine oxidase inhibitors. The treatment of rats for 4 or 7 days with nialamide (40 mg/kg, twice daily) resulted in a suppression of the hypothermic effect of the 5-HT_1A agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT, 0.05–0.25 mg/kg, SC). The decrease in body temperature elicited by a low dose of 5-methoxy-N, N-dimethyltryptamine (5MeODMT, 1 mg/kg) also was diminished in rats treated chronically with nialamide. The administration of a high dose of 5MeODMT (5 mg/kg) resulted in a hyperthermic response, which was also attenuated after the repeated administration of nialamide. The repeated administration of clorgyline (a selective inhibitor of type A MAO) or deprenyl (a selective inhibitor of type B MAO) failed to alter the hypothermic effect of 8-OH-DPAT. However, in animals treated chronically with both clorgyline and deprenyl, a suppressed response to 8-OH-DPAT was observed. In view of the concept that the hypo- and hyperthermic responses to 5-HT agonists are mediated by 5-HT_1A and 5-HT₂ receptor subtypes, respectively, it is concluded that the responsiveness of these 5-HT receptor subtypes involved in thermoregulatory responses is decreased following chronic treatment of rats with monoamine oxidase inhibitors. It appears that inhibition of both type A and B MAO is necessary for this desensitization process.     

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.     

The effects of quipazine on 5-HT metabolism in the rat brain

Summary Since quipazine is a potent 5-HT agonist in peripheral organs, its possible stimulatory effects on serotoninergic receptors in the rat brain were investigated. Quipazine administration (10 mg/kg, i.p.) induced a significant decrease in the synthesis and turnover rates of serotonin in the brain stem as well as in the forebrain. It is not likely that these changes were mediated by a negative feed-back mechanism triggered by adirect action of quipazine on central 5-HT postsynaptic receptors. Indeed, in contrast to LSD and 5-methoxy-N,N-dimethyltryptamine, this compound failed to activate the 5-HT sensitive adenylate cyclase in colliculi homogenates of newborn rats. However, quipazine exerted direct effects on serotoninergic terminals. It inhibited competitively the reuptake process in synaptosomes (Ki =1.38×10^–7 M) and stimulated the K⁺ evoked release of newly synthesized³H-5-HT in slices of the brain stem. Injected in vivo in a dose which affected 5-HT uptake and release, quipazine did not modify MAO activity. However, this activity was noncompetitively inhibited by high concentration of the drug in vitro (Ki=3.0×10^–5 M). These actions are very likelyindirectly responsible for the stimulation of central 5-HT receptors.     

The role of 5-HT receptor subtypes in the anxiolytic effects of selective serotonin reuptake inhibitors in the rat ultrasonic vocalization test

 We evaluated whether the anxiolytic effects of selective serotonin reuptake inhibitors (SSRIs) in the rat ultrasonic vocalization (USV) test are preferentially mediated by (indirect) activation of 5-HT_1A, 5-HT_1B/1D, 5-HT_2A, 5-HT₃ or 5-HT₄ receptors. The SSRIs, paroxetine (ED₅₀ in mg/kg, IP: 6.9), citalopram (6.5), fluvoxamine (11.7) and fluoxetine (>30), dose dependently reduced shock-induced USV. The effects of paroxetine (3.0 mg/kg, IP) were not blocked by the selective 5-HT_1A receptor antagonist, WAY-100635 (3.0 mg/kg, IP), the 5-HT_1B/1D receptor antagonist, GR 127935 (30 mg/kg, IP), the nonselective 5-HT_2A receptor antagonists, ritanserin (3.0 mg/kg, IP) and ketanserin (1.0 mg/kg, IP), the 5-HT₃ receptor antagonist, ondansetron (0.1 mg/kg, IP), or the 5-HT₄ receptor antagonist, GR 125487D (3.0 mg/kg, SC). In contrast, the selective 5-HT_2A receptor antagonist, MDL 100,907 (0.1 mg/kg, IP), completely prevented the paroxetine-induced reduction of USV. Under similar conditions, WAY-100635 blocked the anxiolytic-like effects of the selective 5-HT_1A receptor agonist, 8-OH-DPAT [(±)-8-hydroxy-2-(di-n-propylamino)tetralin, 1.0 mg/kg, IP], and ritanserin, ketanserin, and MDL 100,907 blocked the anxiolytic-like effects of the mixed 5-HT_2A/2C receptor agonist, DOI [1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane, 3.0 mg/kg, IP]. WAY-100635 (1.0 mg/kg, IP) in combination with ritanserin (3.0 mg/kg, IP), but not ondansetron (0.1 mg/kg, IP), GR 125487D (3.0 mg/ kg, SC), or GR 127935 (30 mg/kg, IP), attenuated the USV reducing effects of paroxetine. Although the results suggest that selective stimulation of 5-HT_1A and 5-HT_2A receptors produces a decrease of USV, we postulate that only 5-HT_2A receptors play a pivotal role in the effects of SSRIs in this model of anxiety. Received: 19 May 1997 / Final version: 21 July 1997     

Interaction between 5-HT uptake inhibition and activation of 5-HT autoreceptors by exogenous agonists in rat cerebral cortex slices and synaptosomes

Summary Rat cerebral cortex slices or synaptosomes were labelled with ³H-5-hydroxytryptamine (³H-5-HT) and subsequently superfused. They were depolarized by electrical stimulation (slices) or with high K⁺ (slices and synaptosomes). Continuous electrical stimulation (2 Hz, 24 mA, 2 ms) and continuous or discontinuous K⁺ depolarization (15–25 mM) were used. 1. Continuous electrical stimulation or continuous K⁺-depolarization of slices evoked a steady overflow of tritium that slowly decayed with time. 2. Exposure to increasing concentrations of 5-methoxy-3(1,2,3,6-tetrahydropyridin-4-yl)-1H-indole succinate (RU 24969) (0.001–0.1 M) during continuous electrical stimulation produced a concentration-dependent decrease in tritium overflow. Citalopram (1 M) counteracted the effect of RU 24969. 3. RU 24969 inhibited the evoked ³H-overflow and citalopram reduced the effect of RU 24969 also during continuous depolarization of slices with 20 mM K⁺. Similar results were obtained by using 5-methoxytryptamine or LSD. 4. In slices 1 M citalopram increased significantly the tritium overflow evoked by electrical stimulation or by 20 mM K⁺-depolarization. 5. Increasing the K⁺ concentration from 20 mM to 25 mM mimicked the effects of 1 M citalopram both on the RU 24969 activity and on the evoked tritium overflow. 6. RU 24969 (0.001–0.1 M) decreased in a concentration-dependent way the release of tritium from cortical synaptosomes depolarized with K⁺ (15–20 mM). The presence of 1 M citalopram did not modify significantly the effect of the agonist. Citalopram was ineffective also when the serotonin uptake carrier in superfused synaptosomes was activated by tryptamine. In conclusion, in slices of rat cerebral cortex, the action of exogenous 5-HT autoreceptor agonists is inhibited by 5-HT uptake blockers independently of the depolarizing agent (electrical stimulation or high-K⁺) used to elicit ³H-5-HT release. Increasing K⁺-concentration, which probably increases serotonin in the biophase, mimics the presence of the reuptake inhibitor. These data together with the finding that, in superfused synaptosomes, 5-HT uptake inhibition did not affect the potency of autoreceptor agonists, favours the idea that, in cerebral cortex slices, inhibitors of 5-HT reuptake prevent activation of autoreceptors by exogenous agonists by increasing the concentration of 5-HT in the autoreceptor biophase. Send offprint requests to M. Raiteri at the above address     

Mechanism of tolerance development to 2,5-dimethoxy-4-iodoamphetamine in rats: down-regulation of the 5-HT2A, but not 5-HT2C, receptor

  Rationale: Defining the mechanism of tolerance development to hallucinogenic drugs will help to explain their mechanism of action. Objectives: The present study was conducted to determine first, if tolerance develops to the discriminative stimulus (DS) properties of the hallucinogen, 2,5 dimethoxy-4-iodo-amphetamine (DOI) and second, the mechanism mediating tolerance. Methods: Rats were trained to discriminate 0.75 mg/kg DOI from saline on a concurrent VI-30-min schedule of reinforcement with a 15-min time-out for incorrect responses. To evaluate tolerance development, rats were assigned to one of four groups and treated with either chronic saline or chronic DOI. Prior to chronic treatment, two groups were tested for choice behavior following vehicle administration while the remaining two groups were tested following the administration of 0.375 mg/kg DOI. One group from each pre-test condition was injected with either saline or DOI (1 mg/kg) for 8 days. Twenty-four hours after the last chronic injection the pre-test treatments were replicated. Using receptor autoradiography, the density of 5-HT_2A and 5-HT_2C receptors was measured in independent groups of rats that had received identical treatment conditions. Results: Animals receiving chronic DOI showed a 60% decrease in DOI lever responding (from 100% to 40%) when tested on 0.375 mg/kg DOI, while animals receiving chronic saline showed no change in percent choice (100%) on the DOI lever. Significant changes in binding were observed in 5-HT_2A receptors but not 5-HT_2C receptors. The results of tests with antagonists were consistent with the changes in binding. Conclusions: These results suggest that behavioral tolerance to DOI reflects neuroadaptive changes in 5-HT_2A receptors. Received: 17 July 1998 / Final version: 19 January 1999     

Local modulation of the 5-HT release in the dorsal striatum of the rat: an in vivo microdialysis study

Using in vivo microdialysis in freely moving rats, we examined the involvement of major striatal transmitters on the local modulation of the 5-HT release. Tetrodotoxin reduced the striatal 5-HT output to 15–20% of baseline. The selective 5-HT_1B receptor agonist CP 93129 (50 μM) reduced (50%) and the 5-HT_2A/2C receptor agonist DOI (1–100 μM) increased (220%) the 5-HT output. Neither GABA nor baclofen (100 nM–100 μM) altered the 5-HT output. The glutamate reuptake inhibitor -trans-PDC (1–4 mM) raised 5-HT to 280% of baseline. This effect was not antagonized by the NMDA receptor antagonist MK-801 (0.5 mg/kg i.p.). Local MK-801 (10–100 μM) did not significantly alter the 5-HT output. Finally, neither carbachol (10–100 μM) nor quipirole (10 μM–1 mM) affected 5-HT. These data suggest that the striatal 5-HT release is influenced by local serotonergic and glutamatergic (but not GABAergic) inputs.