Nuria Bel

Fluvoxamine preferentially increases extracellular 5-hydroxytryptamine in the raphe nuclei: an in vivo microdialysis study

The effects of systemic administration of fluvoxamine on extracellular serotonin (5-hydroxytryptamine, 5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) concentrations in the frontal cortex and raphe nuclei of freely moving rats were examined. Fluvoxamine significantly increased extracellular 5-HT concentrations in both regions at the two doses used (1 and 10 mg/kg i.p.). However, the increase in the raphe nuclei was several-fold that in the frontal cortex. Dialysate 5-HIAA concentrations decreased after treatment with fluvoxamine. These results confirm that 5-HT uptake inhibitors preferentially increase extracellular concentrations of 5-HT in the vicinity of cell bodies and dendrites of serotonergic neurones.

Effect of pindolol on the function of pre- and postsynaptic 5-HT1A receptors : In vivo microdialysis and electrophysiological studies in the rat brain

In microdialysis studies, somatodendritic 5-HT1A receptors in the dorsal raphe nucleus (DRN) were activated by the local infusion of 50 μM citalopram, a selective 5-HT reuptake inhibitor (SSRI). This reduced extracellular 5-HT by about 50% in dorsal striatum, an area receiving 5-HT afferents exclusively from the DRN. (−)Pindolol dose-dependently attenuated this citalopram-induced reduction of striatal extracellular 5-HT. Consistent with its 5-HT reuptake blocking properties, single doses of the SSRI paroxetine (1 and 3 mg/kg IP) and citalopram (1 mg/kg IP) significantly elevated extracellular 5-HT in the dorsal striatum. Pretreatment with (−)pindolol (15 mg/kg IP) potentiated the effect of 3 mg/kg paroxetine and 1 mg/kg citalopram on striatal extracellular 5-HT. A 2-day treatment with 10 mg/kg/day (SC) of paroxetine reduced by 60% the spontaneous activity of 5-HT neurons of the DRN. However, 5-HT neurons displayed normal activity in rats treated with paroxetine and (−)pindolol for 2 days. The inhibitory effect of LSD on 5-HT neuronal firing activity was also markedly attenuated in (−)pindolol-treated rats, indicating that somatodendritic 5-HT1A receptors were blocked by (−)pindolol. To determine whether (−)pindolol also blocked postsynaptic 5-HT1A receptors in hippocampus, 5-HT and the prototypical 5-HT1A agonist 8-OH-DPAT were applied by microiontophoresis onto CA3 pyramidal neurons following the same treatment. (−)Pindolol did not modify the responsiveness of these neurons to 5-HT and 8-OH-DPAT. Taken together, these results indicate that (−)pindolol can potentiate the effects of an SSRI on extracellular 5-HT concentration by preventing the activation of somatodendritic 5-HT1A autoreceptors resulting from the blockade of the 5-HT transporter in the raphe. This presumably leads to enhanced 5-HT neurotransmission because (−)pindolol would not alter the responsiveness of certain postsynaptic 5-HT1A receptors, such as those located on hippocampal CA3 pyramidal neurons. These results provide a neurobiological basis for the reported potentiation of certain antidepressant drugs by pindolol in major depression.

Reduction of Serotonergic Function in Rat Brain by Tryptophan Depletion: Effects in Control and Fluvoxamine-Treated Rats

Abstract: The administration of tryptophan (Trp)‐free amino acid mixtures to depressed patients responding to serotonin [5‐hydroxytryptamine (5‐HT)] uptake inhibitors (SSRIs) worsens their clinical state. This procedure reduces Trp availability to brain and thus impairs 5‐HT synthesis. We have examined the influence of Trp depletion on extracellular 5‐HT and 5‐hydroxyindoleacetic acid (5‐HIAA) concentrations in the rat brain using in vivo microdialysis. The treatment with the SSRI fluvoxamine significantly increased 5‐HT content in dialysates from frontal cortex, as compared with control rats (10.2 ± 2.7 vs. 3.1 ± 0.4 fmol per fraction), whereas 5‐HIAA was unaffected. Food deprivation for 20 h reduced dialysate 5‐HT content to almost control values in fluvoxamine‐treated rats (10.2 ± 2.7 vs. 4.3 ± 0.6 fmol per fraction) but did not alter dialysate 5‐HIAA content (7.8 ± 0.4 vs. 7.2 ± 0.5 pmol per fraction). The administration of Trp‐free amino acid mixtures to fluvoxamine‐treated rats significantly attenuated the release of 5‐HT in frontal cortex (∼50%) and, to a lesser extent, in the midbrain raphe nuclei. This effect was more marked in rats not deprived from food before the experiments (67% reduction of dialysate 5‐HT content in frontal cortex) and was absent in control rats (treated with saline). In contrast, dialysate 5‐HIAA was markedly affected by Trp depletion in all groups, including controls (65–75% reductions). These data show that the administration of an amino acid mixture with the same composition and dose (in milligrams per kilogram of body weight) as those inducing a severe mood impairment in depressed patients reduces 5‐HT and 5‐HIAA concentrations in brain dialysates. The reduction of 5‐HT release, however, occurs only in animals previously treated with the antidepressant fluvoxamine for 2 weeks, which would be consistent with a marked reduction of 5‐HT‐mediated transmission in treated depressed patients but not in healthy controls.

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-HT1A receptors and it displays a regional pattern comparable to that of selective 5-HT1A agonists, i.e., preferentially in forebrain areas innervated by the dorsal raphe nucleus (DRN). However, despite a comparatively lower 5-HT1A-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-HT1B/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-HT1B/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-HT1B autoreceptors in hippocampus, an area with a low inhibitory influence of somatodendritic 5-HT1A receptors. Methiothepin-sensitive autoreceptors (possibly 5-HT1B) 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.

Antidepressant drugs inhibit a glial 5-hydroxytryptamine transporter in rat brain

We assessed the role of glial cells in the uptake of serotonin (5‐hydroxytryptamine, 5‐HT). Primary cultures of rat and mouse cortical astrocytes took up and deaminated 5‐HT. The antidepressants citalopram, clomipramine, fluoxetine, fluvoxamine, paroxetine and sertraline inhibited this process. The presence of the mRNAs for the 5‐HT transporter and monoamine oxidase‐A (MOA‐A) was established in cultured astrocytes and in adult rat brain areas with (midbrain and brainstem) and without (frontal cortex) serotonergic cell bodies after reverse transcription‐polymerase chain reaction and hybridization with probes complementary to the cloned neuronal 5‐HT transporter and MAO‐A. To examine in vivo the role of astrocytes in the elimination of 5‐HT from the extracellular brain space, 5‐HT was perfused through dialysis probes implanted in the frontal cortex of conscious rats and its concentration was measured at the probe outlet. Tissue 5‐HT recovery was dose‐dependently inhibited by the concurrent perfusion of citalopram, fluoxetine and paroxetine, showing that it essentially measured uptake through the high‐affinity 5‐HT transporter. Rats lesioned with 5,7‐dihydroxytryptamine (5,7‐DHT; 88% reduction of tissue 5‐HT) displayed tissue 5‐HT recovery slightly higher than sham‐operated rats (55 ± 2 vs. 46 ± 3%, P < 0.001), a finding perhaps attributable to the astrogliosis induced by 5,7‐DHT denervation. Rats lesioned with 6‐hydroxydopamine showed tissue 5‐HT uptake similar to controls, suggesting negligible reuptake of 5‐HT by catecholaminergic terminals. These results are consistent with the presence of a glial component of 5‐HT uptake in the rodent brain, sensitive to antidepressants, which takes place through a 5‐HT transporter very similar or identical to that present in neurons.

Basal and stimulated extracellular serotonin concentration in the brain of rats with altered serotonin uptake.

We examined the relationship between the density of serotonergic (5‐hydroxytryptamine [5‐HT]) uptake sites and extracellular 5‐HT concentration in the rat brain using microdialysis with two different models, lesions with 5,7‐dihydroxytryptamine (50 μg in the dorsal raphe nucleus (DRN) 15 days before) and sublines of rats genetically selected displaying extreme values of platelet 5‐HT uptake. Compared to controls, lesioned rats had a reduced cortical concentration of 5‐hydroxyindoles (45%), unchanged basal extracellular 5‐HT in the DRN and ventral hippocampus (VHPC), and reduced basal 5‐hydroxyindoleacetic acid (5‐HIAA) concentrations (46%, DRN; 22%, VHPC). Yet the perfusion of 100 mmol/L KCl or 1 μmol/L citalopram elevated dialysate 5‐HT significantly more in the DRN and VHPC of controls. In genetically selected rats, platelet 5‐HT content and uptake were highly correlated (r2 = 0.9145). Baseline dialysate 5‐HT (VHPC) was not different between high and low 5‐HT rats and from normal Wistar rats. However, KCl or citalopram perfusion increased dialysate 5‐HT significantly more in high 5‐HT than in low 5‐HT rats, and the former displayed a greater in vivo tissue 5‐HT recovery. Significant but small differences in the same direction were noted in [3H]citalopram binding in several brain areas, as measured autoradiographically. Thus, basal extracellular 5‐HT (but not 5‐HIAA) concentrations are largely independent on the density of serotonergic innervation and associated changes in uptake sites. However, marked differences emerge during axonal depolarization or reuptake blockade. The significance of these findings for the treatment of mood disorders in patients with neurological disorders is discussed. Synapse 28:313–321, 1998.

Relationship between blood serotonergic variables, melancholic traits, and response to antidepressant treatments

The relationship between peripheral serotonergic variables, melancholic traits, and clinical improvement after antidepressant treatment was examined in 83 drug-free major depressive patients. Plasma serotonin (5-HT) concentrations was lower in untreated melancholic patients (1.00 +/- 0.11 vs. 1.84 +/- 0.28 ng/mL, p < 0.008; N = 40 and 43, respectively). A tendency was observed for plasma 5-hydroxyindoleacetic acid (p < 0.06), whereas platelet 5-HT and plasma tryptophan did not differ between groups. After blood sampling and clinical ratings, treatment began with fixed doses of 5-HT uptake inhibitors (clomipramine or fluvoxamine), monoamine oxidase inhibitors, or tianeptine, a 5-HT uptake enhancer. There was no significant difference in response rates between patients with and without melancholic traits. The relationship between the clinical response at 6 weeks (>50% reduction of baseline Hamilton score) and the pretreatment values of biochemical variables was examined. Responders had a lower pretreatment platelet 5-HT (530 +/- 36 vs. 664 +/- 50 ng/10(9) platelets, p < 0.03; N = 44 and 39, respectively). Patients with a platelet 5-HT concentration above 800 ng/10(9) platelets had a lower response rate than those below this value (p < 0.003). This difference was maximal in the subgroup of patients treated with 5-HT uptake inhibitors (N = 49). In this subgroup, the response rates of patients with 5-HT concentrations below and above the cutoff point were, respectively, 70% and 17% (p < 0.001). A pretreatment platelet 5-HT value above 800 ng/10(9) platelets had a predictive value for a negative response of 92%. These results suggest the presence of biochemical differences in the peripheral serotonergic system between melancholic and nonmelancholic patients. The inverse relationship between the pretreatment platelet 5-HT content and clinical response may be useful in the investigation of the relationship between the 5-HT system and antidepressant response.

Adaptative Changes of the Serotonergic System After Antidepressant Treatments

The search for an adequate treatment of major depression is one of the main challenges of Neuropharmacology. To date, no single drug is effective in all patients treated, probably due to the heterogeneity of the disease and to individual differences in the response to the agents used. Despite provoking pharmacological effects when given at once, all treatments need to be administered for several weeks before amelioration signs begin to emerge. This has led to the notion that —rather than the inhibition of uptake or monoamine oxidase (MAO) activity per se— adaptative changes of aminergic receptors mediate the clinical effects.

Two actions are better than one : avoiding self-inhibition of serotonergic neurones enhances the effects of serotonin uptake inhibitors

The serotonin (5-HT)-increasing action of 5-HT uptake or monoamine oxidase inhibitors is limited by a negative feedback at somatodendritic level. The excess 5-HT produced by these antidepressant drugs in the interstitial space of the midbrain raphe activates somatodendritic 5-HT1A autoreceptors, thereby attenuating terminal 5-HT release. This effect is maximal in forebrain areas innervated by the dorsal raphe nucleus and can be prevented by the administration of non-selective [(-)pindolol, (-)tertatolol] and selective (WAY-100635) 5-HT1A antagonists. In keeping with these observations, the combined administration of selective serotonin reputake inhibitors (SSRIs) and 5-HT1A antagonists increase the cortical and striatal extracellular 5-HT concentration more than the former alone. Also, concurrent inhibition of the 5-HT and noradrenaline transporters with 20 mg/kg imipramine increases cortical extracellular 5-HT concentration more than SSRI doses which maximally block the 5-HT transporter. Moreover, the effects of fluoxetine on frontal cortex 5-HT are potentiated by a dose of desipramine that does not modify extracellular 5-HT by itself. Given the relevance of increased serotonergic transmission in the treatment of depression, these experimental data indicate that dual-action antidepressant treatments may be more effective than those which selectively inhibit the 5-HT transporter.

In vivo evidence for the reversible action of the monoamine oxidase inhibitor brofaromine on 5-hydroxytryptamine release in rat brain.

We have used intracerebral microdialysis to examine the reversibility of the action of brofaromine, a selective inhibitor of monoamine oxidase-A (MAO, E.C. 1.4.3.4.), on 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) output in rat frontal cortex. Brofaromine significantly increased the 5-HT output to about 200% of basal values 4 h after the s.c. administration of 10 and 30 mg/kg (but not 3 mg/kg) and reduced the concentration of 5-HIAA in the dialysate dose-dependently (61%, 53% and 41% of basal value with doses of 3, 10 and 30 mg/kg, respectively). At this time, cortical 5-HT concentration was increased and cortical 5-HIAA concentration was decreased in a dose-dependent manner.Treatment of rats with 10 mg/kg brofaromine plus 2.5 mg/kg of the irreversible MAO-B inhibitor l-deprenyl increased the concentration of 5-HT in the dialysate more than did brofaromine alone (503% vs 206% of the basal value, 4 h after administration). Similarly, clorgyline (5 mg/kg) plus l-deprenyl (2.5 mg/kg) increased the concentration of 5-HT in the dialysate to 461 % of the control value. This indicates that the concurrent inhibition of both types of MAO increases 5-HT output more than the selective blockade of either enzyme subtype. We have used this characteristic to examine, in vivo, the reversibility of the interaction of brofaromine with MAO-A. The output of 5-HT and 5-HIAA was examined 19–21 h after treatment with l-deprenyl plus clorgyline or l-deprenyl plus brofaromine. At this time, the concentration of 5-HT in the dialysate was increased six-fold in the clorgyline plus l-deprenyl group, as compared to pre-treatment values, but did not differ significantly from these in the brofaromine plus L-deprenyl group. Also, 5-HIAA concentration in the dialyste was still reduced (48% of basal value) in the clorgyline plus l-deprenyl group but not in the brofaromine plus l-deprenyl group. Cortical 5-HT and 5-HIAA concentrations in these animals, measured at the end of the microdialysis experiments (21 h after treatment), displayed changes that paralleled those in the extracellular compartment.These results indicate that: (a) as with clorgyline, the inhibition of MAO-A with brofaromine has a more pronounced effect on tissue 5-HT concentrations than on extracellular 5-HT concentrations; (b) the simultaneous inhibition of both forms of MAO with l-deprenyl and either brofaromine or clorgyline increased the concentration of 5-HT in the dialysate shortly (1–4 h) after administration more than did treatment with either of the MAO-A inhibitors alone; (c) dialysate and tissue 5-HT concentrations were greatly increased one day after the irreversible inhibition of MAO by clorgyline plus l-deprenyl, whereas they had returned to pretreatment values in those animals treated with brofaromine plus l-deprenyl. These transient effects on 5-HT and 5-HIAA in vivo provide further support for the conclusion that the interaction of brofaromine with brain MAO-A is reversible.