Regulation of central corticosteroid receptors following short-term activation of serotonin transmission by 5-hydroxy-L-tryptophan or fluoxetine.

Alterations of the hypothalamic-pituitary-adrenal (HPA) axis function characterized by a decreased negative feedback capacity are often associated with affective disorders and are corrected by treatment with antidepressant drugs. To gain a better understanding of the effects of the antidepressant drug fluoxetine, a specific serotonin (5-HT) reuptake inhibitor, on central corticosteroid receptors, the effects of short-term activation of serotonin transmission on central corticosteroid receptor expression were analysed in adrenalectomized (ADX) rats either supplemented or not with corticosterone. Serotonin transmission was stimulated either by a single injection of the 5-HT precursor, 5-hydroxy-L-tryptophan (5-HTP), or by a 2-day treatment with fluoxetine. In ADX rats, administration of 5-HTP decreased hippocampal mineralocorticoid (MR) and glucocorticoid (GR) receptor numbers 24 h later, while their respective mRNAs were unchanged and these effects of 5-HTP were mediated by 5-HT2 receptors. In the hypothalamus, GR mRNAs and binding sites decreased 3 h and 24 h after 5-HTP, respectively. By contrast, fluoxetine treatment increased hippocampal MR and GR mRNAs and MR binding sites while GR number remained unchanged. In ADX rats supplemented with corticosterone, 5-HTP and fluoxetine treatment had the same effects on corticosteroid receptors compared to those observed in non supplemented ADX rats: 5-HTP decreased hippocampal MR and GR and hypothalamic GR while fluoxetine treatment increased hippocampal MR. These results show that short-term stimulation of 5-HT transmission by 5-HTP decreases hippocampal and hypothalamic corticosteroid receptor numbers through a corticosterone-independent mechanism. It is hypothesized that the delayed maximal increase in extracellular 5-HT contents after fluoxetine treatment, due to negative feedback regulations induced by the activation of 5-HT1A and 5-HT1B autoreceptors, is not the primary cause for the delayed normalization of corticosteroid receptor numbers that regulates the HPA axis functioning.

Central 5-HT(1) and 5-HT(2) binding sites in transgenic mice with reduced glucocorticoid receptor number.

Transgenic mice bearing a transgene coding for a glucocorticoid receptor antisense mRNA, which partially blocks glucocorticoid receptor expression, were used in order to clarify the role of glucocorticoid receptors in the regulation of 5-HT(1A), 5-HT(1nonA) and 5-HT(2) binding sites labelled by quantitative autoradiography in the frontal and prefrontal cortex, striatum, hypothalamus, amygdala and raphe nuclei. We found that 1 nM [3H]8-hydroxy-2-[di-N-propylamino]tetralin ([3H]8-OH-DPAT) binding to 5-HT(1A) sites was decreased in strata oriens (-15.1+/-3.5%) and radiatum-lacunosum-moleculare (-13.3+/-4.3%) of the hippocampal CA(3) area, and 2 nM [3H]5-hydroxytryptamine binding to 5-HT(1nonA) sites in the presence of 100 nM 8-OH-DPAT and mesulergine was decreased in the dorsal subiculum (-17.8+/-6.9%). By contrast, 5-HT(2) sites labelled by 0.5 nM of (+/-)-1-(2, 5-dimethoxy-4-[125I]iodophenyl)-2-aminopropane was increased in the dorsal subiculum (+35.2+/-11.5%) and CA(2) area (+29.2+/-11.3%). The observed differences in binding to 5-HT(1) and 5-HT(2) sites were all located in areas of the hippocampus that contain both gluco- and mineralo-corticoid receptors, and no difference was observed in anatomical structures which contain only glucocorticoid receptors. Therefore, it seems that the important factor for the regulation of these 5-HT receptors is the interaction between gluco- and mineralo-corticoid receptors rather than the absolute density of glucocorticoid receptors. These results suggest that some of the alterations of the serotonergic neurotransmission observed in depressed patients might be secondary to an altered glucocorticoid receptor function.

The effects of serotonin on glucocorticoid receptor binding in rat raphe nuclei and hippocampal cells in culture.

The raphe-hippocampal serotonin (5-HT) system is involved in the regulation of the hypothalamus-pituitary-adrenal axis. The purpose of this study was to determine and compare the roles of 5-HT in the regulation of glucocorticoid receptor (GR) binding in the raphe nuclei and in the hippocampus. The effects of 5-HT, 5-HT agonists, and the 5-HT reuptake inhibitor citalopram on GR binding sites were studied in primary cultures of the fetal raphe nuclei and the hippocampus. Exposure of hippocampal cells to 5-HT, (+/-)-2,5-dimethoxy-4-iodoamphetamine (DOI; a 5-HT2 agonist), or citalopram resulted in an increase in number of GR binding sites. The effect of DOI was blocked by ketanserin (a 5-HT2 antagonist). Specific and saturable GR binding was found in raphe cells. Exposure of raphe cells to 5-HT, (+/-)-8 hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT; a 5-HT1A agonist), or citalopram induced a significant decrease in number of GR binding sites. The effect of 8-OH-DPAT was reversed by WAY 100135 [N-tert-butyl-3-[1-[1-(2-methoxy)phenyl]piperazinyl]-1-phenylpropiona mide; a 5-HT1A antagonist]. These results show that the regulation of GRs during fetal life is structure-dependent and involves different 5-HT receptor subtypes. Moreover, the regulation of hippocampal GRs by citalopram suggests an action of antidepressants independent of their effects on monoamines.

Effects of bilateral olfactory bulbectomy on the anterior pituitary corticotropic cell activity in male rats.

Bilateral olfactory bulbectomy (OB) has drastic biochemical and behavioral effects and is often associated with an increase in plasma corticosterone concentrations. This experiment examined the effects of OB on adrenocorticotropin (ACTH) and corticosterone release under basal and stress conditions and on proopiomelanocortin (POMC) gene expression. Bulbectomy potentiated hypophysal ACTH and adrenal corticosterone release induced by ether stress but had no effect on ACTH release under basal conditions, despite a significant increase of circulating corticosterone. POMC gene expression was stronger (+60%) in OB rats than in sham-operated rats. These results suggest that olfactory bulbectomy substantially altered the negative feed-back exerted by glucocorticoids on anterior pituitary corticotropic cells in the male rat.

An investigation of serotonergic involvement in the regulation of ACTH and corticosterone in the olfactory bulbectomized rat.

The bilateral olfactory bulbectomy resulted in significantly higher plasma concentration of corticosterone, but not of ACTH in basal conditions and much higher plasma ACTH and corticosterone concentrations after 15 min of immobilization stress than were observed in sham-operated animals. Daily treatment with fluoxetine-a specific serotonin reuptake inhibitor-(15 mg/kg/day) had no effect on basal ACTH and corticosterone concentrations in OB rats. Fluoxetine treatment caused lower levels of ACTH, but not of corticosterone secretion, in response to immobilization stress. Bulbectomy significantly reducing 5-HT concentration in the amygdala. Stress increased serotonergic activity in the hypothalamus but not in the amygdala of OB rats. Chronic fluoxetine treatment of both unstressed and stressed OB rats resulted in a lower turnover rate in the two structures. Our results suggest that the hypercorticosteronemia observed after bulbectomy in unstressed OB rats is independent of the serotonergic system in both hypothalamus and amygdala. In contrast, they also demonstrate hypothalamic 5-HT changes in the HPA hyperactivity of OB rats in response to stress. Chronic fluoxetine treatment may normalize pituitary ACTH secretion in response to stress, possibly desensitization of the 5-HT receptors in the hypothalamus due to 5-HT being move available at the synapses.

Indirect evidence for an association of 5-HT(1B) binding sites with retinal and geniculate axon terminals in the rat suprachiasmatic nucleus.

The purpose of the present study was to investigate the possible cellular location of 5-HT(1B) receptors on retinal and geniculate afferents in the rat suprachiasmatic nucleus (SCN). Biocular enucleation significantly decreased 5-HT(1B) binding site labeling (35%), specifically in the ventral part of the SCN, while monocular enucleation produced a decrease of smaller magnitude (12%), limited to the ventral part of the contralateral SCN, these results being consistent with the known distribution of retinal afferents in the nucleus. By contrast, bilateral geniculate lesion did not induce any significant variation of 5-HT(1B) binding site labeling in the SCN. Previously, we reported that serotonin (5-HT) synthesis inhibition by parachlorophenylalanine increases 5-HT(1B) binding site labeling in the SCN. Using saturation studies, we have now demonstrated that this upregulation reflected an increase in the total number of 5-HT(1B) binding sites (+41% in the dorsal and +67% in the ventral part of the SCN). Furthermore, we evaluated the effects of bilateral geniculate lesion after 5-HT stores depletion in order to overcome problems of technical resolution limits. The magnitude of upregulation was significantly decreased (27%) after bilateral geniculate lesion, suggesting that part of the 5-HT(1B) receptor population was located on geniculate axon terminals within the SCN. The possible involvement of 5-HT(1B) receptors, according to their cellular locations evidenced in the present study, in photic and nonphotic entrainment of the circadian clock is discussed.

Identification and role of serotonin 5-HT1A and 5-HT1B receptors in primary cultures of rat embryonic rostral raphe nucleus neurons.

Autoregulatory mechanisms affecting serotonin [5-hydroxytryptamine (5-HT)] release and synthesis during the early period of development were investigated in dissociated cell cultures raised from embryonic rostral rat rhombencephalon. The presence of 5-HT1A and 5-HT1B receptors in serotoninergic neurons was assessed using binding assays. The involvement of 5-HT1A and 5-HT1B receptors in the control of the synthesis and release of [3H]5-HT was studied using biochemical approaches with several serotoninergic receptor ligands. A mean decrease of 30% in [3H]5-HT synthesis and release was observed in the presence of 5-HT (10(-8) M), the 5-HT1A agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), the 5HT1B/1A agonist 5-methoxy-3-(1,2,5,6-tetrahydro-4-pyridinyl)-1H-indole (RU 24969), the 5-HT1B agonist 3-(1,2,5,6-tetrahydropyrid-4-yl)pyrrolo[3,2-b]pyrid-5-one (CP-93,129), and the 5-HT(1D/1B) agonist sumatriptan. Inhibition of 5-HT synthesis and release induced by 8-OH-DPAT was blocked by chiral N-tert-butyl-3-[1-[1-(2-methoxy)phenyl]piperazinyl]-1-phenylpropionam ide dihydrochloride quaternary-hydrate (WAY 100135) (10(7) M) or methyl 4-[4-[4-(1,1,3-trioxo-2H-1,2-benzoisothiazol-2-yl)butyl]-1-p iperazinyl]-1Hindole-2-carboxylate (SDZ 216-525) (10(-7)M), and that of CP-93,129 was blocked by methiothepin (10(-7) M). Paradoxically, extracellular levels of [3H]5-HT increased in the presence of 8-OH-DPAT and RU 24969 at 10(-6) M. 5-HT uptake experiments showed that these two agonists interacted with the 5-HT transporter. 5-HT1 binding sites (620 fmol/mg of protein) and 5-HT1A (482 fmol/mg of protein) and 5-HT1B (127 fmol/mg of protein) receptors were detected in 12-day in vitro cell cultures. Experiments carried out with tetrodotoxin suggested that 5-HT1A receptors are located on nerve cell bodies, whereas 5-HT1B receptors are located on the nerve terminals. We concluded that autoregulatory mechanisms involving 5-HT1A and 5-HT1B autoreceptors are functionally mature in cells from rostral raphe nuclei during the early period of development.

Olfactory bulbectomy increases vasopressin, but not corticotropin-releasing hormone, content in the external layer of the median eminence of male rats.

Removal of the olfactory bulbs results in numerous physiological and behavioral changes in rats. The most frequent and characteristic change is an abnormally high level of corticosterone in the blood, possibly due to changes in the activity of the hypothalamic neurons which synthesize corticotrophin-releasing hormone (CRH). Some of these neurons also synthesize vasopressin (AVP). They are located in the parvocellular part of the paraventricular nucleus of the hypothalamus, which projects into the external layer of the median eminence. We investigated whether there was such a change in activity by studying the synthesis and storage activity of CRH neurons in bulbectomized rats. CRH and AVP axon terminals in frozen sections of the external layer of the median eminence were labeled by immunofluorescence techniques and the degree of labeling was analyzed semi quantitatively. There was no difference in the area or intensity of CRH-labeling in control and bulbectomized rats. However, a significantly larger area was stained for AVP in the bulbectomized than in control rats. We also used in situ hybridization, with single- and double-labeling, to study the effects of bulbectomy on expression of the genes encoding CRH and AVP. No significant difference was found in the levels of mRNA for CRH and the number of CRH+/AVP+ cell bodies was similar in the parvocellular part of the paraventricular nucleus in bulbectomized and normal rats. Our results suggest that the hypothalamo-pituitary-adrenal (HPA) axis changes observed after olfactory bulbectomy may be due to plastic changes in hypothalamic CRH neurons, resulting in greater storage of increased AVP in CRH neurosecretory nerve terminals in the external layer of the median eminence.

Effect of serotonin inhibition on glucocorticoid and mineralocorticoid expression in various brain structures.

Many studies have shown the existence of functional interactions between central neurotransmitter systems and the hypothalamo-pituitary adrenal axis. Mineralocorticoid receptors (MR) and glucocorticoid receptors (GR) are regulated by multiple factors including glucocorticoids themselves. Neurotransmitters such as serotonin (5-hydroxytryptamine: 5-HT) can regulate brain corticosteroid receptors in a complex way. The present study examined the short-term (48 h) effects of parachlorophenylalanine (PCPA), a drug which specifically inhibits 5-HT synthesis, on corticosteroid receptor levels and on the expression of their respective messenger ribonucleic acids (mRNA) in the rat hippocampus, hypothalamus and brain stem. The study was performed in bilaterally adrenalectomized animals, in order to avoid potential drug-induced changes in plasma corticosterone levels, which could secondarily regulate MR and GR. Short-term inhibition of 5-HT synthesis by PCPA significantly increased the number of hippocampal MR-binding sites. PCPA treatment did not alter the number of GR-binding sites in the hippocampus, hypothalamus and brain stem. We observed no change in the affinities of GR and MR sites in all the structures studied. In PCPA-treated rats, restoration of control 5-HT levels by injection of its immediate precursor, 5-hydroxytryptophan (5-HTP) brings the number of hippocampal MR-binding sites back to control levels. It can therefore be concluded that the increase in number of MR-binding sites induced by acute PCPA treatment is dependent on the decrease in 5-HT levels. The increase in hippocampal MR binding sites was correlated with an induction of their messengers, suggesting that 5-HT modulates the synthesis of MR protein. Although PCPA did not modify the number of hippocampal GR-binding sites, a decrease in hippocampal GR mRNA expression was observed. This study shows that 5-HT inhibits hippocampal mineralocorticoid receptor synthesis and that this effect is not mediated by changes in corticosterone hormone secretion, and illustrates the existence of complex mechanisms for corticosteroid receptor regulation in the hippocampus.

Regional serotonin metabolism under basal and restraint stress conditions in the brain of transgenic mice with impaired glucocorticoid receptor function.

Transgenic (TG) mice deficient in glucocorticoid receptors (GR) were used in order to study the effects of a reduced GR function on adrenocorticotropin hormone and corticosterone plasma levels and on serotonin metabolism in different brain areas under basal resting conditions, after a 30-min restraint stress and 60 min after the end of the restraint stress. There was no difference in basal or stress-induced levels of either adrenocorticotropin hormone or corticosterone in control and TG mice, but the return of adrenocorticotropin hormone to basal values after the end of the stress was delayed in TG mice. Under basal conditions, the ratio 5-hydroxyindoleacetic acid/5-hydroxytryptamine was decreased only in the hippocampus of TG mice compared to controls. In the brain stem, the ratio 5-hydroxyindoleacetic acid/5-hydroxytryptamine increased compared to basal values after a 30-min restraint stress and values were still high 60 min after the end of the restraint stress in both control and TG mice. In the hippocampus, the ratio 5-hydroxyindoleacetic acid/5-hydroxytryptamine increased at the end of the stress and returned to basal levels 60 min later in control mice, whereas there was no change at the end of the stress but an increase 60 min later in TG mice. Finally there was no change in serotonin metabolism in the cortex, striatum or hypothalamus in either group or situation. Our results support the hypothesis of a tonic activation of serotonin turnover by corticosterone through GR in the mouse hippocampus. Moreover, stress-induced stimulation of serotonin metabolism in the brain stem and hippocampus appears to be delayed in TG mice compared to control mice. These results are particularly relevant for mood disorders such as depression where alterations of serotoninergic transmission might be secondary to an impairment of GR functions.

Post-lesion up-regulation of 5-HT1B binding sites in the suprachiasmatic nucleus may be reversed after spontaneous or graft-induced serotonin reinnervation.

We have previously reported that selective axotomy of serotoninergic neurons produced by an intraventricular injection of 5, 7-dihydroxytryptamine is followed by an increase in 5-HT1B binding sites in the suprachiasmatic nucleus of the hypothalamus. This post-lesion up-regulation is shown here to be spontaneously reversed after long-term survival in spite of an incomplete reinnervation of the nucleus. Recovery may be accelerated by fetal raphe transplants that produce more rapid reinnervation.

Effects of bilateral olfactory bulbectomy on circadian rhythms of ACTH, corticosterone, motor activity and body temperature in male rats.

Bilateral olfactory bulbectomy (BOX) has major biochemical and behavioral effects, and is one of the most widely investigated of animal models of depression. We studied the consequences of BOX in male rats, on the organization of endogenous circadian rhythms for ACTH, corticosterone (Cort), motor activity (MA) and body temperature (BT). Mean levels were increased for Cort and MA, whereas no significant changes were observed for ACTH and BT. Significantly higher plasma Cort morning values were evidenced in BOX than sham-operated animals. In addition, compared with the single prominent power spectrum for the 24 hours period of control rats, the BOX animals displayed substantially lower 24 hours spectral power for the MA and BT circadian rhythms. These alterations suggest that olfactory bulbectomy, by disruption of the afferences and efferences, induced drastic changes in the function of the endogenous clock or of its regulating systems. From this point of view, bulbectomized rats may therefore be a valuable model to studying the etiology of psychiatric disorders with rhythm disturbance.

Serotonin directly stimulates luteinizing hormone-releasing hormone release from GT1 cells via 5-HT7 receptors.

Luteinizing hormone-releasing hormone (LHRH release, which serves as the primary drive to the hypothalamic-pituitary gonadal axis, is controlled by many neuromediators. Serotonin has been implicated in this regulation. However, it is unclear whether the central effect of serotonin on LHRH secretion is exerted directly on LHRH neurosecretory neurons or indirectly via multisynaptic pathways. The present studies were undertaken in order to examine whether LHRH secretion from immortalized LHRH cell lines is directly regulated by serotonin and, if so, to identify the receptor subtype involved. 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), a 5-HT1A/7 receptor agonist, stimulated LHRH release from GT1-1 cells. This effect was blocked by ritanserin, a 5-HT2/7 receptor antagonist, but not by SDZ-216-525, a 5-HT1A antagonist. Basal LHRH release was not affected by the 5-HT2 agonist DOI. Reverse transcription and polymerase chain reaction technique (RT-PCR) was used in order to identify 5-HT1A and 5-HT7 receptor mRNA in immortalized LHRH cell lines. GT1-1 cells express mRNA for the 5-HT7, but not the 5-HT1A receptor subtypes. These results demonstrate a direct stimulatory effect of serotonin on LHRH release via 5-HT7 receptor.

Resolution of multiple affinity states of 5-hydroxytryptamine-1A receptors labelled by [3H]-8-hydroxy-2-(di-n-propilamino) tetralin in rat hippocampal membranes.

1. We examined the binding of [3H]-8-hydroxy-2-(DI-n-propilamino)tetralin ([3H]-8OH-DPAT) to 5-hydroxytriptamine-1A (5-HT1A) receptors in rat hippocampal membranes. 2. Computer analysis of [3H]-8OH-DPAT displacement curves in the absence and in the presence of 100 microM guanosine-5'-O-(3-thiotriphosphate) (GTP gamma S) were best fitted with a three-site model with apparent dissociation constants (Kd) of 0.45, 2.8 and 30 nM; the corresponding binding capacity (Bmax) existing in the three affinity states were 4, 2 and 12 pmol/g of tissue (wet weight), respectively. 3. These results suggest that [3H]-8OH-DPAT binding can be resolved as complex isotherms and we provided evidence that [3H]-8OH-DPAT labels 2 high-affinity GTP gamma S-sensitive and one low-affinity GTP gamma S-sensitive state of 5-HT1A receptors.

Adrenergic control of alpha-melanocyte-stimulating hormone release in frog pituitary is mediated by both beta- and a nonconventional alpha 2-subtype of adrenoreceptors.

Previous studies have shown that melanotrope cells of the pars intermedia of Rana ridibunda are inhibited by dopaminergic D2 agonists and stimulated by beta-adrenergic agonists. In the present study, we have examined the possible involvement of alpha-adrenoreceptors in the regulation of frog melanotrope cells. Reversed-phase HPLC analysis combined with electrochemical detection revealed the presence of both dopamine and noradrenaline in pars intermedia extracts (74.1 and 3.2 ng/mg protein, respectively), while adrenaline was undetectable. Administration of graded doses of noradrenaline and adrenaline (from 0.1 to 10 microM) to perifused frog neurointermediate lobes induced a dose-dependent inhibition of alpha-MSH release. The inhibitory effect of adrenaline was partially blocked by the D2-dopaminergic antagonist sulpiride and totally suppressed by concomitant administration of sulpiride and yohimbine (an alpha 2-adrenergic antagonist). Conversely, in the presence of sulpiride, noradrenaline provoked a strong stimulation of alpha-MSH secretion which was totally blocked by the beta-adrenergic antagonist propranolol. Taken together, our results indicate that endogenous catecholamines may exert a complex regulatory action on frog melanotrope cells through activation of dopaminergic D2, alpha 2- and beta-adrenergic receptors.

Stimulatory effects of 5HT1A receptor agonists on luteinizing hormone-releasing hormone release from cultured fetal rat hypothalamic cells: interactions with progesterone.

Previous works have suggested an interactive stimulatory effect of progesterone (P) and serotonin (5-HT) on luteinizing hormone release. The purpose of the present study was to determine whether 5-HT via 5-HT1A receptors interacts with P in the process of luteinizing hormone-releasing hormone (LHRH) release. Using fetal hypothalamic neurons in primary cell cultures the first goal of this study was to determine the effects of 5-HT1A receptor agonists on LHRH secretion. 8-Hydroxy-2 (di-n-propylamino) tetralin (8-OH-DPAT) or ipsapirone (10(-5) M) significantly stimulated LHRH release. Pharmacological studies have allowed to rule out the possible involvement of alpha 2- or beta-adrenoreceptors, or 5-HT uptake sites, in the stimulatory effect of 8-OH-DPAT on LHRH release, thus demonstrating the specific involvement of 5-HT1A receptors in the stimulation of LHRH release. The second goal was to test the ability of P to stimulate LHRH release from fetal hypothalamic neurons. P (10(-6) M) applied for 30 or 120 min significantly stimulated LHRH secretion. The maintenance of the stimulation of LHRH release by P after a cycloheximide treatment or by an impermeable analog of P, P-3-BSA, has suggested a nongenomic effect of P on LHRH release. The effects of a pretreatment of cells by P on 8-OH-DPAT-induced LHRH release were tested. While 10(-7) M P alone did not stimulate LHRH release, this concentration of steroid potentiated the LHRH response to 10(-5) M 8-OH-DPAT. These findings led to the conclusion that P acting at the level of the plasma membrane potentiates the stimulatory effect of 5-HT1A receptor agonists on LHRH release.

8-OH-DPAT regulates the amplitude and the phase of LH surge in ovariectomized steroid-primed rats.

Precise interactions between ovarian steroids and neurotransmitters are required for the secretion of phasic LH surge. Previous data suggested the existence of an interactive stimulatory effect of progesterone (P) and serotonin (5-HT) on LH release. In the present work the effects of 8-OH-DPAT, a selective 5-HT(1A) agonist, on phasic LH secretion were tested in ovariectomized rats implanted for 6 days with a pellet of 17 ß estradiol (OVX-E(2)) and in OVX-E(2) treated with progesterone (OVX-E(2)-P). Intraperitoneal injection of 8-OH-DPAT at 11.00 h in the morning of the expected LH surge had no effect on circadian plasma levels of LH in OVX-E(2) rats, whereas it induced a phase advance and an increase in LH surge in OVX-E(2)-P rats. Administration of the antiprogestin RU 38486 in OVX-E(2)-P rat, totally abolished the combined effects of P and 8-OH-DPAT on phasic LH release. SDZ 216-525, a specific 5-HT(1A) antagonist administered 60 min before 8-OH-DPAT, inhibited the stimulatory effect of the 5-HT(1A) agonist on the amplitude of LH surge. The present data suggest that progesterone is required for the regulation of phasic LH release by 5-HT(1A) agonists and that under this hormonal condition the activation of 5-HT(1A) receptors induces a phase advance and an increase in LH surge.

Impairment of serotoninergic transmission is followed by adaptive changes in 5HT1B binding sites in the rat suprachiasmatic nucleus.

Serotonin1B (5-HT1B) receptor binding in the suprachiasmatic nucleus (SCN) following impairment of serotoninergic transmission was studied by quantitative autoradiography. Serotonin (5-HT) denervation with 5,7-dihydroxytryptamine (5,7-DHT) caused a significant increase in the density of 5-HT1B receptors in both the ventral (62%) and dorsal (53%) parts of the SCN as early as 3 days after axotomy. The magnitude of this increase did not differ 3, 15 or 21 days post-lesion. An up-regulation of 5-HT1B receptors with similar magnitude was obtained in the two parts of the SCN after inhibition of 5-HT synthesis by chronic parachlorophenylalanine treatment. In this case, up-regulation was shown to be reversible after restoration of 5-HT synthesis with L-5-hydroxytryptophan. These results indicate that 5-HT1B receptor density in the SCN was inversely correlated with 5-HT levels. These plastic properties exhibited by 5-HT1B receptors in the SCN are discussed in relation to the mode of 5-HT transmission and possible localization of the receptors onto the main chemically defined cell populations of the nucleus.

Effect of tianeptine on the hypothalamic somatotropic axis in the conscious sheep.

The action of serotonin on growth hormone (GH) secretion is controversial because of interspecies differences and lack of specificity of serotoninergic drugs. Serotonin (5-HT) appears to inhibit GH release in the sheep and in man. We have investigated the site of action of tianeptine, a 5-HT uptake enhancer, in sheep since it is possible to collect hypophysial portal blood for the simultaneous determination of growth hormone-releasing hormone (GHRH) and somatostatin in this species under conscious, unstressed conditions. Tianeptine injection (10 mg/kg i.v.) resulted in a significant, immediate and short-lasting (30 min) increase in peripheral GH (+750%; P < 0.01) and hypophysial portal GHRH (+180%; P < 0.01). No change in the secretion of somatostatin was recorded during the same time. These data suggest that serotoninergic inputs are inhibitory to GH secretion. Tianeptine acts centrally to stimulate GH secretion in the sheep and its effect is mediated through changes in GHRH but not somatostatin release into hypophysial portal blood.

N-methyl-D-aspartic acid/glycine interactions on the control of 5-hydroxytryptamine release in raphe primary cultures.

Glutamic acid and glycine were quantified in cells and medium of cultured rostral rhombencephalic neurons derived from fetal rats. In the presence of 1 mM Mg2+, NMDA (50 microM) significantly stimulated (by 69%) release of newly synthesized 5-[3H]hydroxytryptamine ([3H]5-HT). D-2-Amino-5-phosphonopentanoate (AP-5; 50 microM) blocked the stimulatory effect of NMDA. AP-5 by itself inhibited [3H]5-HT release (by 25%), suggesting a tonic control of 5-HT by glutamate. In the absence of Mg2+, basal [3H]5-HT release was 60% higher as compared with release with Mg2+. AP-5 blocked the increased [3H]5-HT release observed without Mg2+, suggesting that this effect was due to the stimulation of NMDA receptors by endogenous glutamate. Glycine (100 microM) inhibited [3H]5-HT release in the absence of Mg2+. Strychnine (50 microM) blocked the inhibitory effect of glycine, indicating an action through strychnine-sensitive inhibitory glycine receptors. The [3H]5-HT release stimulated by NMDA was unaffected by glycine. In contrast, when tested in the presence of strychnine, glycine increased NMDA-evoked [3H]5-HT release (by 22%), and this effect was prevented by a selective antagonist of the NMDA-associated glycine receptor, 7-chlorokynurenate (100 microM). 7-Chlorokynurenate by itself induced a drastic decrease in [3H]5-HT release, indicating that under basal conditions these sites were stimulated by endogenous glycine. These results indicate that NMDA stimulated [3H]5-HT release in both the presence or absence of Mg2+. Use of selective antagonists allowed differentiation of a strychnine-sensitive glycine response (inhibition of [3H]5-HT release) from a 7-chlorokynurenate-sensitive response (potentiation of NMDA-evoked [3H]5-HT release).