Chronic effects of fluoxetine,a selective inhibitor of serotonin uptake,on neurotransmitter receptors

Summary Fluoxetine administration to rats at a dose of 10 mg/kg i.p. daily up to 12 or 24 days failed to change the concentration-dependent binding of [³H]WB4101, [³H]clonidme and [³H]dihydroalprenolol to ₁ ^–, ₂ ^– and-adrenergic receptors, respectively; [³H]quinuclidinyl benzilate to muscarinic receptors; [³H]pyrilamine to histamine H₁ receptors and [³H]naloxone to opiate receptors. Persistent and significant decreases in receptor number (B_max value) without changes in the dissociation constant (K_D value) of [³H]5-HT binding in cortical membranes were observed upon chronic treatment with fluoxetine administered either by intraperitoneal injection or incorporation in the diet, A detectable reduction of 5-HT₁ receptor number occurred after once-daily injections of fluoxetine at 10 mg/kg i.p. within 49 hours. After pretreatment for 3 days with p-chlorophenylalanine, an inhibitor of 5-HT synthesis, followed by repeated administration of fluoxetine, 5-HT₁ receptor numbers were higher than those of normal rats, suggesting a dependence on synaptic concentration of 5-HT for fluoxetine to affect a receptor down-regulation. These studies provide further evidence for the selectivity of fluoxetine as an inhibitor of 5-HT reuptake, resulting in a selective down-regulation of 5-HT₁ receptors in the cerebral cortex of rat brain.     

Different regulation of serotonin receptors following adrenal hormone imbalance in the rat hippocampus and hypothalamus

Summary Adrenal influence on serotonin (5-HT) transmission in the hippocampal and hypothalamic areas was studied in adrenalectomized rats receiving or not corticosterone replacement. After adrenalectomy, the 5-HT presynaptic receptors were desensitized both in hippocampus and hypothalamus: a significant increase in 5-HT 1 and 5-HT 2 receptor binding numbers took place in membranes from the hippocampus, but not in hypothalamus, while no changes in affinity of receptors to radioligands were observed in either brain area. Corticosterone treatment restored the adrenalectomy-impaired 5-HT autoreceptor sensitivity in hippocampus and hypothalamus and 5-HT density receptor sites in the hippocampus. Serotonin autoreceptor down-regulation following adrenalectomy may increase 5-HT release to maintain the constancy of serotonergic transmission in the brain and 5-HT modulated CRH-ACTH release to compensate the plasma corticosteroid drop. Corticosterone seems to display a distinct tonic control on serotonin transmission in both hippocampus and hypothalamus, the diversity being due to the different roles played by the hormone in these brain regions.     

Regulation of corticosteroid receptors in the rat brain: the role of serotonin and stress

It has been suggested that physiological resistance to repeated stress is associated with increased 5-hydroxytryptamine (5-HT) release in the dorsal hippocampus and that dysregulation of this neuroadaptation may be implicated in the psychopathology of depression. This study used 5,7-dihydroxytryptamine lesions to investigate the role of 5-HT projections to the hippocampus in physiological responses to repeated stress and putative changes in corticosteroid receptor immunoreactivity in the brain. Repeated exposure to elevated open platform stress (1 h/day) caused regionally selective changes in glucocorticoid and mineralocorticoid receptor immunoreactivity in the dorsal hippocampus that were not observed in ventral hippocampus, frontal cortex, hypothalamus or parietal cortex. Glucocorticoid receptor immunoreactivity in the dorsal hippocampus was decreased after 5 days but increased after 20 days of stress. Mineralocorticoid receptor immunoreactivity was increased after 5 or 10 days of stress. The increases in glucocorticoid and mineralocorticoid receptor immunoreactivity, evoked by repeated stress, were abolished by lesions of the principal 5-HT projections to the hippocampus. The lesions abolished the increased defecation observed in stressed animals, but had no effects on the plasma corticosterone response to the stressor or the habituation of this response observed following repeated stress. The experiments have revealed a dissociation in the regulation of corticosteroid receptor expression in the dorsal and ventral hippocampus by repeated stress and 5-HT. The data suggest that adaptation to inescapable stress is associated with regionally selective changes in corticosteroid receptor expression in dorsal hippocampus that are largely 5-HT-dependent, although these changes do not mediate habituation of the pituitary adrenocortical response to the stressor.     

Transitional states of central serotonin receptors in Parkinson's disease

Summary Crude membrane preparations from the frontal cortex of controls and parkinsonian patients were used to demonstrate affinity changes of the specific³H-5-hydroxytryptamine (5-HT) binding sites. Two such sites were noteable in controls, a finding consistent with earlier observations. In Parkinson's disease, both high- and low-affinity sites are significantly decreased. Additional experiments either with prolonged incubation times or pre-incubation with N-ethylmaleimide change the two affinities to a single high-affinity or low-affinity constant. The concept of transitional states of 5-HT receptors is discussed and seems to have important implications in the treatment of parkinsonism.     

Chronic fluoxetine treatment accelerates kindling epileptogenesis in mice independently of 5‐HT2A receptors

Patients with epilepsy often have mood disorders, and these are commonly treated with antidepressant drugs. Although these drugs are often successful in mitigating depressive symptoms, how they affect the epileptogenic processes has been little studied. Recent evidence has demonstrated that treatment with selective serotonin reuptake inhibitor (SSRI) antidepressant drugs adversely promotes epileptogenesis, which may be of great concern considering the number of patients exposed to these drugs. This study investigated 5‐HT_2A receptor signaling as a potential mechanism driving the pro‐epileptogenic effects of the prototypical SSRI fluoxetine. Male homozygous 5‐HT_2A receptor knockout mice or wild‐type littermates (n = 9‐14/group) were treated with continuous fluoxetine (10 mg kg^?1d^?1, sc) or vehicle and subjected to electrical kindling of the amygdala. Compared to vehicle, fluoxetine treatment accelerated kindling epileptogenesis (P < .001), but there was no effect of genotype (P = .75), or any treatment x genotype interaction observed (P = .90). Of interest, fluoxetine treatment increased afterdischarge thresholds in both genotypes (P = .007). We conclude that treatment with fluoxetine promotes epileptogenesis in mice, but this effect is not mediated by 5‐HT_2A receptors. This suggests that antidepressants may accelerate the onset of acquired epilepsy in patients who have experienced epileptogenic cerebral insults.     

Nerve growth factor and p75NGFR factor receptor mRNA change in rodent CNS following stress activation of the hypothalamo-pituitary-adrenocortical axis

The synthesis of nerve growth factor (NGF) by the hippocampus raises the possibility that NGF may play a role in the regulation of the hypothalamic-pituitary-adrenal axis (HPAA). Subchronic cold stress has been shown to activate the HPAA in a mild noninvasive manner, to stimulate serum glucocorticoid levels, and to perturb NGF binding in hippocampus and basal forebrain. One or repeated episodes of cold stress increased NGF mRNA levels in the hippocampus and p75^NGFR mRNA levels in the basal forebrain. These changes were not due to elevated serum glucocorticoid levels since treatment with exogenous corticosterone had no effect on NGF and p75^NGFR mRNA levels. Adrenalectomy did not prevent the stress induced increases in NGF and p75^NGFR mRNA. © 1993 Wiley-Liss, Inc.     

Age-dependent loss of corticosterone modulation of central serotonin 5-HT1A receptor binding sites

A loss of endocrine and neurotransmitter system interactions, including corticosterone regulation of 5-HT_1Areceptors, may underlie the age-related deficits in the hypothalamic-pituitary-adrenal (HPA) axis including adapting to stress. In this study, female Fischer 344 rats, (ages 3, 13, and 18 months), were bilaterally adrenalectomized and supplemented for 3 weeks with placebo or corticosterone (200 mg or 600 mg) containing 21 day sustained-release pellets implanted subcutaneously (LC, MC, or HC, respectively). Scatchard analysis using the 5-HT_1A receptor agonist [³H]8-hydroxy-2-(di-N-propylamino) tetralin (8-OH-DPAT) demonstrated a significant decrease in hippocampal receptor density in 3 month and 13 month MC groups (-35.2 and -32.1%, respectively) as compared to age-matched LC groups; a significant decline in 5-HT_1A receptor density in 3 month and 13 month HC groups was found compared to age-matched MC groups (−16.7 and −22.0%, respectively). However, these hormone treatments (LC or HC) failed to alter hippocampal 5-HT_1A binding site density in the 18 month groups. Cortical 5-HT_1A receptor densities were altered in a similar age-dependent manner. In contrast, the density of hypothalamic 5-HT_1A receptors in the 18 month LC group was significantly increased above that in the 3 month LC group. An additional indicator of the hippocampal response to corticosterone, the distribution of glial fibrillary acidic protein (GFAP), revealed an age-related decline in responsiveness to hormone treatment in the oldest group. The present study has identified an age-associated deficit in the regulation of hippocampal 5-HT_1A receptors by corticosterone which may underlie the diminished capacity of the aging HPA axis to cope with stress. J. Neurosci. Res. 53:86–98, 1998. © 1998 Wiley-Liss, Inc.     

氟西汀对癫痫合并抑郁大鼠模型海马自噬的作用

目的 探讨氟西汀对癫痫合并抑郁大鼠海马齿状回自噬的影响。方法 将60只SD大鼠随机分为对照组、模型组、氟西汀组、3-甲基腺嘌呤（3-MA）组。采用体重、摄食量、旷场试验评定大鼠抑郁水平;采用免疫组化测定大鼠海马齿状回beclin1、LC3-I、mToR蛋白表达水平,荧光实时定量RT-PCR测定大鼠海马齿状回beclin1、LC3-I、mToR基因表达水平。结果 药物干预后,模型组体重、摄食量、垂直运动次数和水平运动次数明显低于对照组（P<0.01）;氟西汀组、3-MA组经药物治疗后以上指标高于模型组（P<0.01,P<0.05）。模型组海马齿状回beclin1、LC3-I表达显著升高,mToR表达下降,与对照组相比有统计学意义（P<0.01）;氟西汀组、3-MA组大鼠海马齿状回beclin1、LC3-I表达下降,mToR表达升高,与模型组相比差异有统计学意义（P<0.01）。结论 氟西汀可能通过改善癫痫合并抑郁大鼠海马齿状回区beclin1、LC3-I、mToR表达,抑制细胞自噬。     

Sympathetic denervation and chronic serotonin uptake blockade by fluoxetine do not affect pineal gland 5-hydroxyindole acetic acid: evidence that oxidative deamination of pineal serotonin is a property of the pinealocyte

Summary This investigation tested the hypothesis that oxidative deamination of 5HT in the pineal gland occurs primarily in cellular compartments other than the pinealocyte (i.e., noradrenergic nerve terminals and glia). Following sympathetic denervation of the pineal gland by bilateral superior cervical sympathectomy, pineal levels of 5HT and its oxidative metabolite, 5HIAA, were measured by HPLC from samples collected at six time points over the 24 h photoperiod. The role of glia in 5HT deamination was further examined by chronic treatment with the 5HT uptake blocker, fluoxetine (10 mg/kg). Sympathectomy abolished the circadian rhythms of both 5HT and 5HIAA, but had no statistically significant effect on the ratio of 5HIAA/5HT compared to shamoperated and intact controls over the 24 h period. Pineal daytime levels of both 5HT and 5HIAA were unaffected by fluoxetine treatment. These findings indicate that the pinealocyte is an important cellular compartment for 5HT oxidative metabolism.     

Endogenous 5-HT, released by MDMA through serotonin transporter- and secretory vesicle-dependent mechanisms, reduces hippocampal excitatory synaptic transmission by preferential activation of 5-HT1B receptors located on CA1 pyramidal neurons

A multitude of different serotonin (5-HT) receptor types are expressed in the hippocampus, but the identity of receptors actually mediating the physiological response to endogenous 5-HT has not been determined. We combined pharmacologically induced release of 5-HT with patch-clamp recordings on disinhibited rat CA1 minislices to determine effects of endogenous 5-HT on the excitability of pyramidal neurons and synaptic transmission among them. We found that application of 5-HT releasers, 3,4-methylenedioxy-methamphetamine (MDMA) or p-methylthioamphetamine, at concentrations ranging from 2 to 50 microm, reduced the excitatory synaptic transmission between CA1 pyramidal neurons without altering their basal electrical properties. This effect of MDMA was blocked by the selective 5-HT1B antagonist GR 55562, was dependent on endogenous 5-HT content and was mediated by presynaptically located, pertussis-toxin sensitive mechanisms. We found no other MDMA effects in our preparation, which indicates that the release of endogenous 5-HT preferentially stimulates 5-HT1B receptors on CA1 pyramidal neurons. Therefore, 5-HT1B receptor activation may represent a predominant component of the physiological response to endogenous 5-HT in the CA1. The high sensitivity of the 5-HT1B receptor-mediated reduction of polysynaptic excitatory responses to the extracellular 5-HT level enabled us to study mechanisms of the 5-HT releasing action of MDMA. Block of the serotonin transporter (SERT) with citalopram slowed the time course and reduced overall 5-HT release by MDMA. Depletion of vesicular 5-HT, by inhibition of vesicular monoamine transporter type 2 with tetrabenazine prevented the release. Thus although the SERT reversal contributes, a direct vesicle-depleting action is essential for MDMA release of 5-HT.     

Hypersensitivity to central serotonin receptor activation in scrapie-infected hamsters and the effect of serotonergic drugs on scrapie symptoms

Low doses of the serotonin agonist, quizapine, and the serotonin precursor,l-5-hydroxytryptophan methyl ester, produce small but significant reductions in scrapie-induced ataxia and action jerks in hamsters.Higher doses of both drugs elicit a behavioral syndrome specific for central serotonin receptor activation. Scrapie-infected hamsters show a dramatic hypersensitivity to both drugs compared to control animals. This suggests that scrapie infection in hamsters causes a disturbance in the serotonergic pathway of the brain stem.     

The Effect of Corticosterone on Reactivity to Spatial Novelty is Mediated by Central Mineralocorticosteroid Receptors

Corticosterone, secreted by the adrenal glands, binds to central mineralocorticoid receptors with high affinity and to glucocorticoid receptors with a tenfold lower affinity. In previous studies we have shown that the selective activation of either mineralocorticoid receptors or glucocorticoid receptors exerts distinctly different behavioural effects. In this study we examined in particular the mineralocorticoid receptor-mediated effect of corticosterone on the control of the behavioural response of male Wistar rats to spatial novelty. This analysis was based on our observation that in adrenal-intact rats the presence of an object in the centre of an open field alters the time spent and distance walked in the centre compared to the peripheral area, i.e. the pattern of reactive locomotor activity is changed. Using this paradigm we found that 1 day after removal of the adrenals the rats increased their behavioural reactivity towards the object. Treatment of adrenalectomized rats with a low dose of corticosterone (50 μg/kg s.c.) 1 h prior to testing restored the behavioural reactivity to the level of sham-operated, intact rats. Surprisingly, a high dose of corticosterone (1000 μg/kg s.c.) also increased the rat's reactivity towards the object. The same high dose of corticosterone given to adrenal-intact rats also increased behavioural reactivity. Pretreatment of these rats with an intracerebroventricular injection of the selective mineralocorticoid receptor antagonist RU28318 (100 ng/μl) prevented the corticosterone-induced increase in behavioural reactivity, while the blockade of glucocorticoid receptors with the antagonist RU38486 (100 ng/μl) was not effective. Administration of the mineralocorticoid receptor antagonist without corticosterone to adrenal-intact rats also increased behavioural reactivity, but this increase did not reach statistical significance. General locomotor activity was not affected by either treatment. In conclusion, we found a U-shaped relationship between the pattern of behavioural reactivity in a novel environment and the circulating plasma corticosterone level. The response to spatial novelty appeared to be sensitive with respect to the activation and blockade of central, presumably hippocampal mineralocorticoid receptors.     

Quantitative autoradiographic mapping of serotonin 5-HT1 and 5-HT2 receptors and uptake sites in the neocortex of the rhesus monkey

The in vitro autoradiographic technique was used to characterize the distribution of serotonin 5-HT1 and 5-HT2 receptors and uptake sites in 11 cortical areas of frontal, parietal, and occipital lobes in the rhesus monkey; 5-HT1 receptors were labeled with [3H]5-HT; 5-HT2 receptors were labeled with [3H]ketanserin; and 5-HT uptake sites were labeled with [3H]citalopram. Five-HT1 and 5-HT2 receptors and 5-HT uptake sites were found in every cortical area examined with the absolute concentration of 5-HT1 receptors higher than that of 5-HT2 receptors in all areas. In eight regions of prefrontal and parietal as well as in prestriate cortex, 5-HT1 and 5-HT2 receptors had complementary distribution profiles: 5-HT1 receptors were concentrated in layers I and II and the upper strata of layer III, while 5-HT2 receptors had their highest concentration throughout layers III and IV. Only the primary motor and visual cortex had receptor distributions different from that described above. Thus, in the primary visual cortex, both 5-HT1 and 5-HT2 receptors were found in high concentration in sublayer IVc beta, though the density of 5-HT1 receptor was also high in other subdivisions of layer IV and in layers III, V, and VI. In the primary motor cortex, both receptor subtypes were concentrated in layers I and II and the upper strata of layer III. The pattern of distribution of serotonin uptake sites did not match the patterns of distribution of either 5-HT1 or 5-HT2 receptors alone; rather it approximated the combined patterns of distribution of both receptor subtypes. The complementary patterns of distribution of 5-HT1 and 5-HT2 receptors in most areas of the monkey cerebral cortex suggest that these two receptor subtypes may make differential contributions to cortical functions.     

The role of hypothalamic serotonin (5HT) before ovulation in immature rats treated with pregnant mare serum (PMS).

(1) PCPA methyl ester (10 mg/rat i.p.) inhibits induced ovulation in immature rats treated with pregnant mare serum (PMS). It also suppresses the preovulatory surges of LH and FSH, but not those of oestradiol or progesterone. (2) There is an increase in hypothalamic 5HT levels in the aftermoon and hypothalamic 5HIAA levels in the evening of the two days studied (days 28 and 29 of life). This occurs whether or not PMS was given on day 27. (3) The antiovulatory effects of PCPA are only seen when it is given on the afternoon or evening of the day before the pre-ovulatory gonadotrophin surge, i.e. on day 28 over the period of raised hypothalamic 5HT metabolism. (4) PCPA reduces 5HT metabolism in the hypothalamus within 2 hr of administration and its anti-ovulatory effect can be overcome by 5-hydroxytryptophan. This indicates that hypothalamic 5HT activity is essential for the gonadotrophin surge. (5) The anti-ovulatory effect of PCPA can be overcome by progesterone, LH and FSH but not oestradiol.     

Serotonin 5- HT (2C) receptor knockout mice: autoradiographic analysis of multiple serotonin receptors.

Quantitative receptor autoradiography was used to study possible alterations of the densities of multiple serotonin (5-HT) receptor subtypes and of serotonin transporter in the brain of 5-HT(2C) receptor knockout mice. The radioligands employed were [(3)H]citalopram, [(3)H]WAY100,635, [(3)H]8-OH-DPAT, [(3)H]GR125743, [(3)H]sumatriptan, [(3)H]MDL100,907, [(125)I](+/-)DOI, [(3)H]mesulergine, [(3)H]5-HT, [(3)H]GR113808, and [(3)H]5-CT. As expected, radioligands that label 5-HT(2C) receptors showed a complete absence of labeling in mutant mice choroid plexus and significantly reduced densities in other brain regions expressing 5-HT(2C) receptors. With the rest of the radioligands, no significant alterations in the densities of labeled sites were found in any brain region. In situ hybridization showed no changes in 5-HT(2A) receptor and serotonin transporter mRNA levels, whereas 5-HT(2C) receptor mRNA levels were reduced in certain brain regions. The present results indicate that the mouse serotonergic system does not exhibit compensatory up- or down-regulation of the majority of its components (serotonin transporter and most 5-HT receptor subtypes) in response to the absence of 5-HT(2C) receptors.     

An immunocapture/scintillation proximity analysis of Gαq/11 activation by native serotonin (5‐HT)2A receptors in rat cortex: Blockade by clozapine and mirtazapine

Though transduction mechanisms recruited by heterologously expressed 5‐HT_2A receptors have been extensively studied, their interaction with specific subtypes of G‐protein remains to be directly evaluated in cerebral tissue. Herein, as shown by an immunocapture/scintillation proximity analysis, 5‐HT, the prototypical 5‐HT_2A agonist, DOI, and Ro60,0175 all enhanced [³⁵S]GTPγS binding to Gαq/11 in rat cortex with pEC₅₀ values of 6.22, 7.24 and 6.35, respectively. No activation of Go or Gs/olf was seen at equivalent concentrations of DOI. Stimulation of Gαq/11 by 5‐HT (30 μM) and DOI (30 μM) was abolished by the selective 5‐HT_2A vs. 5‐HT_2C/5‐HT_2B antagonists, ketanserin (pK_B values of 9.11 and 8.88, respectively) and MDL100,907 (9.82 and 9.68). By contrast, 5‐HT‐induced [³⁵S]GTPγS binding to Gαq/11 was only weakly inhibited by the preferential 5‐HT_2C receptor antagonists, RS102,221 (6.94) and SB242,084 (7.39), and the preferential 5‐HT_2B receptor antagonist, LY266,097 (6.66). The antipsychotic, clozapine, which had marked affinity for 5‐HT_2A receptors, blocked the recruitment of Gαq/11 by 5‐HT and DOI with pK_B values of 8.54 and 8.14, respectively. Its actions were mimicked by the “atypical” antidepressant and 5‐HT_2A receptor antagonist, mirtazapine, which likewise blocked 5‐HT and DOI‐induced Gαq/11 protein activation with pK_B values of 7.90 and 7.76, respectively. In conclusion, by use of an immunocapture/scintillation proximity strategy, this study shows that native 5‐HT_2A receptors in rat frontal cortex specifically recruit Gαq/11 and that this action is blocked by clozapine and mirtazapine. Quantification of 5‐HT_2A receptor‐mediated Gαq/11 activation in frontal cortex should prove instructive in characterizing the actions of diverse classes of psychotropic agent. Synapse 63:95–105, 2009. © 2008 Wiley‐Liss, Inc.     

Effects of short-term serotonin depletion on the efficacy of serotonin neurotransmission: electrophysiological studies in the rat central nervous system

The effects of short-term serotonin (5-HT) depletion by p-chlorophenylalanine (PCPA) on the firing activity of dorsal raphe nucleus 5-HT neurons, on the responsiveness of dorsal hippocampus pyramidal neurons to microiontophoretically applied 5-HT and on the efficacy of the electrical stimulation of the ascending 5-HT pathway in suppressing the firing activity of CA3 dorsal hippocampus pyramidal neurons were assessed in chloral hydrate-anesthetized rats. PCPA (250 mg/kg/day i.p. for 2 days) reduced the 5-HT content of the dorsal hippocampus by 90%. However, the number of spontaneously active 5-HT neurons per microelectrode trajectory through the dorsal raphe or their average rate of firing was unaltered. The effect of afferent 5-HT pathway stimulation was reduced in only 40% of treated rats, whereas the sensitivity of CA3 pyramidal neurons to microiontophoretic 5-HT was not modified. The function of the terminal 5-HT autoreceptor was assessed using methiothepin, an autoreceptor antagonist. Methiothepin (1 mg/kg, i.v.) significantly enhanced the efficacy of the stimulation in PCPA-treated rats, although the degree of enhancement was much less than in controls. A greater reduction of the effectiveness of the stimulation was obtained by increasing the dose of PCPA (350 mg/kg/day i.p. for 2 days). This regimen reduced the 5-HT content of the dorsal hippocampus by 95%. In these rats, the sensitivity of the terminal 5-HT autoreceptor was assessed by increasing the frequency of the stimulation from 1 to 5 Hz. This procedure reduced to a similar extent the efficacy of the stimulation in treated and control rats, suggesting that the reduced effectiveness of methiothepin in enhancing 5-HT synaptic transmission in PCPA-treated rats is due to a lower degree of activation of the terminal 5-HT autoreceptor. The present results showing that the 350 mg/kg/day regimen of PCPA, but not the 250 mg/kg/day regimen, reduced the efficacy of the stimulation of the ascending 5-HT pathway suggest that a greater than 90% depletion is required to affect 5-HT neurotransmission significantly. The reduced level of activation of terminal 5-HT autoreceptors in rats treated with the lower dose of PCPA may facilitate the release of the remaining 5-HT per stimulation-triggered action potential, ensuring a virtually unaltered synaptic efficacy.