Downregulated hypothalamic 5-HT3 receptor expression and enhanced 5-HT3 receptor antagonist-mediated improvement in fatigue-like behaviour in cholestatic rats

Abstract  The serotonin neurotransmitter system, including the 5-HT₃ receptor, has been implicated in the genesis of fatigue in patients with liver disease. Therefore, we examined the possible role of 5-HT₃ receptors in cholestasis-associated fatigue. Rats were either bile duct resected (BDR) or sham resected and studied 10 days postsurgery. A significant decrease in hypothalamic 5-HT₃ receptor expression was detected by immunohistochemistry and Western blot in BDR vs sham rats, coupled with increased hypothalamic serotonin turnover identified by an elevated 5-hydroxyindoleacetic acid (5-HIAA) to 5-HT ratio in BDR vs sham rats. To examine fatigue-like behaviour, an activity meter was used. BDR rats exhibited significantly lower locomotor activity than did sham animals. Subcutaneous injection of the 5-HT₃ receptor antagonist tropisetron (0.1 mg kg⁻¹) resulted in significantly increased locomotor activity in BDR rats compared to the activity in saline-treated controls, but was without effect in sham rats. However, a 10-fold higher dose of tropisetron significantly increased locomotor activity in both BDR and sham rats compared to saline-injected controls. These findings indicate that cholestasis in the rat is associated with increased hypothalamic serotonin turnover, decreased hypothalamic 5-HT₃ receptor expression, and enhanced sensitivity to locomotor activation induced by 5-HT₃ receptor antagonism, thereby implicating the 5-HT₃ receptor system in cholestasis associated fatigue.     

Increased binding at 5-HT(1A), 5-HT(1B), and 5-HT(2A) receptors and 5-HT transporters in diet-induced obese rats

5-Hydroxytryptamine (5-HT, serotonin), synthesized in midbrain raphe nuclei and released in various hypothalamic sites, decreases food intake but the specific 5-HT receptor subtypes involved are controversial. Here, we have studied changes in the regional density of binding to 5-HT receptors and transporters and the levels of tryptophan hydroxylase, in rats with obesity induced by feeding a palatable high-energy diet for 7 weeks. We mapped binding at 5-HT receptor subtypes and transporters using quantitative autoradiography and determined tryptophan hydroxylase protein levels by Western blotting. In diet-induced obese (DiO) rats, specific binding to 5-HT(1A) receptors ([3H]8-OH-DPAT) was significantly increased in the dorsal and median raphe by 90% (P<0.01) and 132% (P<0.05), respectively, compared with chow-fed controls. 5-HT(1B) receptor binding sites ([125I]cyanopindolol) were significantly increased in the hypothalamic arcuate nucleus (ARC) of DiO rats (58%; P<0.05), as were 5-HT(2A) receptor binding sites ([3H]ketanserin) in both the ARC (44%; P<0.05) and lateral hypothalamic area (LHA) (121%; P<0.05). However, binding to 5-HT(2C) receptors ([3H]mesulgergine) in DiO rats was not significantly different from that in controls in any hypothalamic region. Binding to 5-HT transporters ([3H]paroxetine) was significantly increased (P<0.05) in both dorsal and median raphe, paraventricular nuclei (PVN), ventromedial nuclei (VMH), anterior hypothalamic area (AHA) and LHA of DiO rats, by 47%-165%. Tryptophan hydroxylase protein levels in the raphe nuclei were not significantly different between controls and DiO rats. In conclusion, we have demonstrated regionally specific changes in binding to certain 5-HT receptor subtypes in obesity induced by voluntary overeating of a palatable diet. Overall, these changes are consistent with reduced 5-HT release and decreased activity of the 5-HT neurons. Reduction in the hypophagic action of 5-HT, possibly acting at 5-HT(1A), 5-HT(1B) and 5-HT(2A) receptors, may contribute to increased appetite in rats presented with highly palatable diet.     

5-HT1B receptor knockout mice show a compensatory reduction in 5-HT2C receptor function

Although null mutant ('knockout') mice have provided valuable animal models to complement traditional approaches to psychopharmacology, such animals may also show complex adaptations to the induced mutation. Here we demonstrate that serotonin1B (5-HT1B) receptor knockout (KO) mice show adaptations in serotonin2C (5-HT2C) receptor-mediated functions. They show smaller reductions in food intake and locomotor activity in response to administration of 5-HT2C receptor agonists that are not accounted for by altered drug disposition. These effects are not mimicked by pretreatment of wildtype (WT) mice with a 5-HT1B receptor antagonist showing that they result from a longer term adaptation to the loss of 5-HT1B receptor function and not from a short-term interaction between 5-HT1B- and 5-HT2C-mediated functions. In addition, we show that 5-HT1B receptor KO mice have a lowered hypothalamic c-fos response to the administration of 5-HT2C receptor agonists. These results demonstrate that compensatory adaptations to the constitutive loss of 5-HT1B receptors may be an important determinant of the altered response of 5-HT1B KO mice to a variety of pharmacological challenges.     

Modulatory role of 5-HT3 receptors in mediation of apomorphine-induced aggressive behaviour in male rats

We have studied the effects of serotonin (5-HT) 5-HT3 receptor agonists 1-phenylbiguanide (1-PBG) and 1-(m-chlorophenyl)biguanide (mCPBG), and antagonists 3-tropanyl-3,5-dichlorobenzoate (MDL-72222) and tropisetron (3-tropanyl-indole-3-carboxylate HCl; ICS-205930) on apomorphine-induced aggressive behaviour in normal or DSP-4 [N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine hydrochloride] pre-treated male Wistar rats. DSP-4 (50 mg/kg) pre-treatment significantly accelerated the development of apomorphine-induced aggressive behaviour. mCPBG (1.0 and 10 mg/kg) did not modify the aggressiveness, but 1-PBG (3.0 and 30 mg/kg) attenuated the aggressiveness in normal but not DSP-4 pre-treated rats. MDL-72222 (0.4 and 4.0 mg/kg) attenuated the aggressive behaviour in normal rats, tropisetron (0.3 mg/kg) had an antiaggressive effect only by citalopram (10 mg/kg) challenge. MDL-72222 and tropisetron were ineffective in DSP-4 pre-treated rats. In conclusion, our results indicate that the 5-HT3 receptors modulate the apomorphine-induced aggressive behaviour and the 5-HT3 receptor antagonists have moderate antiaggressive effect in this test.     

Study of mechanisms of calcitonin analgesia in mice. Involvement of 5-HT3 receptors.

The analgesic effect of calcitonin when serotonin (5-HT) concentration is increased and the involvement of some 5-HT receptors were studied using the writhing test in mice. 5-hydroxytryptophan (5-HTP) administration increased both 5-HT levels in the central nervous system (CNS) and calcitonin analgesia. The 5-HT(1A) agonist (+/-)-8-hydroxy-2-dipropylaminotetralin hydrobromide (8-OH-DPAT) diminished calcitonin analgesia, this effect being antagonised by the 5-HT(1A) antagonist (WAY 100, 135). As the stimulation of 5-HT(1A) autoreceptors reduces the turnover of 5-HT, the effect of 8-OH-DPAT on calcitonin analgesia may be attributed to this decrease. The 5-HT(2A-2C) agonist (+/-)-1-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane hydrochloride (DOI) diminished calcitonin analgesia. A sub-analgesic dose of the 5-HT(2A) antagonist ketanserin failed to prevent this effect. The 5-HT(3) agonist (+/-)-2-methyl-5-hydroxytryptamine maleate (2-methyl-5-HT) potentiated calcitonin analgesia, whereas it was significantly reduced by the 5-HT(3) antagonist tropisetron. The effect of 2-methyl-5-HT on calcitonin analgesia was also reversed by tropisetron, This result suggests that the 5-HT(3) receptor may play an important role in the relationship between calcitonin and the serotonergic system. Tropisetron also reversed the analgesia induced by calcitonin plus 5-HTP corroborating importance of the 5-HT(3) receptors.     

Intrastriatal serotonin 5-HT2 receptors mediate dopamine D1-induced hyperlocomotion in 6-hydroxydopamine-lesioned rats

Striatal dopamine (DA) and serotonin (5-HT) functions are altered following DA denervation. Previous research indicates that intrastriatal coadministration of D1 and 5-HT2 receptor agonists synergistically increase locomotor behavior in DA-depleted rats. In the present study, we examined whether striatal 5-HT2 mechanisms also account for supersensitive D1-mediated locomotor behavior following DA denervation. Adult male Sprague-Dawley rats were subjected to bilateral striatal cannulation and then received either intracerebroventricular (i.c.v.) or intrastriatal 6-hydroxydopamine (6-OHDA; 200 microg or 20 microg/side, respectively). After at least 3 weeks, i.c.v.-lesioned rats received intrastriatal infusions of the 5-HT2 receptor antagonist ritanserin (2.0 microg/side) or its vehicle (DMSO) followed by systemic SKF 82958, a D1 agonist (1.0 mg/kg, i.p.) and locomotor activity was monitored. In another experiment, intrastriatal sham and 6-OHDA-lesioned rats received bilateral intrastriatal infusions of ritanserin (2.0 microg/side) or its vehicle (DMSO) followed by intrastriatal infusions of SKF 82958 (5.0 microg/side) or vehicle (0.9% saline). Rats with DA loss demonstrated supersensitive locomotor responses to both systemic and intrastriatal SKF 82958. Ritanserin pretreatment blunted systemic SKF 82958-induced hyperlocomotion and returned intrastriatal D1-mediated hyperactivity to sham lesion levels. The results of this study suggest that striatal 5-HT2 receptors contribute to D1-mediated hyperkinesias resulting from DA loss and suggest a pharmacological target for the alleviation of dyskinesia that can develop with continued DA replacement therapy.     

5-HT precursor loading, but not 5-HT receptor agonists, increases motor function after spinal cord contusion in adult rats

Serotonergic (5-HT) receptors are upregulated following spinal cord transection. Stimulation by administration of serotonergic receptor agonists has been successful in improving hindlimb function. We tested whether this strategy would be successful in incomplete injury models (moderate or severe thoracic contusion) where descending projections are partially spared which should produce less denervation-induced receptor upregulation. Adult rats received midthoracic moderate (MOD: 25 mm drop) or severe (SEV: 50 mm drop) contusion injuries. Distribution of 5-HT and its transporter and expression of 5-HT_2C receptors were evaluated in lumbar spinal cord and motor response to 5-HT receptor activation was assessed using open field locomotion (BBB) score, percent weight supported treadmill stepping (%WS) and evaluation of hindlimb muscle activation (tremor and serotonin syndrome).5-HT immunostaining 3 months post-contusion revealed few 5-HT fibers caudal to the severe contusion, and more spared caudal to the moderate contusion. The distribution of 5-HT transporter paralleled 5-HT staining, but was more greatly reduced. Thus serotonin reuptake may be less efficient in the injured spinal cord. Immunostaining for the 5-HT_2C receptor in the dorsal and ventral horns at L5 showed significant upregulation in SEV, compared to sham or MOD rats.Neither 5-HT_2C nor 5-HT_1A receptor agonists, alone or in combination, nor the serotonin transporter inhibitor d-fenfluramine modified BBB scores or %WS in either group. Despite the increased sensitivity of post-synaptic targets, agonist treatment did not improve function in SEV rats. We conclude that selective 5-HT_2C or 5-HT_1A receptor activation was not effective in improving hindlimb function after incomplete lesions. In contrast, the 5-HT precursor 5-hydroxytryptophan (l-5-HTP), which leads to activation of all classes of 5-HT receptors, increased both %WS and hindlimb activity in the MOD group. While no side effects were observed in normal or MOD rats, SEV rats displayed hindlimb tremors and 33% mortality, indicating hypersensitivity to the precursor.     

Biochemical characterization of phosphoinositide hydrolysis stimulated by 5-HT3 receptor agonists.

Serotonin (5-HT) stimulates phosphoinositide (PI) turnover in rat fronto-cingulate cortical slices and is probably mediated through the activation of both 5-HT2 and 5-HT3 receptors. We have extended these findings and have assessed whether the increased stimulation of PI turnover is secondary to 5-HT stimulated arachidonate metabolism or to the release of another neurotransmitter. Incubation of the cortical slices by the two 5-HT3 receptor agonists, 2-methyl-serotonin (2-Me-5-HT) and phenylbiguanide (PBG), significantly decreases serotonin-stimulated phosphoinositide turnover, indicating that activation of 5-HT3, receptors by 2-Me-5-HT and PBG caused the desensitized PI hydrolysis to 5-HT. Indomethacin did not affect the increased PI hydrolysis induced by 5-HT, 2-Me-5-HT and PBG, suggesting that neither cyclooxygenase nor lipoxgenase activity is required for the PI response and that it is independent of arachidonic acid metabolism. The stimulation in PI turnover induced by 5-HT and the 5-HT3 receptor agonists was not potentiated by proteinase inhibitors suggesting that the release of a peptide neurotransmitter is not involved in the PI response. In addition, the effects of 5-HT, 2-Me-5-HT and PBG on PI turnover are additive to the effect of KCl and veratrine. In conclusion, our results indicate that the action of 2-Me-5-HT and PBG on PI turnover is direct.     

5-HT(3) receptor mediates the dose-dependent effects of citalopram on pentylenetetrazole-induced clonic seizure in mice: Involvement of nitric oxide

Citalopram is a selective serotonin reuptake inhibitor (SSRI), widely used in the treatment of depressive disorders. It has been shown that citalopram affects seizure susceptibility. Although the exact mechanism of these effects are not yet fully understood, recent data suggest that 5HT(3) receptors and nitric oxide (NO) might participate in the central effects of SSRIs. In this study in a mouse model of clonic seizure induced by pentylenetetrazole we investigated whether 5-HT(3) receptors are involved in the effects of citalopram on seizure threshold. In our experiments, citalopram at lower doses (0.5 and 1mg/kg, i.p) significantly increased the seizure threshold and at higher doses (≥25mg/kg) showed proconvulsive effects. Moreover, mCPBG (a 5-HT(3) receptor agonist) at low and non-effective doses augmented while non-effective doses of tropisetron prevented the anticonvulsive properties of citalopram. On the other hand, Low doses of nitric oxide synthase inhibitors l-NAME and 7-NI alone or in combination with lower doses of 5-HT(3) receptor agonist enhanced the anticonvulsive property of citalopram, while l-arginine (NO precursor) alone or in combination with tropisetron blocked the protective effect of citalopram. In summary, our findings demonstrate that 5-HT(3) receptor mediates the anticonvulsant properties of low doses of citalopram, whereas it seems that the proconvulsive effect is mostly mediated through the NO pathway and can be totally blocked by NOS inhibitors. This could propose a new approach toward finding the mechanism of citalopram activity, curtailing the adverse effects of citalopram and perhaps managing the convulsions as a vicious consequence of citalopram overdose.     

Intrathecal administration of 5-HT(3) receptor agonist modulates jaw muscle activity evoked by injection of mustard oil into the temporomandibular joint in the rat

The effect of intrathecal administration of the 5-HT(3) receptor agonist 2-methyl-5-hydroxytryptamine (2m-5HT) on jaw muscle activity evoked by mustard oil (MO) injection into the temporomandibular joint of anesthetized rats was examined. One microgram or 100 microg of 2m-5HT significantly enhanced or suppressed jaw muscle responses, respectively. Pre-administration of tropisetron, a 5-HT(3) receptor antagonist, attenuated the effect of 2m-5HT. These results indicate that activation of 5-HT(3) receptors can modulate trigeminal nociceptive responses.     

Increased binding at 5-HT1A, 5-HT1B, and 5-HT2A receptors and 5-HT transporters in diet-induced obese rats

5-Hydroxytryptamine (5-HT, serotonin), synthesized in midbrain raphe nuclei and released in various hypothalamic sites, decreases food intake but the specific 5-HT receptor subtypes involved are controversial. Here, we have studied changes in the regional density of binding to 5-HT receptors and transporters and the levels of tryptophan hydroxylase, in rats with obesity induced by feeding a palatable high-energy diet for 7 weeks. We mapped binding at 5-HT receptor subtypes and transporters using quantitative autoradiography and determined tryptophan hydroxylase protein levels by Western blotting. In diet-induced obese (DiO) rats, specific binding to 5-HT_1A receptors ([³H]8-OH-DPAT) was significantly increased in the dorsal and median raphe by 90% (P<0.01) and 132% (P<0.05), respectively, compared with chow-fed controls. 5-HT_1B receptor binding sites ([¹²⁵I]cyanopindolol) were significantly increased in the hypothalamic arcuate nucleus (ARC) of DiO rats (58%; P<0.05), as were 5-HT_2A receptor binding sites ([³H]ketanserin) in both the ARC (44%; P<0.05) and lateral hypothalamic area (LHA) (121%; P<0.05). However, binding to 5-HT_2C receptors ([³H]mesulgergine) in DiO rats was not significantly different from that in controls in any hypothalamic region. Binding to 5-HT transporters ([³H]paroxetine) was significantly increased (P<0.05) in both dorsal and median raphe, paraventricular nuclei (PVN), ventromedial nuclei (VMH), anterior hypothalamic area (AHA) and LHA of DiO rats, by 47%–165%. Tryptophan hydroxylase protein levels in the raphe nuclei were not significantly different between controls and DiO rats. In conclusion, we have demonstrated regionally specific changes in binding to certain 5-HT receptor subtypes in obesity induced by voluntary overeating of a palatable diet. Overall, these changes are consistent with reduced 5-HT release and decreased activity of the 5-HT neurons. Reduction in the hypophagic action of 5-HT, possibly acting at 5-HT_1A, 5-HT_1B and 5-HT_2A receptors, may contribute to increased appetite in rats presented with highly palatable diet.     

Partial 5-HT(1A) receptor agonist activity by the 5-HT(2C) receptor antagonist SB 206,553 is revealed in rats spinalized as neonates

Modification of spinal serotonergic receptors caudal to spinal injury occurs in rats that received spinal cord transections as neonates. Evaluation of the serotonin syndrome, a group of motor stereotypies elicited by serotonergic (5-HT) agents in 5-HT-depleted animals, and open field locomotor behavior were used to assess behavioral consequences of injury and treatment. We extend these findings to show that a partial 5-HT(1A) agonist activity is revealed by the 5-HT(2C) receptor antagonist (SB 206,553) in this animal model, as measured by evaluation of serotonin syndrome behavior. Treadmill stimulation enhances this motor response, caudal to the injury, in the hindlimbs and tail. These results imply a broader modification of serotonergic receptors than previously thought and suggest a potential strategy by which serotonergic agents may enhance functional recovery following neonatal injury.     

Effect of a 5-HT(1A) receptor agonist, flesinoxan, on the extracellular noradrenaline level in the hippocampus and on the locomotor activity of rats

We have studied effects of 5-hydroxytryptamine 1A (5-HT(1A)) receptor-selective compounds on the extracellular noradrenaline (NA) level in the hippocampus of rats using microdialysis and on their locomotor activity. A selective 5-HT(1A) receptor agonist, flesinoxan (5 mg/kg, i.p.) increased the extracellular NA level in the hippocampus, and increased the locomotor activity. Both responses were blocked by pretreatment with a 5-HT(1A) receptor antagonist, WAY100635 (1 mg/kg, i.p.) and an alpha(2) adrenoceptor agonist, clonidine (50 microg/kg, i.p.). Bilateral intrahippocampal injection of flesinoxan (200 nmol in 2 microl, respectively) increased the locomotor activity of rats and the intrahippocampal perfusion of flesinoxan (1 mM, 2 microl/min) increased the extracellular NA level in the hippocampus. Bilateral intrahippocampal injections of a small amount of WAY100635 (0.1 nmol in 2 microl, respectively) blocked the flesinoxan (5 mg/kg, i.p.)-induced hyperactivity. Flesinoxan (5 mg/kg, i.p.) did not significantly influence the level of serotonin or its major metabolite in the hippocampus, or dopamine or its metabolites in the striatum. In conclusion, these behavioral as well as pharmacological results indicate that postsynaptic 5-HT(1A) receptor activation by flesinoxan increase the extracellular NA level in the hippocampus, which may be the cause of the increase of the locomotor activity.     

Effects of 5-HT2A receptor antagonist on blood flow in chronically compressed nerve roots

Neurogenic intermittent claudication (NIC) can be caused by compression of the cauda equina by spinal stenosis and is a major clinical problem. A reduction of blood flow is an important mechanism for inducing NIC and may be caused by a vasoconstrictive effect mediated by the serotonin 5-HT2A receptor in chronic cauda equina compression lesions. This study assessed the effects of the 5-HT2A receptor antagonist on nerve vasculature in chronically compressed nerve roots. A plastic balloon was placed under the lamina of L7 and inflated to 10 mmHg and left for 1 week in several cauda equina compression models. All experimental animals received an acute administration of serotonin. One group received sarpogrelate hydrochloride (5-HT2A receptor antagonist: 5-HTRA) before administration of serotonin, and another group was administered 5-HTRA after administration of serotonin. Diameters and blood flow in the vasculature of S2 or S3 nerve roots were measured after injection of serotonin. In animals without compression of the cauda equina (sham), blood vessels contracted and the blood flow was reduced after administration of serotonin. In sham and compression animals receiving both serotonin and 5-HTRA, blood vessel diameter was not reduced and was significantly larger than that in the compression group receiving only serotonin (p<0.05). Likewise, the blood flow was not reduced in sham and compression animals receiving serotonin and 5-HTRA and was significantly greater in the compression group treated only with serotonin. 5-HTRA inhibited vasoconstriction and the reduction of blood flow in chronically compressed nerve roots challenged with serotonin. This fact suggests that 5-HTRA might be effective at improving blood flow in chronically compressed nerve roots in patients with spinal canal stenosis and changes in circulation levels of serotonin.     

5-HT(1A) receptor-mediated inhibition and 5-HT(2) as well as 5-HT(3) receptor-mediated excitation in different subdivisions of the rat amygdala

The techniques of extracellular single cell recording and microiontophoresis were used to study the effects of serotonin (5-HT) and of 5-HT(1A), 5-HT(2A/2C) and 5-HT(3) receptor agonists on the spontaneous activity of amygdaloid neurons in rats anesthetized with urethane. The background discharge rate was modified by 5-HT as well as by 5-HT agonists in about two-thirds of neurons tested in different nuclei of the amygdaloid complex. Whereas the 5-HT(2) and 5-HT(3) agonists significantly increased the neuronal discharge rate in nearly all subdivisions of the amygdala, the 5-HT(1A) agonist significantly inhibited the firing rate. Co-administration of bicuculline and 5-HT receptor agonists prevented the 8-OH-DPAT-induced increases in the firing rate in most cases tested, as well as the inhibitory effects of DOI or 2-methyl-5HT. Therefore, GABAergic interneurons seem to be involved in the mediation of serotonergic effects. The action of 5-HT agonists on the neuronal discharge rate was blocked by different receptor-specific antagonists. The results support the hypothesis that 5-HT exerts control throughout the amygdala by acting at least on 5-HT(1A), 5-HT(2A/2C) and 5-HT(3) receptors seemingly located both on projection and interneurons.     

Potent serotonin (5-HT)(2A) receptor antagonists completely prevent the development of hyperthermia in an animal model of the 5-HT syndrome

The serotonin (5-HT) syndrome is the most serious side effect of antidepressants, and it often necessitates pharmacotherapy. In the present study, the efficacy of several drugs was evaluated in an animal model of the 5-HT syndrome. When 2 mg/kg of clorgyline, a type-A monoamine oxidase inhibiting antidepressant, and 100 mg/kg of 5-hydroxy-L-tryptophan, a precursor of 5-HT, were administered intraperitoneally to rats to induce the 5-HT syndrome, the rectal temperature of the rats increased to more than 40 degrees C, and all of the animals died by 90 min after the drug administration. The noradrenaline (NA) levels in the anterior hypothalamus, measured by microdialysis, increased to 15.9 times the preadministration level. Pretreatment with propranolol (10 mg/kg), a 5-HT(1A) receptor antagonist as well as a beta-blocker, and dantrolene (20 mg/kg), a peripheral muscle relaxant, did not prevent the death of the animals, even though these two drugs suppressed the increase in rectal temperature to some extent. Chlorpromazine and cyproheptadine prevented the lethality associated with the 5-HT syndrome only at high doses. By contrast, pretreatment with ritanserin (3 mg/kg) and pipamperone (20 mg/kg), both potent 5-HT(2A) receptor antagonists, completely prevented the increase in rectal temperature and death of the animals, and the hypothalamic NA levels in these two groups increased less than that in the other groups. These results suggest that potent 5-HT(2A) receptor antagonists are the most effective drugs for treatment of the 5-HT syndrome, and that NA hyperactivity occurs in the 5-HT syndrome.     

Evidence that spinal 5-HT1, 5-HT2 and 5-HT3 receptor subtypes modulate responses to noxious colorectal distension in the rat

This study examined whether the antinociception produced following the intrathecal (i.t.) administration of serotonin (5-hydroxytryptamine, 5-HT) and other 5-HT receptor agonists in a model of visceral pain that utilizes colorectal distension (CRD) as the noxious visceral stimulus is mediated through interaction with spinal 5-HT1, 5-HT2, or 5-HT3 receptor subtypes. CRD in conscious rats reliably elicits two pseudaffective reflexes: a vigorous pressor response and a visceromotor response. Antinociception is characterized by inhibition of both pseudaffective responses. The effects of 5-HT receptor agonists and antagonists on resting blood pressure were also examined. The i.t. administration of 5-HT resulted in a dose-dependent elevation of the visceromotor threshold and inhibition of the pressor response to CRD. The 5-HT1A receptor agonist 8-OH-DPAT, the 5-HT1B receptor agonist RU-24969, the 5-HT2 receptor agonists DOI, MK-212 and alpha-methyl-5-HT and the 5-HT3 agonist 2-methyl-5-HT all dose-dependently inhibited the pressor response and dose-dependently elevated the visceromotor threshold to noxious CRD. The rank order of potency of these agonists was the same for both pseudaffective responses to CRD: DOI greater than or equal to 8-OH-DPAT greater than or equal to MK-212 = RU-24969 greater than or equal to alpha-methyl-5-HT = 2-methyl-5-HT much greater than 5-HT. The antinociceptive effects of 5-HT, RU-24969, alpha-methyl-5-HT and DOI were antagonized by i.t. pretreatment with methysergide. Intrathecal pretreatment with ketanserin antagonized the antinociceptive effects of MK-212 and MDL-72222 antagonized the effects produced by 2-methyl-5-HT in response to CRD. The antinociceptive effects produced by 8-OH-DPAT were not antagonized by i.t. pretreatment with methysergide. These results demonstrate that 5-HT1, 5-HT2 and 5-HT3 receptors in the spinal cord mediate antinociception in response to noxious CRD in conscious rats.     

Altered expression and functions of serotonin 5-HT1A and 5-HT1B receptors in knock-out mice lacking the 5-HT transporter

By taking up serotonin (5-hydroxytryptamine, 5-HT) released in the extracellular space, the 5-HT transporter (5-HTT) regulates central 5-HT neurotransmission. Possible adaptive changes in 5-HT neurotransmission in knock-out mice that do not express the 5-HT transporter were investigated with special focus on 5-HT1A and 5-HT1B receptors. Specific labelling with radioligands and antibodies, and competitive RT-PCR, showed that 5-HT1A receptor protein and mRNA levels were significantly decreased in the dorsal raphe nucleus (DRN), increased in the hippocampus and unchanged in other forebrain areas of 5-HTT-/- vs. 5-HTT+/+ mice. Such regional differences also concerned 5-HT1B receptors because a decrease in their density was found in the substantia nigra (-30%) but not the globus pallidus of mutant mice. Intermediate changes were noted in 5-HTT+/- mice compared with 5-HTT+/+ and 5-HTT-/- animals. Quantification of [35S]GTP-gamma-S binding evoked by potent 5-HT1 receptor agonists confirmed such changes as a decrease in this parameter was noted in the DRN (-66%) and the substantia nigra (-30%) but not other brain areas in 5-HTT-/- vs. 5-HTT+/+ mice. As expected from actions mediated by functional 5-HT1A and 5-HT1B autoreceptors, a decrease in brain 5-HT turnover rate after i.p. administration of ipsapirone (a 5-HT1A agonist), and an increased 5-HT outflow in the substantia nigra upon local application of GR 127935 (a 5-HT1B/1D antagonist) were observed in 5-HTT+/+ mice. Such effects were not detected in 5-HTT-/- mice, further confirming the occurrence of marked alterations of 5-HT1A and 5-HT1B autoreceptors in these animals.     

Reduction in 5-HT1A receptor density, 5-HT1A mRNA expression, and functional correlates for 5-HT1A receptors in genetically defined aggressive rats

The present experiments tested the hypothesis that one of the critical mechanisms underlying genetically defined aggressiveness involves brain serotonin 5-HT1A receptors. 5-HT1A receptor density, the receptor mRNA expression in brain structures, and functional correlates for 5-HT1A receptors identified as 8-OH-DPAT-induced hypothermia and lower lip retraction (LLR) were studied in Norway rats bred for 59 generations for the lack of aggressiveness and for high affective aggressiveness with respect to man. Considerable differences between the highly aggressive and the nonaggressive rats were shown in all three traits. A significant decrease in B(max) of specific receptor binding of [3H]8-OH-DPAT in the frontal cortex, hypothalamus, and amygdala and a reduction in 5-HT1A receptor mRNA expression in the midbrain of aggressive rats were found. 5-HT1A receptor agonist 8-OH-DPAT (0.5 mg/kg, i.p.) produced a distinct hypothermic reaction in nonaggressive rats and did not affect significantly the body temperature in aggressive rats. Similar differences were revealed in 8-OH-DPAT-induced LLR: LLR was expressed much more in nonaggressive than in aggressive animals. Additionally, 8-OH-DPAT (0.5 mg/kg i.p.) treatment significantly attenuated the aggressive response to man. The results demonstrated an association of aggressiveness with reduced 5-HT1A receptor expression and function, thereby providing support for the view favoring the idea that brain HT1A receptor contributes to the genetically defined individual differences in aggressiveness.