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The medial prefrontal cortex has been shown to participate in pain processing and to undergo structural and functional changes in chronic visceral pain syndromes. Its infralimbic region (IL) is directly involved in central autonomic regulation and provides extensive projections to pain-related brain centers, being most capable of modulating visceral nociception. However, the precise neuronal and molecular mechanisms underlying IL-exerted descending control of visceral pain and their probable alterations in pathology remain unknown. Considering the high levels of serotonin 5-HT3 receptors in the IL and their important role in visceral pain regulation, in this study performed in anesthetized adult male Wistar rats, we aimed to determine whether 5-HT3 receptor-dependent mechanisms are involved in the IL influence on visceral pain-responsive neurons of the caudal ventrolateral medulla (CVLM) and whether intestinal inflammation affects the process. Our results showed that electrical stimulation of the IL caused a facilitation of both excitatory and inhibitory CVLM neuron responses to noxious colorectal distension (CRD). Both effects were 5-HT3-dependent, being decreased following intracerebroventricular injection of a 5-HT3-antagonist, granisetron (20 µg in 10 µL saline). In trinitrobenzenesulfonic acid-induced colitis, the IL stimulating influence on CRD-excited CVLM neurons intensified, while the cortex-driven facilitation of CRD-inhibited cells reduced; both appeared resistant to the 5-HT3 receptor blockade. These data provide an insight to the specific mechanisms underlying cortical modulation of acute and gut pathology-associated visceral nociception, promoting the development of differentiated strategies for their treatment. [ABSTRACT FROM AUTHOR]

Rationale: 2-Amino-6-chloro-3,4-dihydroquinazoline (e.g., A6CDQ) represents a novel putative antidepressant originally thought to act through a 5-HT3 serotonin receptor antagonist mechanism. Here, we investigated this further by examining a positional isomer of A6CDQ (i.e., A7CDQ). Materials and methods: 5-HT3 receptor and transporter activity (uptake-1 and uptake-2) were investigated using a variety of in vitro assays and the in vivo mouse tail suspension test (TST). Results: Although A7CDQ binds at 5-HT3 receptors with low affinity (Ki = 1975 nM) compared to A6CDQ (Ki = 80 nM), it retained 5-HT3 receptor antagonist action (IC50 = 5.77 and 0.26 μM, respectively). In the mouse TST A7CDQ produced antidepressant-like actions (ED50 = 0.09 mg/kg) comparable to that of A6CDQ. In addition, A6CDQ was found to be a 5-HT releasing agent (Km = 2.8 μM) at hSERT and a reuptake inhibitor (IC50 = 1.8 μM) at hNET, whereas A7CDQ was a weak reuptake inhibitor (Km = 43.6 μM) at SERT but a releasing agent (EC50 = 3.3 μM) at hNET. Moreover, A6CDQ and A7CDQ were potent inhibitors of uptake-2 (e.g.; OCT3 IC50 = 3.9 and 5.9 μM, respectively). Conclusions: A simple shift of a substituent in a common quinazoline scaffold from one position to another (i.e., a chloro group from the 6- to the 7-position) resulted in a common action in the TST but via a somewhat different mechanism. A6CDQ and A7CDQ might represent the first members of a new class of potential antidepressants with a unique multi-modal mechanism of action. [ABSTRACT FROM AUTHOR]

The caudal ventrolateral reticular formation of the medulla oblongata is the first layer of visceral nociceptive processing. In experiments on rats, neuronal responses in this zone to nociceptive stimulation of the large intestine were examined and the effects of selective blockade of 5-HT3 receptors on these responses were assessed. By the character of responses to nociceptive colorectal stimulation (CRS), the recorded medullary neurons were divided into three groups-excited, inhibited and indifferent. Intravenous injection of 5-HT3 antagonist granisetron (1 and 2 mg/kg) as well as local application of this agent on the surface of the medulla oblongata (1.25 and 2.5 nmole) suppressed the background and evoked firing of CRS-excited reticular neurons in a dose-dependent manner but did not exert as pronounced influence on the cells inhibited by visceral nociceptive stimulation. Spike activity in the group of CRS-indifferent neurons under similar conditions was 5-HT3-independent. The results obtained provide evidence that 5-HT3 receptors mediate the facilitating effect of serotonin on supraspinal transmission of the abdominal nociceptive stimulus which, at least in part, is realized via selective activation of visceral medullary nociceptive neurons. A shutdown of this mechanism may underlie the analgesic effect of 5-HT3 antagonists in abdominal pain syndromes. [ABSTRACT FROM AUTHOR]

Background and Objective: Although 5-fluorouracil (5-FU) is a widely used as chemotherapy agent, severe mucositis develops in approximately 80 % of patients. 5-FU-induced small intestinal mucositis can cause nausea and vomiting. The current study was designed to investigate peripheral alterations due to the 5-FU-induced mucositis of neuronal and non-neuronal 5-HT 3 and NK 1 receptor expression by immunohistochemical analysis. Methods: 5-FU was administered by i.p. injection to C57BL/6 mice. After 4 days, segments of the jejunum were removed. The specimens were analyzed by immunohistochemistry, real-time PCR, and enzyme immunoassay. Results: The numbers of 5-HT 3 receptor immunopositive cells and nerve fibers in mucosa were increased by 5-FU treatment. The 5-HT 3 receptor immunopositive cell bodies were found only in jejunal submucosa and myenteric plexus in the 5-FU-treated mice. The numbers of NK 1 receptor cells in mucosa and immunopositive expression of NK 1 receptors in deep muscular plexus were dramatically increased in 5-FU-treated mice. Real-time PCR demonstrated that 5-FU treatment significantly increased mRNA levels of 5-HT 3A, 5-HT 3B, and NK 1 receptors. The amounts of 5-HT and substance P increased after 5-FU treatment. The 5-HT 3 or NK 1 receptor immunopositive cells colocalized with both 5-HT and substance P. Furthermore, 5-HT 3 and NK 1 receptors colocalized with CD11b. Conclusions: The 5-HT 3 and NK 1 immunopositive macrophages and mucosal mast cells in lamina propria release 5-HT and substance P, which in turn activate their corresponding receptors on mucosal cells in autocrine and paracrine manners. It is assumed to result in the release of 5-HT and substance P in mucosa. [ABSTRACT FROM AUTHOR]

Rationale and objectives: We previously found that systemic injections of the 5-HT uptake blocker fluoxetine attenuate intermittent footshock stress-induced reinstatement of alcohol seeking in rats, while inhibition of 5-HT neurons in the median raphe induces reinstatement of alcohol seeking. In this study, we further explored the role of 5-HT in footshock stress-induced reinstatement of alcohol seeking by determining the effects of the 5-HT releaser and reuptake blocker dexfenfluramine, and the 5- HT receptor antagonists ondansetron and tropisetron, which decrease alcohol self-administration and anxiety-like responses in rats, on this reinstatement. Methods: Different groups of male Wistar rats were trained to self-administer alcohol (12% v/v) for 28-31 days (1 h/day, 0.19 ml per alcohol delivery) and then their lever responding for alcohol was extinguished over 9-10 days. Subsequently, the effect of systemic injections of vehicle or dexfenfluramine (0.25 or 0.5 mg/kg, i.p), ondansetron (0.001, 0.01, or 0.1 mg/kg, i.p), or tropisetron (0.001, 0.01, and 0.1 mg/kg, i.p) on reinstatement induced by 10 min of intermittent footshock (0.8 mA) was determined. Results: Systemic injections of dexfenfluramine, ondansetron or tropisetron attenuated footshock-induced reinstatement of alcohol seeking. Injections of dexfenfluramine, ondansetron, or tropisetron had no effect on extinguished lever responding in the absence of footshock. Conclusions: The present results provide additional support for the hypothesis that brain 5-HT systems are involved in stress-induced reinstatement of alcohol seeking. The neuronal mechanisms that potentially mediate the unexpected observation that both stimulation of 5-HT release and blockade of 5-HT3 receptors attenuate footshock-induced reinstatement are discussed. [ABSTRACT FROM AUTHOR]

Rationale: Temporal differentiation refers to animals' ability to regulate their behaviour during an ongoing interval. Striatal dopaminergic mechanisms are purported to be involved in temporal differentiation, and recent evidence also implicates 5-hydroxytryptaminergic (5-HTergic) mechanisms, possibly mediated by 5-HT2A receptors. There is evidence that 5-HT3 receptors contribute to the regulation of dopamine release in the basal ganglia; however, it is not known whether 5-HT3 receptor stimulation can influence temporal differentiation. Objective: We examined the effects of a selective 5-HT3 receptor agonist m-CPBG, a mixed 5-HT2A/3 receptor agonist quipazine, and selective 5-HT3 and 5-HT2A receptor antagonists (MDL- 72222 and ketanserin, respectively) on temporal differentiation in a free-operant psychophysical procedure. Methods: Twenty-four rats were trained to respond on two levers (A and B) under a free-operant psychophysical schedule, in which sucrose reinforcement (0.6 M, 50 µl) was provided intermittently for responding on A during the first half and on B during the second half of 50-s trials. Logistic psychometric functions were fitted to the relative response rate data [percent responding on B (%B) vs time from trial onset (t)], and quantitative indices of timing performance [T50 (value of t corresponding to %B=50), Weber fraction, and mean time of switching from A to B, S50] were derived. Results: Quipazine (0.5, 1, and 2 mg kg-1) altered timing performance, dose-dependently reducing T50 and S50; m-CPBG (2.5, 5, and 10 mg kg-1) had no significant effect. The effect of quipazine was antagonized by ketanserin (2 mg kg-1), but not by MDL-72222 (1 mg kg-1). Conclusions: The present results provide no evidence for the involvement of 5- HT3 receptors in temporal differentiation and indicate that the effect of quipazine on performance was mediated by 5-HT2A receptor stimulation. The results are consistent with previous evidence for the involvement of 5-HT2A receptors in interval timing behaviour. [ABSTRACT FROM AUTHOR]

The patch-clamp technique was used on excised (outside-out) patches to characterize h5-HT3A receptors stably transfected in HEK 293 cells and to compare the effects of the barbiturate anaesthetics methohexital and pentobarbital on this ligand-gated cation channel. At negative membrane potentials 5-HT induced inward currents in a concentration-dependent manner (EC50=8.6 µM, Hill coefficient =1.5). The mean peak current induced by 30 µM 5-HT was –110 pA at –100 mV. The 5-HT3A receptor antagonist ondansetron (0.3 nM) reversibly inhibited the 5-HT (30 µM) signal by 70% and at 3 nM it abolished the response. Methohexital and pentobarbital inhibited 5-HT-induced (30 µM) currents in a concentration-dependent manner. The maximal inhibition with a given methohexital or pentobarbital concentration was reached when the respective drug was applied 45 s prior to and during the 2-s 5-HT pulse (IC50 values=95 µM and 127 µM, Hill coefficient =–1.0 and –1.6, respectively). Although the barbiturates were, thus, equipotent, their effects differed substantially with respect to the dependence on the time schedule of application to the patches: the potency of methohexital was virtually maximal when the drug was applied exclusively 45 s before the agonist pulse, but its inhibitory potency decreased considerably when it was exclusively applied during the 2-s 5-HT pulse (IC50=380 µM). Conversely, pentobarbital was almost maximally potent in inhibiting the 5-HT signal when it was exclusively coapplied with this agonist, but its inhibitory potency was considerably lower (IC50 ~500 µM) when applied exclusively 45 s before 5-HT. Another difference between both barbiturates involves the rate of inactivation of 5-HT3 receptor-mediated currents: whereas high concentrations of methohexital (≥300 µM) were necessary to induce moderate (≤ twofold) acceleration of this parameter, pentobarbital produced such an effect at all concentrations and the extent of acceleration increased with increasing concentration (1.5- to fivefold). In conclusion, two barbiturates, chemically closely related but of different lipophilicity, clearly differ with respect to the kinetics of their effect on 5-HT3 receptor channels; one possible explanation involves drug access to an amphipathic site of action via both an aqueous and a hydrophobic pathway. Pentobarbital, in contrast to methohexital, inhibits h5-HT3A receptor-mediated currents at anaesthetic concentrations (~90 µM). [ABSTRACT FROM AUTHOR]

Nearly 30% of patients treated with metformin experience gastrointestinal side effects. Since release of 5-hydroxytryptamine (5-HT) from the intestine is associated with nausea, vomiting, and diarrhea, we examined whether metformin induces 5-HT release from the intestinal mucosa. In 40% of tissue biopsy specimens of human duodenal mucosa, metformin (1, 10, and 30 µM) caused an increase in 5-HT outflow by 35, 70, and 98%, respectively. Peak increases in 5-HT outflow were observed after 10–15 min exposure to metformin, returning to baseline levels after 25 min. Tetrodotoxin (1 µM) reduced by about 50% the metformin-evoked increase in 5-HT outflow (P<0.05). Metformin-evoked release was not affected by scopolamine + hexamethonium, propranolol, the 5-HT3 receptor antagonist dolasetron, naloxone, or the NK1 receptor antagonist L703606. In the presence of tetrodotoxin (1 µM), somatostatin (1 µM) further reduced metformin-induced 5-HT release by 15–20%. In view of the 5-HT releasing effects of selective 5-HT3 receptor agonists to which metformin (N-N-dimethylbiguanide) is structurally related, we investigated whether metformin directly interacts with 5-HT3 receptors. Receptor binding (inhibition of [3H]-GR65630 binding) and agonist effects (stimulation of [14C]-guanidinium influx) at 5-HT3 receptors were studied in murine neuroblastoma N1E-115 cells, which express functional 5-HT3 receptors. Metformin up to 0.3 mM failed to inhibit [3H]-GR65630 binding and to modify displacement of [3H]-GR65630 binding induced by 5-HT. 5-HT (3 µM) stimulated the influx of [14C]-guanidinium in intact N1E-115 cells. Metformin up to 1 mM failed to modify basal influx, 5-HT-induced influx, and 5-HT+ substance P-induced influx of [14C]-guanidinium. Our results indicate that metformin induces 5-HT3 receptor-independent release of 5-HT from human duodenal mucosa via neuronal and non-neuronal mechanisms. Part of the gastrointestinal side effects observed during treatment with metformin could, thus, be produced by the release of 5-HT and other neurotransmitter substances within the duodenal mucosa. [ABSTRACT FROM AUTHOR]

The influence of diabetic hyperglycemia on solid gastric emptying in rats was examined. Diabetes was produced by streptozotocin (STZ, 40mg/kg i.v.), and diabetic hyperglycemia was observed from 1 day after the STZ injection. The gastric emptying of glass beads in the diabetic rats was significantly delayed compared with that in age-matched control rats at 1, 3 and 7 days after diabetes induction. A slight decrease in gastric emptying was observed in the diabetic rats from 2 to 52 weeks after the diabetes induction. We also investigated the influence of gastroprokinetic agents on STZ-induced diabetic gastroparesis and subdiaphragmatic vagotomy-induced gastroparesis in rats. The selective 5-HT3 receptor antagonists ramosetron (YM060), YM114 (KAE-393), granisetron and ondansetron, and the substituted benzamides (5-HT4 receptor agonist/5-HT3 receptor antagonists) cisapride mosapride and SC-53116 dose-dependently enhanced gastric emptying in normal rats. These compounds also reversed the impairment of diabetic gastroparesis rats at 7 days after the STZ injection, but higher doses were required. The solid gastric emptying in subdiaphragmatic vagotomized rats was also delayed. Ramosetron and the substituted benzamides cisapride and zacopride partially reversed the gastroparesis in the vagotomized rats. These results suggest that acute hyperglycemia is important mechanism for the delay of solid gastric emptying in diabetic rats. It is also suggested that selective 5-HT3 receptor antagonists and substituted benzamides enhance gastric emptying not only in normal rats but also in diabetic and vagotomized rats. [ABSTRACT FROM AUTHOR]

We investigated the effect of ICS 205-930 [(3α-tropanyl)-1H-indole-3-carboxylic acid ester], a selective antagonist at 5-HT receptors, on the cardiogenic hypertensive chemoreflex in the anaesthetized dog. The reflex was elicited by injection of 5-HT (12.5-1600 μg) into the left cardiac ventricle and consisted of a dose-dependent systemic hypertension associated with a decrease in heart rate. ICS 205-930 (10, 30, and 100 μg/kg i.v.) caused a displacement to the right of both the dose-response curves of 5-HT-induced blood pressure increase and heart rate reduction. Its blocking effects upon the action of 5-HT could be surmounted by increasing the dose of the agonist. The selective 5-HT receptor antagonist, ketanserin (0.1 mg/kg i.v.) and the combined 5-HT and 5-HT receptor antagonist, methiothepin (0.1 mg/kg i.v.) had no influence on the hypertensive reflex. When the reflex was elicited by the ganglionic stimulant, 1,1-dimethyl-4-phenyl-piperazinium (DMPP; 100-1600 μg), ICS 205-930 had no blocking effect. The results suggest that the 5-HT-induced cardiogenic hypertensive chemoreflex is mediated by 5-HT receptors. [ABSTRACT FROM AUTHOR]