Central 5HT Research Articles

Live or heat-killed probiotic administration reduces anxiety and central cytokine expression in BALB/c mice, but differentially alters brain neurotransmitter gene expression

While the potential for probiotic supplements to act as adjunctive treatments for mood disorders has been widely demonstrated, the precise mode of action remains unclear. To investigate the psychotropic effects of a multi-species probiotic on emotional behaviour in male BALB/c mice, we explored the potential mechanisms of action relating to the temporal changes in the mRNA expression of brain cytokines, BDNF, central 5HT receptor and serotonin transporter (SERT) and GABA receptor in the context of probiotic induced behavioural changes. The effects of a heat-killed probiotic, independent of microbial metabolic processes were also evaluated on the same outcomes to understand whether the host response to the bacteria is more or less important than the contribution of the metabolic activity of the bacteria themselves. Results showed that probiotic supplementation reduced anxiety-like behaviours, increased time spent in the light area of the light-dark box, and decreased the expression of pro-inflammatory cytokines in the brain. Furthermore, probiotic administration elevated hippocampal BDNF and decreased GABAB1β expression. Interestingly, the heat-killed probiotic and its membrane fraction had similar effects on emotional behaviours and gene expression in the brain. The ingestion of live and heat-killed probiotic preparations also reduced TLR2 expression in the gut. Thus, the present study reveals that the anxiolytic action of a multispecies probiotic in BALB/c mice is independent of bacterial viability. This suggests that it is the host response to probiotics, rather than microbial metabolism that facilitates the molecular changes in the brain and downstream behaviours.This article is part of the Special Issue on “Microbiome & the Brain: Mechanisms & Maladies”.

P.2.027 - Inducible tryptophan hydroxylase 2 knockout mice as model for anxiety and fear like behaviour

Background The amygdaloid nuclei are involved in expression of anxiety and fear responses. Malfunctioning of these nuclear complexes may be due in part to dysregulation serotonin (5-HT) neurotransmission. However the neural mechanisms involved are yet to be fully deciphered. Constitutive tryptophan hydroxylase 2 knockout mice (Tph2-/-) which lack 5-HT from embryonic stages of development show increased aggression and altered anxiety and fear responses in aversive contexts[1]. Still, it is unclear whether the observed phenotypes are attributed to abnormal development resulting from 5-HT deficiency or to a specific requirement for 5-HT in adulthood [2]. Inducible Cre-mediated recombination of Tph2 during adulthood, which permits time specific depletion of brain 5-HT may decipher the specific role of 5-HT in the regulation of emotional responses. Aim Characterization of a mouse model for effect of acute brain serotonin depletion on anxiety and fear-like behavior. Methods Mice (10-12weeks old) expressing Cre-recombinase under the Tph2 specific promoter and with Exon V of Tph2 flanked by loxP sites were injected with tamoxifen (2mg, daily for 5 consecutive days) to induce conditional Tph2 deletion (Tph2 icko). Four weeks after injection, immunohistochemistry and high performance liquid chromatography were performed (HPLC) to assess the efficiency of Cre-mediated recombination of Tph2 in raphe nuclei. The mice were subjected to behavioural testing including elevated plus maze, light-dark box, open-field and fear conditioning in an automated fear conditioning chamber. Afterwards, brains were fixed in 4% PFA and used for immunofluorescence staining of 5HT. Data are represented as mean±SEM and Students’ t-test was use to compared means between Tph2 icko mice and sham controls (Tph2CON). A p-value less than 0.05 was regarded as significant. Results The Tph2 icko mice showed significant (p Conclusion Cre-mediated inducible Tph2 deletion may be able to model the impact of central 5HT depletion on anxiety- and fear-like behavior in a time-specific manner.

Spinal Cord Injury Leads to Reductions in Central Nervous System 5HT Coupled with Blunting of Arousal and Genioglossus Muscle Responses to Hypercapnia in Mice

PurposeThe present investigation was designed to examine if spinal cord injury (SCI) leads to reductions in central nervous system serotonin (5HT). The investigation was also designed to investigate if this reduction was coupled to blunting of arousal responses (AR) and genioglossus muscle sensitivity (GS) to hypercapnia.MethodsTelemetry transmitters were surgically implanted in wild type mice (Tph2+/+) (n = 4) and tryptophan hydroxylase 2 knockout mice (Tph2−/−) (n = 4) to measure electroencephalography, genioglossus muscle activity, core body temperature and gross motor activity. Following recovery, the Tph2+/+ and Tph2−/− mice were placed in whole body plethysmographs and exposed to episodes of 7 % carbon dioxide to determine the AR and GS to hypercapnia during non‐rapid eye movement (NREM) sleep. Following these measures a C2 hemisection of the spinal cord was completed and AR and GS were measured subsequently. In addition, high‐performance liquid chromatography was performed to measure 5HT levels in three brainstem sections (i.e. dorsal respiratory group and hypoglossal nucleus; raphe magnus, obscurus and pallidus; raphe median and paramedian) obtained from 4 intact TPH2+/+ mice and 4 TPH2−/− mice fourteen days after SCI.ResultsTotal 5HT levels in the brainstem were reduced following SCI (3.2 ± 0.1 vs. 1.5 ± 0.3 ng/mg; P < 0.001). In particular, the levels of 5HT in the dorsal respiratory group and hypoglossal nucleus (1.2 ± 0.2 vs. 0.4 ± 0.1 ng/mg; P < 0.01) and in the raphe median and paramedian (1.3 ± 0.2 vs. 0.8 ± 0.1 ng/mg; P < 0.08) were reduced following SCI. In conjunction with the reduction in 5HT the AR (Before SCI: 8.3 ± 4.4 vs. After SCI: 33.8 ± 8.7 s; P < 0.01) and the GS (Before SCI: 180 ± 58 vs. After SCI: 33 ± 10 microvolts; P < 0.02) to hypercapnia was blunted following SCI in the Tph2+/+ but not the Tph2−/− mice.ConclusionsSCI in a spontaneously breathing non‐anesthetized sleeping mouse model was accompanied by a reduction in central nervous system 5HT. This reduction could be responsible for the blunted AR and GS to hypercapnia during NREM sleep. Ultimately, depletion of 5HT could result in an increase in apnea frequency and blunting of the AR and GS could lead to increases in apnea duration following SCI.

Evaluation of antidepressant activity of ondansetron alone and in combination with fluoxetine-an experimental study

Background: Direct antagonism at 5HT 3 receptor site may be associated with antidepressant activity as conventional antidepressants also possess affinity for central 5HT 3 binding site. So in this study, an effort is made to investigate the antidepressant effect of ondansetron (OND), a selective 5HT 3 antagonist, alone and in combination with fluoxetine (FLX). Methods: Acute and chronic models of Forced swimming test (FST), tail suspension test (TST), open field test (OFT) and mice brain serotonin estimation by UV spectrophotometry were applied for the evaluation of antidepressant activity. Results: In FST and TST (acute and chronic models), ondansetron showed statistically significant antidepressant activity (p<0.05) as compared to control. The combination groups (OND 0.25 mg/kg + FLX 5mg/kg & OND 0.5 mg/kg + FLX 5 mg/kg) showed statistically significant antidepressant activity (p<0.05) as compared to fluoxetine (5 mg/kg) and ondansetron (0.25 mg/kg and 0.5 mg/kg) in FST and TST (acute and chronic models). Open field test in chronic study showed ondansetron (0.25 mg/kg and 0.5 mg/kg) increases number of central squares crossed as compared to control which is statistically significant (p<0.05). The combination group (OND 0.25 mg/kg + FLX 5 mg/kg) showed significantly increased (p<0.05) number of central squares crossed as compared to fluoxetine (5 mg/kg). The combination group (OND 0.5 mg/kg + FLX 5 mg/kg) showed significantly increased (p<0.05) brain serotonin compared to control, ondansetron (0.25 mg/kg and 0.5 mg/kg) and fluoxetine (5 mg/kg). Conclusions: Our study concludes that ondansetron alone and in combination with fluoxetine possesses significant antidepressant activity in animal models of depression.

Anorexigenic effect of serotonin is associated with changes in hypothalamic nuclei activity in an avian model

The anorexigenic effect of serotonin (5HT) has been documented for decades; however, its central mechanism has not been fully elucidated, especially so in non-mammalian vertebrates. Therefore, we centrally injected 5HT to chicks and measured several appetite-associated parameters. Chicks that received central 5HT dose- and time-dependently decreased food intake while water intake was not affected. To determine which hypothalamic nuclei were associated with this effect c-Fos immunoreactivity was measured in appetite-associated nuclei. Only the ventromedial hypothalamus and arcuate nucleus were activated. Whole blood glucose was measured after 5HT injection but was not affected. From the hypothalamus, several appetite-associated mRNAs were measured by real-time PCR after 5HT injection but not one of these showed any difference in expression. Lastly, a comprehensive behavior analysis demonstrated that 5HT caused reducing pecking and increased deep rest. Together we interpret these results as exogenous 5HT injection causes short term satiety that is likely a secondary effect to an increase in the amount of time spent in deep rest.

Functional Recovery from Neonatal Breathing Abnormalities in 5HT‐Deficient Pet‐1 Knockout Mice Following Developmental Exposure to Nicotine

Serotonin (5HT) is known to modulate central neural circuits controlling breathing in mammals. Pet-1 gene knockout (KO) leads to a 70% loss of central 5HT neurons and breathing deficits in neonates, including reduced breathing frequency, more spontaneous apneas, delayed autoresuscitation responses, and higher neonatal mortality compared to wild type (WT) littermates. We tested the effects of nicotine (NIC) on breathing in a 5HT-deficient context by exposing fetuses to saline (SAL) or NIC (60 mg/kg dam body weight) from day 5 of gestation via osmotic mini-pump implants in pregnant dams and compared resting ventilation and autoresuscitation in WT and KO neonates. Relative to WT SAL-treated mice, Pet-1 KO neonates suffered depressed resting ventilation, more frequent apneas, and delayed autoresuscitation responses, as expected. However, these breathing deficits were reversed in KO mice following NIC exposure despite continued high neonatal mortality in this group. Analysis of the caudal raphe nuclei showed a similar (~70%) loss of 5HT neurons in both NIC- and SAL-treated KO mice compared to WT controls. We conclude that a full complement of 5HT neurons normally promotes but is not absolutely necessary for producing normal resting ventilation and autoresuscitation responses in mouse neonates, although the underlying mechanism by which NIC exposure can compensate for a 5HT neuron deficit is unclear. In addition, we speculate that the high neonatal mortality of Pet-1 KO mice may be due to abnormal cardiac function, rather than breathing dysfunction per se. Supported by the CJ Foundation for SIDS and a TCNJ SOSA award.

Serotonergic Correlation with Anger and Aggressive Behavior in Acute Stroke Patients: An Intensity Dependence of Auditory Evoked Potentials (IDAP) Study

Anger and aggressive behavior (AB) are two of the main post-stroke behavioral manifestations, which could imply both an anger trait (TA) or a state condition of anger (SA). Serotonergic system is thought to play an inhibitory control on aggressive impulse. Nevertheless, whether 5HT has the same role in TA and in SA, is still debated. Intensity dependence of auditory evoked potentials (IDAP) is thought to be inversely related to the central 5HT tone. The aim of this study was to evaluate, in acute stroke patients, the 5HT system involvement in AB by IDAP. Consecutive stroke patients were evaluated and compared with healthy controls. The Spielberger Trait Anger Scale (STAS) was used to assess AB, SA and TA. Patients with AB and TA showed a significantly increased IDAP value, whereas patients with SA had a significantly lower IDAP; this indicates an increased 5HT tone. In acute stroke patients with AB, there is a decreased central 5HT tone. Surprisingly, we found an opposite 5HT feature between patients with TA and those showing SA, suggesting that the hypothesis of aggression based on 5HT deficiency requires further investigations. This might open new strategies in the treatment of post-stroke AB.

Expression of 5HT-related genes after perinatal treatment with 5HT agonists

Abstract Serotonin (5HT) is a biologically active amine with diverse roles in the mammalian organism. Developmental alterations in 5HT homeostasis could lead to exposure of the developing brain to non-optimal serotonin concentrations that may result in developmental and behavioral deficits. In order to explore the molecular basis of the effects of developmental disturbances on 5HT metabolism on adult central 5HT homeostasis, observed in our previous studies, we measured changes in gene expression of the neuronal 5HT-regulating proteins in adult animals after perinatal treatment with the immediate 5HT precursor 5-hydroxytryptophan (5HTP, 25 mg/kg), or monoamine oxidase (MAO) inhibitor tranylcypromine (TCP 2 mg/kg), during the period of the most intensive development of 5HT neurons — from gestational day 12 until postnatal day 21. Adult animals were sacrificed and the relative mRNA levels for tryptophan hydroxylase 2, MAO A, MAO B, receptors 5HT1A and 5HT2A, 5HT transporter (5HTT) and vesicular monoamine transporter (VMAT) were determined in the raphe nuclei region and prefrontal cortex using Real-Time Relative qRT-PCR. In comparison to the saline treated animals, treatment with 5HTP caused mild but significant increase in MAO A and MAO B mRNA abundance. TCP-treated animals, besides an increase in mRNA abundance for both MAO genes, displayed significantly increased 5HTT and VMAT2 mRNA levels and significantly decreased 5HT1A receptor mRNA levels. Our results suggest that perinatal exposure of rats to 5HTP, and especially TCP, induces long-lasting/permanent changes in the expression of 5HT-regulating genes, that presumably underlie 5HT-related neurochemical and behavioral changes in adult animals.

The relation of developmental changes in brain serotonin transporter (5HTT) and 5HT1A receptor binding to emotional behavior in female rhesus monkeys: Effects of social status and 5HTT genotype

The goal of the present study was to examine how social subordination stress and 5HTT polymorphisms affect the development of brain serotonin (5HT) systems during the pubertal transition in female rhesus monkeys. We also examined associations with developmental changes in emotional reactivity in response to a standardized behavioral test, the Human Intruder (HI). Our findings provide the first longitudinal evidence of developmental increases in 5HT1A receptor and 5HTT binding in the brain of female primates from pre- to peripuberty. The increase in 5HT1A BPND in these socially housed female rhesus monkeys is a robust finding, occurring across all groups, regardless of social status or 5HTT genotype, and occurring in the left and right hemispheres of all prefrontal regions studied, as well as the amygdala, hippocampus, hypothalamus, and raphe nuclei. 5HTT BPND also showed an increase with age in raphe, anterior cingulate cortex, and dorsolateral prefrontal cortex. These changes in brain 5HT systems take place as females establish more adult-like patterns of social behavior, as well as during the HI paradigm. Indeed, the main developmental changes in behavior during the HI (increase in freezing and decrease in submission/appeasement) were related to neurodevelopmental increases in 5HT1A receptors and 5HTT, because the associations between these behaviors and 5HT endpoints emerge at peripuberty. We detected an effect of social status on 5HT1A BPND in the hypothalamus and on 5HTT BPND in the orbitofrontal cortex, with subordinates showing higher BPND than dominants in both cases during the pubertal transition. No main effects of 5HTT genotype were observed for 5HT1A or 5HTT BPND. Our findings indicate that adolescence in female rhesus monkeys is a period of central 5HT reorganization, partly influenced by exposure to the social stress of subordination, that likely functions to integrate adrenal and gonadal systems and shape the behavioral response to emotionally challenging social situations.

The effects of the perinatal treatment with 5-hydroxytryptophan or tranylcypromine on the peripheral and central serotonin homeostasis in adult rats

Serotonin (5HT) is a biologically active amine present in mammals in the brain and the peripheral tissues. Autism is a neurodevelopmental disorder in which 5HT homeostasis is disturbed both centrally and peripherally, but the relationship between the 5HT disturbances in the two compartments is not understood. In an attempt to explore the relationship between the disturbed peripheral 5HT homeostasis and central 5HT functioning, we exposed the developing rat brain to increased 5HT concentrations, by treatment of rats with subcutaneous injections of the immediate 5HT precursor 5-hydroxy-l-tryptophan (5HTP, 25mg/kg), or the non-selective MAO inhibitor tranylcypromine (TCP, 2mg/kg), during the period of the most intensive development of 5HT neurons – from gestational day 13 to post-natal day 21. The effects of the mentioned treatments on peripheral and central 5HT levels were then studied in adult rats. Platelet and plasma 5HT concentrations (measured by ELISA), as well as cortical and midbrain 5HT, tryptophan and 5-hydroxyindoleacetic acid levels (measured by HPLC) were determined in twelve 5HTP treated and eight TCP treated rats, and compared with the values measured in 10 control, saline treated rats. Treatment with 5HTP significantly raised peripheral but not central 5HT concentrations. At adult age, peripheral 5HT homeostasis was re-established, while modest decrease in 5HT concentration was observed in frontal cortex, presumably due to hyperserotonemia-induced loss of 5HT terminals during brain development. Treatment with TCP induced significant 5HT elevations in both compartments. At adult age, permanent changes in 5HT homeostasis were observed, both peripherally (as hyperserotonemia) and centrally (as altered 5HT metabolism with decreased 5HT concentrations). Further studies are planned in order to explore the nature of the different disturbances of 5HT homeostasis induced by the two compounds, and their results are expected to shed some light on the role of hyperserotonemia in autism.

Methods to Quantify Pharmacologically Induced Alterations in Motor Function in Human Incomplete SCI

Spinal cord injury (SCI) is a debilitating disorder, which produces profound deficits in volitional motor control. Following medical stabilization, recovery from SCI typically involves long term rehabilitation. While recovery of walking ability is a primary goal in many patients early after injury, those with a motor incomplete SCI, indicating partial preservation of volitional control, may have the sufficient residual descending pathways necessary to attain this goal. However, despite physical interventions, motor impairments including weakness, and the manifestation of abnormal involuntary reflex activity, called spasticity or spasms, are thought to contribute to reduced walking recovery. Doctrinaire thought suggests that remediation of this abnormal motor reflexes associated with SCI will produce functional benefits to the patient. For example, physicians and therapists will provide specific pharmacological or physical interventions directed towards reducing spasticity or spasms, although there continues to be little empirical data suggesting that these strategies improve walking ability.In the past few decades, accumulating data has suggested that specific neuromodulatory agents, including agents which mimic or facilitate the actions of the monoamines, including serotonin (5HT) and norepinephrine (NE), can initiate or augment walking behaviors in animal models of SCI. Interestingly, many of these agents, particularly 5HTergic agonists, can markedly increase spinal excitability, which in turn also increases reflex activity in these animals. Counterintuitive to traditional theories of recovery following human SCI, the empirical evidence from basic science experiments suggest that this reflex hyper excitability and generation of locomotor behaviors are driven in parallel by neuromodulatory inputs (5HT) and may be necessary for functional recovery following SCI. The application of this novel concept derived from basic scientific studies to promote recovery following human SCI would appear to be seamless, although the direct translation of the findings can be extremely challenging. Specifically, in the animal models, an implanted catheter facilitates delivery of very specific 5HT agonist compounds directly onto the spinal circuitry. The translation of this technique to humans is hindered by the lack of specific surgical techniques or available pharmacological agents directed towards 5HT receptor subtypes that are safe and effective for human clinical trials. However, oral administration of commonly available 5HTergic agents, such as selective serotonin reuptake inhibitors (SSRIs), may be a viable option to increase central 5HT concentrations in order to facilitate walking recovery in humans. Systematic quantification of how these SSRIs modulate human motor behaviors following SCI, with a specific focus on strength, reflexes, and the recovery of walking ability, are missing.This video demonstration is a progressive attempt to systematically and quantitatively assess the modulation of reflex activity, volitional strength and ambulation following the acute oral administration of an SSRI in human SCI. Agents are applied on single days to assess the immediate effects on motor function in this patient population, with long-term studies involving repeated drug administration combined with intensive physical interventions.

Methods to Quantify Pharmacologically Induced Alterations in Motor Function in Human Incomplete SCI

Spinal cord injury (SCI) is a debilitating disorder, which produces profound deficits in volitional motor control. Following medical stabilization, recovery from SCI typically involves long term rehabilitation. While recovery of walking ability is a primary goal in many patients early after injury, those with a motor incomplete SCI, indicating partial preservation of volitional control, may have the sufficient residual descending pathways necessary to attain this goal. However, despite physical interventions, motor impairments including weakness, and the manifestation of abnormal involuntary reflex activity, called spasticity or spasms, are thought to contribute to reduced walking recovery. Doctrinaire thought suggests that remediation of this abnormal motor reflexes associated with SCI will produce functional benefits to the patient. For example, physicians and therapists will provide specific pharmacological or physical interventions directed towards reducing spasticity or spasms, although there continues to be little empirical data suggesting that these strategies improve walking ability. In the past few decades, accumulating data has suggested that specific neuromodulatory agents, including agents which mimic or facilitate the actions of the monoamines, including serotonin (5HT) and norepinephrine (NE), can initiate or augment walking behaviors in animal models of SCI. Interestingly, many of these agents, particularly 5HTergic agonists, can markedly increase spinal excitability, which in turn also increases reflex activity in these animals. Counterintuitive to traditional theories of recovery following human SCI, the empirical evidence from basic science experiments suggest that this reflex hyper excitability and generation of locomotor behaviors are driven in parallel by neuromodulatory inputs (5HT) and may be necessary for functional recovery following SCI. The application of this novel concept derived from basic scientific studies to promote recovery following human SCI would appear to be seamless, although the direct translation of the findings can be extremely challenging. Specifically, in the animal models, an implanted catheter facilitates delivery of very specific 5HT agonist compounds directly onto the spinal circuitry. The translation of this technique to humans is hindered by the lack of specific surgical techniques or available pharmacological agents directed towards 5HT receptor subtypes that are safe and effective for human clinical trials. However, oral administration of commonly available 5HTergic agents, such as selective serotonin reuptake inhibitors (SSRIs), may be a viable option to increase central 5HT concentrations in order to facilitate walking recovery in humans. Systematic quantification of how these SSRIs modulate human motor behaviors following SCI, with a specific focus on strength, reflexes, and the recovery of walking ability, are missing. This video demonstration is a progressive attempt to systematically and quantitatively assess the modulation of reflex activity, volitional strength and ambulation following the acute oral administration of an SSRI in human SCI. Agents are applied on single days to assess the immediate effects on motor function in this patient population, with long-term studies involving repeated drug administration combined with intensive physical interventions.

Neuroendocrine effects of Citalopram, a selective serotonin re-uptake inhibitor, during lifespan in humans

Serotonergic system contributes to the regulation of hypothalamus-pituitary-adrenal axis. In humans, serotonergic agonists increase PRL, ACTH, and cortisol, while serotonin (5HT) influence on GH is controversial. Central 5HT activity and neuroendocrine function change during lifespan. To clarify the neuroendocrine response to 5HT across lifespan, we assessed ACTH, cortisol, DHEA, PRL, and GH responses to citalopram (CT) in young adults (YA) (no.=12, 29.2±1.7 yr mean±SEM), middle aged (MA) (no.=12, 54.3±0.9 yr), and elderly (ES) (no.=12, 69.3±0.9 yr) males. All the subjects received placebo (saline iv over 120 min) or CT (20 mg iv over 120 min). Blood samples were taken every 15 min up to 240 min. During placebo, ACTH, cortisol, GH, and PRL were similar in all groups while DHEA showed an age-dependent reduction from middle age (p<0.001). During CT, ACTH, and cortisol were higher than during placebo in YA (p<0.05) and even more in MA (p<0.01 vs placebo, p<0.05 vs YA); in ES, the increase of both ACTH and cortisol (p<0.05 vs placebo) was lower than in MA (p<0.05) and higher than in YA (p<0.05 for cortisol only). No changes were observed for DHEA, GH, and PRL in any group. Corticotrope response to CT is age-dependent in normal men, being amplified starting from middle age, suggesting precocious changes in the serotonergic neuroendocrine control during lifespan. CT is a useful tool to evaluate the age-dependent serotonergic function in humans.

5HT-1A receptors and anxiety-like behaviours: Studies in rats with constitutionally upregulated/downregulated serotonin transporter

Altered activity of brain serotonergic (5HT) system has been implicated in a wide range of behaviours and behavioural disorders, including anxiety. Functioning of 5HT-1A receptor has been suggested as a modulator of emotional balance in both, normal and pathological forms of anxiety. Here, we studied serotonergic modulation of anxiety-like behaviour using a genetic rat model with constitutional differences in 5HT homeostasis, named Wistar-Zagreb 5HT (WZ-5HT) rats. The model, consisting of high-5HT and low-5HT sublines, was developed by selective breeding of animals for extreme activities of peripheral (platelet) 5HT transporter, but selection process had affected also central 5HT homeostasis, as evidenced from neurochemical and behavioural studies.Anxiety-like behaviour in WZ-5HT rats was evaluated by two commonly used paradigms: open field and elevated-plus maze. The involvement of 5HT-1A receptors in behavioural response was assessed by measuring mRNA expression in cell bodies (raphe nuclei) and projection regions (frontal cortex, hippocampus) by use of RT-PCR and in situ hybridization, and by measuring functionality of cortical 5HT-1A receptors by use of [3H]8-OH-DPAT radioligand binding.Animals from the high-5HT subline exhibit increased anxiety-like behaviour and decreased exploratory activity when exposed to novel environment. No measurable differences in constitutional (baseline) functionality or expression of 5HT-1A receptors between sublines were found.The results support contribution of increased serotonergic functioning to the anxiety-like behaviour. They also validate the high-5HT subline of WZ-5HT rats as a potential model to study mechanisms of anxiety, especially of its nonpathological form, while the low-5HT subline may be useful to model sensation seeking phenotype.

Perinatal treatment

Abstract Serotonin (5-hydroxytryptamine, 5HT) regulates the development of 5HT neurons and target tissues during neurogenesis, while later it assumes the function of a neurotransmitter. Alterations in serotonin neurotransmission are indicated as biological substrates in several neuropsychiatric disorders, including autism. The most consistent 5HT-related finding in autistic disorder is hyperserotonemia, but the mechanism of its development and its relation to central 5HT dysfunction are still unclear. In an attempt to pharmacologically induce hyperserotonemia during the period of most intensive development of 5HT neurons, and to later investigate its effects on central 5HT functions, we have treated rats from gestational day 13 until postnatal day 21 with 2 mg/kg of the nonselective irreversible MAO inhibitor tranylcypromine (TCP). The control group received saline in the same manner. TCP treated rats displayed a long-lasting significant increase in platelet 5HT concentrations compared to the control rats. The TCP treated group had smaller litters, significantly lower pup survival rate, and slower weight gain during the post-weaning free-feeding period than the control group. Pups from the TCP group returned to their dams significantly slower than the control pups suggesting lower separation anxiety. Our results indicate that the perinatal treatment of rats with tranylcypromine has induced both, disregulation of the peripheral 5HT homeostasis and disturbances in central 5HT physiology in pups and young rats. The extent of the changes in the central serotonergic compartment in adult rats will be explored in our further studies.

Serotonin and bulimia nervosa.

Serotonin (5HT) is one of several neuromodulators of feeding. Experimentally reducing 5HT activity in animals increases food intake, while increasing 5HT activity has the opposite effect. Studies suggest that women with bulimia nervosa show signs of reduced 5HT activity, which may be related to binge eating. Data supporting the theory that reduced central nervous system 5HT activity may play a role in the pathophysiology of bulimia nervosa is reviewed. Disturbances of 5HT activity and the relationship to other psychopathology in bulimia nervosa, such as depression, substance abuse, and impulsivity, are also reviewed.

Expression of TRPV4 in the Stimulated Rat Oral Mucous Membrane-Nociceptive Mechanisms of Lingual Conical Papillae-

The study was supported by 2006-2007 Aid Program for Overseas Training of the Promotion and Mutual Aid Corporation for Private School of Japan and International Exchange Grant, Osaka Dental University. We studied the function of TRPV4 expression and its neuronal activation in response to noxious stimulation of oral mucosa. The intermolar region of dorsal lingual eminence (IDLE) of rats was stimulated with 10 microl of either normal saline or 5% formalin. Immunohistological studies of the TRPV4, pERK and serotonin (5HT) expression in designated regions of tongues and brainstems were performed for studying the descending pain modulatory system in response to nociception. Specimens of the experimental IDLE demonstrated a significant increase of TRPV4 activity in particular in stratum basale of conical papillae (p < 0.01). pERK-IR positive neurons were significantly increased in the RMg (p < 0.05), Sp5C (p < 0.05) and Md (p < 0.01); TRPV4-IR neurons were found to show a similar distribution with pERK-IR cells in the peripheral Sp5C (p < 0.05). A significant increase of 5HT expression was observed in the RMg (p < 0.01), RPa (p < 0.01) and ROb (p < 0.05). The results suggest that TRPV4 in the oral mucosa is nociceptor of peripheral hyperalgesia, and pERK expression in the Sp5C is closely related with central hyperalgesia of the nociception. Furthermore, pERK-IR cells of the central 5HT nervous system are activated to accelerate 5HT release for neuronal modulation of the descending pain modulatory system in response to nociception.

Central aminopeptidase and serotonin system activities: Possible relationship

The co-localization of serotonin (5-hydroxytryptamine, 5HT) and neuroactive peptides in the same neuron points to the importance of interactions between serotonergic and peptidergic systems in maintaining body homeostasis. In this work, we used an original genetic rat model to search for possible interrelations between 5HT system functioning and the activities of aminopeptidases, i.e. enzymes which are the key regulators of (neuro)peptides level/function. The activities of three cytosolic exopeptidases: alanyl aminopeptidase (alanyl-AP), arginyl aminopeptidase (arginyl-AP) and dipeptidyl peptidase III (DPP III) were measured in brains and peripheral tissues of the sublines of rats with constitutionally upregulated/downregulated 5HT transporter activity. These rat sublines, termed as high-5HT and low-5HT subline, have been obtained previously by selective breeding for the extreme values of platelet 5HT level and velocity of 5HT uptake. Besides in the periphery they show marked alterations also in brain 5HT function, indicating the differences in central 5HT transmission/homeostasis. In this study, we have found that animals from the high-5HT subline have significantly lower activity of brain alanyl-AP (p<0.05) and arginyl-AP (p<0.01) as compared to control animals. No other differences were noticed regardless of the 5HT subline, investigated organ or analyzed aminopeptidase. Results suggest that the constitutional upregulation of serotonergic activity may be related to a lowered brain cytosolic aminopeptidase activity which may have an influence on the cleavage of their physiological substrates.

Evidence of an enhanced central 5HT response in irritable bowel syndrome and in the rat maternal separation model

Efforts to define either a biomarker for irritable bowel syndrome (IBS) or a valid animal model have proven disappointing. The aims of this study were to determine if buspirone stimulates prolactin release through the 5-hydroxytryptamine (5HT)1a receptor and whether this response is altered in patients with IBS and in the rat maternal separation model. Buspirone (30 mg) was used to stimulate prolactin release in 40 patients with IBS and in 40 healthy controls. In study 1, 10 IBS patients and 10 controls underwent pretreatment with pindolol (5HT1a antagonist) or placebo followed by buspirone. In study 2, 30 patients with IBS and 30 healthy controls had prolactin release stimulated by buspirone. Maternally separated and nonseparated rats were also treated with buspirone and prolactin monitored. Serotonin metabolites were measured together with the expression of the 5HT1a and serotonin transporter (SERT) gene. Pindolol produced a dose-dependent decrease in the buspirone prolactin response. Patients with IBS and maternally separated rats showed an exaggerated release of prolactin in response to buspirone. In the animal model, an increased turnover of 5HT was found in the brainstem together with a trend toward increased activity of the SERT gene. In conclusion altered central serotonin responses are found in both IBS and in an animal model.

Modulation of neural response to happy and sad faces by acute tryptophan depletion

Central serotonin (5HT) plays a major role in emotional processing. We used functional neuroimaging (fMRI) to investigate the effects of experimental manipulation of central 5HT levels on the regional neural response to happy and sad facial expressions. Ten healthy participants (eight men and two women) were scanned during an implicit emotional processing task after receiving a tryptophan-free (acute tryptophan depletion, ATD) or a balanced amino acid drink in a double-blind design. ATD lowered total plasma tryptophan concentration by 80%. There was no significant effect on subjective mood ratings, on response accuracy and on reaction times. Compared to sham depletion, ATD attenuated activation in the right medial/inferior frontal gyrus, the posterior cingulate cortex, the occipital and parietal cortex bilaterally, the right hippocampus, claustrum and insula. Conversely, ATD was associated with relatively increased activation in the left inferior frontal gyrus. ATD had differential effects on activation during the processing of happy and sad faces in the right putamen and in the left superior temporal gyrus. In both cortical and sub-cortical regions, the neural response associated with processing emotional faces is significantly modulated by 5HT manipulation resulting from ATD. Moreover, in certain areas, this effect of 5HT depends on the emotional valence of the stimulus.