Serotonin Transporter mRNA Expression Research Articles

Serotonin Transporter mRNA Expression Is Reduced in the Peripheral Blood Mononuclear Cells of Subjects with Major Depression but Normal in Fibromyalgia.

Fibromyalgia (FM) and major depression disorder (MDD) frequently co-occur. Both disorders may share common serotonergic alterations, although there is less evidence of such alterations in FM. It is also unclear as to whether these alterations are persistent over time or transient. The objectives of this study were to (i) examine the changes in mRNA expression of serotonin transporter (SERT) on the surface of peripheral blood mononuclear cells (PBMCs) in FM, MDD, and the FM + MDD subjects compared to healthy controls, and to (ii) evaluate the effect of drug treatment on SERT expression. PBMCs were isolated from FM, MDD, FM + MDD, and control subjects. SERT expression was analyzed at the mRNA level via quantitative real-time polymerase chain reaction. Statistical analyses were performed using analyses of variance and linear mixed-effects models. SERT mRNA expression was significantly reduced in MDD subjects compared to controls (p < 0.001), but not in FM nor in FM + MDD subjects. Although the drug treatments improved symptoms in FM, MDD, and FM + MDD subjects, they had no significant effect on SERT mRNA expression. These results corroborate the role of the SERT in the pathophysiology of MDD, but not in FM, and show that the decreased mRNA expression of SERT is a persistent, rather than transient, phenomenon.

Expression Level Changes in Serotonin Transporter are Associated with Food Deprivation in the Pond Snail Lymnaea stagnalis.

In the pond snail Lymnaea stagnalis, serotonin (5-HT) plays an important role in feeding behavior and its associated learning (e.g., conditioned taste aversion: CTA). The 5-HT content in the central nervous system (CNS) fluctuates with changes in the nutritional status, but it is also expected to be influenced by changes in the serotonin transporter (SERT) expression level. In the present study, we identified SERT in Lymnaea and observed its localization in 5-HTergic neurons, including the cerebral giant cells (CGCs) in the cerebral ganglia and the pedal A cluster neurons and right and left pedal dorsal 1 neurons in the pedal ganglia by in situ hybridization. Real-time PCR revealed that the SERT mRNA expression level was lower under severe food deprivation than under mild food deprivation in the whole CNS as well as in a single CGC. These results inversely correlated with previous data that the 5-HT content in the CNS was higher in the severely food-deprived state than in the mildly food-deprived state. Furthermore, in single CGCs, we observed that the 5-HT level was significantly increased in the severely food-deprived state compared with the mildly food-deprived state. Our present findings suggest that changes in the SERT expression level associated with food deprivation may affect 5-HT signaling, probably contributing to learning and memory mechanisms in Lymnaea.

A three-pronged analysis confirms the association of the serotoninergic system with attention deficit hyperactivity disorder.

The serotonin transporter (SERT), encoded by the solute carrier family 6 number 4 (SLC6A4) gene, controls serotonin (5-HT) availability and is essential for the regulation of behavioral traits. Two SLC6A4 genetic variants, 5-HTTLPR and STin2, were widely investigated in patients with various neurobehavioral disorders, including attention deficit hyperactivity disorder (ADHD). We analyzed the association of the 5-HTTLPR (L/S) and STin2 (10/12) variants, plasma 5-HT, and 5-hydroxyindole acetic acid (5-HIAA), as well as SERT messenger RNA (mRNA) with ADHD in the eastern Indian subjects. Nuclear families with ADHD probands (n = 274) and ethnically matched controls (n = 367) were recruited following the Diagnostic and Statistical Manual of Mental Disorders. Behavioral traits, executive function, and intelligence quotient (IQ) of the probands were assessed using the Conner's Parent Rating Scale-Revised, Parental Account of Children's Symptoms (PACS), Barkley Deficit in Executive Functioning-Child and Adolescent Scale, and Wechsler Intelligence Scale for Children-III, respectively. After obtaining informed written consent, peripheral blood was collected to analyze genetic variants, plasma 5-HT, 5-HIAA, and SERT mRNA expression. ADHD probands showed a higher frequency of the 5-HTTLPR "L" allele and "L/L" genotype (P < 0.05), lower 5-HIAA level, and higher SERT mRNA expression. Scores for behavioral problems and hyperactivity were higher in the presence of the "S" allele and "S/S" genotype, while executive deficit was higher in the presence of the "L" allele. IQ score was lower in the presence of the STin2 "12" allele and L-12 haplotype. Data obtained indicate a significant association of the serotoninergic system with ADHD, warranting further in-depth investigation.

Thrombocytopenia after meta-iodobenzylguanidine (MIBG) therapy in neuroblastoma patients may be caused by selective MIBG uptake via the serotonin transporter located on megakaryocytes

BackgroundThe therapeutic use of [131I]meta-iodobenzylguanidine ([131I]MIBG) is often accompanied by hematological toxicity, primarily consisting of severe and persistent thrombocytopenia. We hypothesize that this is caused by selective uptake of MIBG via the serotonin transporter (SERT) located on platelets and megakaryocytes. In this study, we have investigated whether in vitro cultured human megakaryocytes are capable of selective plasma membrane transport of MIBG and whether pharmacological intervention with selective serotonin reuptake inhibitors (SSRIs) may prevent this radiotoxic MIBG uptake.MethodsPeripheral blood CD34+ cells were differentiated to human megakaryocytic cells using a standardized culture protocol. Prior to [3H]serotonin and [125I]MIBG uptake experiments, the differentiation status of megakaryocyte cultures was assessed by flow cytometry. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to assess SERT and NET (norepinephrine transporter) mRNA expression. On day 10 of differentiation, [3H]serotonin and [125I]MIBG uptake assays were conducted. Part of the samples were co-incubated with the SSRI citalopram to assess SERT-specific uptake. HEK293 cells transfected with SERT, NET, and empty vector served as controls.ResultsIn vitro cultured human megakaryocytes are capable of selective plasma membrane transport of MIBG. After 10 days of differentiation, megakaryocytic cell culture batches from three different hematopoietic stem and progenitor cell donors showed on average 9.2 ± 2.4 nmol of MIBG uptake per milligram protein per hour after incubation with 10–7 M MIBG (range: 6.6 ± 1.0 to 11.2 ± 1.0 nmol/mg/h). Co-incubation with the SSRI citalopram led to a significant reduction (30.1%—41.5%) in MIBG uptake, implying SERT-specific uptake of MIBG. A strong correlation between the number of mature megakaryocytes and SERT-specific MIBG uptake was observed.ConclusionOur study demonstrates that human megakaryocytes cultured in vitro are capable of MIBG uptake. Moreover, the SSRI citalopram selectively inhibits MIBG uptake via the serotonin transporter. The concomitant administration of citalopram to neuroblastoma patients during [131I]MIBG therapy might be a promising strategy to prevent the onset of thrombocytopenia.

Effects of Paeonia lactiflora Extract on Estrogen Receptor β, TPH2, and SERT in Rats with PMS Anxiety.

This study is to investigate the effect of Paeonia lactiflora extract on PMS anxiety and on expression of estrogen receptor β (ERβ), tryptophan hydroxylase-2 (TPH2), and serotonin transporter (SERT) in the premenstrual syndrome (PMS) anxiety model rats. The vaginal smear and open field test were used to screen rats in nonreception phase of estrus cycle with similar macroscopic behaviors and regular estrus cycle. PMS anxiety model rats were prepared by electrical stimulation. RT-PCR and immunofluorescence were used to measure the expression of ERβ, TPH2, and SERT. Compared with normal rats, the total distance in the open field test of the model rats was significantly increased (P < 0.05). The model rats showed nervous alertness, irritability, and sensitivity to external stimuli. After treatment with the Paeonia lactiflora extract, the total distance of rats was significantly reduced (P < 0.05). In reception stage, there was no significant difference in the mRNA and protein expression of ERβ, TPH2, and SERT. In nonreception stage, the expression of ERβ and TPH2 in the model group was significantly decreased (P < 0.05) as compared with the control group, but not SERT. Abnormal changes of the above indicators were reversed after the administration of the Paeonia lactiflora extract. In conclusion, Paeonia lactiflora extract can increase the expression of ERβ and TPH2 and decrease SERT in PMS model rats, which may be one of the mechanisms underlying the effect of Paeonia lactiflora extract on PMS.

Serotonin exerts a direct modulatory role on bladder afferent firing in mice.

Functional disorders (i.e. interstitial cystitis/painful bladder syndrome and irritable bowel syndrome) are associated with hyperexcitability of afferent nerves innervating the urinary tract and the bowel, respectively. Various non-5-HT3 receptor mRNA transcripts are expressed in mouse urothelium and exert functional responses to 5-HT. Whilst 5-HT3 receptors were not detected in mouse urothelium, 5-HT3 receptors expressed on bladder sensory neurons plays a role in bladder afferent excitability both under normal conditions and in a mouse model of chronic visceral hypersensitivity. These data suggest that the role 5-HT3 receptors play in bladder afferent signalling warrants further study as a potential therapeutic target for functional bladder disorders. Serotonin (5-HT) is an excitatory mediator that in the gastrointestinal (GI) tract plays a physiological role in gut-brain signalling and is dysregulated in functional GI disorders such as irritable bowel syndrome (IBS). Patients suffering from IBS frequently suffer from urological symptoms characteristic of interstitial cystitis/painful bladder syndrome, which manifests due to cross-sensitization of shared innervation pathways between the bladder and colon. However, a direct modulatory role of 5-HT in bladder afferent signalling and its role in colon-bladder neuronal crosstalk remain elusive. The aim of this study was to investigate the action of 5-HT on bladder afferent signalling in normal mice and mice with chronic visceral hypersensitivity (CVH) following trinitrobenzenesulfonic acid-induced colitis. Bladder afferent activity was recorded directly using ex vivo afferent nerve recordings. Expression of 14 5-HT receptor subtypes, the serotonin transporter (SERT) and 5-HT-producing enzymes was determined in the urothelium using RT-PCR. Retrograde labelling of bladder-projecting dorsal root ganglion neurons was used to investigate expression of 5-HT3 receptors using single cell RT-PCR, while sensory neuronal and urothelial responses to 5-HT were determined by live cell calcium imaging. 5-HT elicited bladder afferent firing predominantly via 5-HT3 receptors expressed on afferent terminals. CVH animals showed a downregulation of SERT mRNA expression in urothelium, suggesting increased 5-HT bioavailability. Granisetron, a 5-HT3 antagonist, reversed bladder afferent hypersensitivity in CVH mice. These data suggest 5-HT exerts a direct effect on bladder afferents to enhance signalling. 5-HT3 antagonists could therefore be a potential therapeutic target to treat functional bladder and bowel disorders.

Analysis of Expression and Functional Activity of Aromatic L-Amino Acid Decarboxylase (DDC) and Serotonin Transporter (SERT) as Potential Sources of Serotonin in Mouse Ovary.

The origin of serotonin in the ovary is the key question for understanding mechanisms of serotonergic regulation of reproductive function. We performed a study of the expression and functional activity of the serotonin transporter (SERT) and the enzyme for the synthesis of serotonin, aromatic l-amino acid decarboxylase (DDC) in mouse ovary. A pronounced peak of SERT mRNA expression occurs at the age of 14 days, but serotonin synthesis enzymes are expressed at the maximum level in the ovaries of newborn mice. SERT is detected immunohistochemically in all cellular compartments of the ovary with a maximum level of immunostaining in the oocytes of growing ovarian follicles. DDC immunolocalization, in contrast, is detected to a greater extent in primordial follicle oocytes, and decreases at the later stages of folliculogenesis. Serotonin synthesis in all cellular compartments occurs at very low levels, whereas specific serotonin uptake is clearly present, leading to a significant increase in serotonin content in the oocytes of growing primary and secondary follicles. These data indicate that the main mechanism of serotonin accumulation in mouse ovary is its uptake by the specific SERT membrane transporter, which is active in the oocytes of the growing ovarian follicles.

Does fluoxetine exposure affect hypoxia tolerance in the Gulf toadfish, Opsanus beta?

Due to ineffective wastewater treatment technologies, pharmaceuticals such as the selective serotonin reuptake inhibitors (SSRIs)—a common class of antidepressants which inhibit the serotonin transporter (SERT)—can be found in surface waters and marine receiving waters near wastewater effluents. Understanding how exposure to these chemicals might impact non-target organisms, especially combined with other environmental stressors like hypoxia, is essential in order to thoroughly evaluate environmental risk. It was hypothesized that both acute and chronic exposure to the SSRI fluoxetine (FLX) would interfere with the metabolic hypoxia response of the Gulf toadfish, Opsanus beta. Here we demonstrate that acute intraperitoneal treatment with 50 μg g−1 FLX significantly reduces the regulation index, or degree of metabolic regulation, in toadfish. Acute FLX exposure significantly reduced SERT mRNA expression in the first and third gill arches, but mRNA expression was not affected in heart tissues or in the second gill arch. In contrast, the regulation index was unaffected by 14–17 day waterborne FLX exposure to environmentally relevant (0.01 μg L−1) and approximately 1000-fold higher (8.5 μg L−1) concentrations. However, the higher concentration was sufficient to induce a systemic elevation in plasma serotonin concentrations. Chronic FLX exposure did not alter SERT mRNA expression in heart or gill tissues. The results of this study implicate the involvement of 5-HT pathways in hypoxia tolerance but demonstrate that current environmental levels of FLX are insufficient to impair the metabolic hypoxia response in marine fish.

Reduced Serotonin Transporter Levels and Inflammation in the Midbrain Raphe of 12 Month Old APPswe/PSEN1dE9 Mice.

Although mood and sleep disturbances are nearly universal among patients with Alzheimer's disease (AD), brain structures involved in non-cognitive processing remain under characterized in terms of AD pathology. This study was designed to evaluate hallmarks of AD pathology in the brainstem of the APPswe/PS1dE9 mouse model of familial AD. Fresh-frozen sections from female, 12 month old, transgenic and control B6C3 mice (n=6/genotype) were examined for amyloid burden and neurofibrillary alterations, by using 6E10 immunohistochemistry and the Gallyas silver stain, respectively. Serotonin transporter (SERT) densities in the dorsal and the median raphe were quantified by [3H]DASB autoradiography. SERT mRNA expression was measured by RT-PCR and visualized by in situ hybridization. Neuroinflammation was evaluated by immunohistochemical staining for microglia and astrocytes, and by measuring mRNA levels of the proinflammatory cytokines TNF-α, IL-1β and IL-6. No amyloid- and tau-associated lesions were observed in the midbrain raphe of 12 month old APPswe/PS1dE9 mice. SERT binding levels were reduced in transgenic animals compared to age-matched controls, and SERT mRNA levels were decreased by at least 50% from control values. Intense microglial, but not astrocytic immunoreactivity was observed in APPswe/PS1dE9 vs. wild-type mice. Levels of TNF-α mRNA were two-fold higher than control and correlated positively with SERT mRNA expression levels in transgenic animals. There was no amyloid accumulation and tau-associated pathology in the midbrain raphe of 12 month old APPswe/PS1dE9 mice. However, there was a local neuroinflammatory response with loss of serotonergic markers, which may partially account for some of the behavioral symptoms of AD.

Effect of Lactobacillus rhamnosus GG supernatant on serotonin transporter expression in rats with post-infectious irritable bowel syndrome.

AIMTo evaluate the effect of Lactobacillus rhamnosus GG supernatant (LGG-s) on the expression of serotonin transporter (SERT) in rats with post-infectious irritable bowel syndrome (PI-IBS).METHODSCampylobacter jejuni 81-176 (1010 CFU/mL) was used to induce intestinal infection to develop a PI-IBS model. After evaluation of the post-infectious phase by biochemical tests, DNA agarose gel electrophoresis, abdominal withdrawal reflex (AWR) test, and the intestinal motility test, four PI-IBS groups received different concentrations of LGG-s for 4 wk. The treatments were maintained for 1.0, 2.0, 3.0 or 4.0 wk during the experiment, and the colons and brains were removed for later use each week. SERT mRNA and protein levels were detected by real-time PCR and Western blot, respectively.RESULTSThe levels of SERT mRNA and protein in intestinal tissue were higher in rats treated with LGG-s than in control rats and PI-IBS rats gavaged with PBS during the whole study. Undiluted LGG-s up-regulated SERT mRNA level by 2.67 times compared with the control group by week 2, and SERT mRNA expression kept increasing later. Double-diluted LGG-s was similar to undiluted-LGG-s, resulting in high levels of SERT mRNA. Triple-diluted LGG-s up-regulated SERT mRNA expression level by 6.9-times compared with the control group, but SERT mRNA expression decreased rapidly at the end of the second week. At the first week, SERT protein levels were basically comparable in rats treated with undiluted LGG-s, double-diluted LGG-s, and triple-diluted LGG-s, which were higher than those in the control group and PBS-treated PI-IBS group. SERT protein levels in the intestine were also comparable in rats treated with undiluted LGG-s, double-diluted LGG-s, and triple-diluted LGG-s by the second and third weeks. SERT mRNA and protein levels in the brain had no statistical difference in the groups during the experiment.CONCLUSIONLGG-s can up-regulate SERT mRNA and protein levels in intestinal tissue but has no influence in brain tissue in rats with PI-IBS.

PT614. Altered Serotonin transporter mRNA expression in the peripheral leukocytes of obsessive-compulsive disorder

PT614. Altered Serotonin transporter mRNA expression in the peripheral leukocytes of obsessive-compulsive disorder

Traumatic brain injury decreases serotonin transporter expression in the rat cerebrum

Objectives: An association has been postulated between traumatic brain injury (TBI) and depression. The serotonin transporter (SERT) regulates the concentration of serotonin in the synaptic cleft and represents a molecular target for antidepressants. We hypothesized that SERT expression in the brain changes following TBI.Methods: We performed immunohistochemistry, real-time polymerase chain reaction analysis for mRNA and western blot analysis for protein to examine the time-dependent changes in SERT expression in the cerebrum during the first 14 days after TBI, using a controlled cortical impact model in rats.Results: SERT immunoreactivity in neuronal fibres within the area adjacent to the cortical contusion decreased 1 to 14 days after injury. Significantly decreased SERT mRNA and protein expression were noted in the area adjacent to the cortical contusion 7 days after injury. There were no significant changes in SERT expression in the cingulum of the injured brain.Discussion: The findings of this study indicate that TBI decreases SERT expression in the cerebral cortex. The decreased levels of SERT expression after TBI may result in decreased serotonin neurotransmission in the brain and indicate a possible relationship with depression following TBI.

RNA‐Seq Analysis Reveals Ectopic Expression of Truncated Non‐Functional Serotonin Transporter in the Hippocampus of SERT‐KO Mice

The serotonergic system innervates the cortex, entorhinal cortex, hippocampus (HPC), nucleus accumbens, amygdala and anterior hypothalamus via serotonergic neurons arising from raphe nuclei in the mid and hindbrain. These structures are part of the limbic system, which is involved in emotions and motivation. Termini of the serotonergic neurons that innervate these limbic regions contain the serotonin transporter (SERT), which is a therapeutic target for many mood disorders. To date, insight into the link between SERT and its impact on these psychiatric conditions has been afforded by the use of a SERT‐deficient mouse line (SERT‐KO). Numerous studies have used this murine model to investigate serotonergic function in the HPC and links to neuropsychiatric disorders. In the present study, RNA‐Seq was performed on isolated HPC from 9 week old male C57BL6/J (WT) and B6.129(Cg)‐Slc6a4tm1Kpl/J (SERT‐KO) mice yielding 28–34 million mappable reads, indicating exceptional sequencing depth of the mouse genome. Reads were aligned to the mouse genome (mm10) using Tophat v2.0.13 and fragments were assigned to genes using Feature Counts v1.4.6. Differential gene expression analysis was performed with R/Bioconductor package DESeq2 v1.10.1 and analyzed with Ingenuity Pathway Analysis (IPA, Qiagen). Unexpectedly, an mRNA species capable of coding for amino acids 115 to 630 of SERT was detected in KO HPC at levels significantly higher than full‐length SERT in WT HPC samples (219 vs. 2 fragment counts, respectively). The truncated SERT transcript parallels the genetic mutation used to create the SERT‐KO mouse where exon 2 of the promoter region was replaced with a PGK neomycin‐polyA cassette. The altered 5′‐UTR region in the SERT‐KO mice appears to allow for ectopic expression of truncated SERT mRNA in the cell bodies of pyramidal, granule and/or glial cells residing in the HPC. Expression of SERT mRNA in the HPC of WT and KO mice was normalized to the reference gene Beta‐2‐microglobulin (B2M), revealing a 155‐fold increase in SERT mRNA in the KO mouse vs WT. IPA analysis of genes showing significant expression changes identified a number of genes involved in the “Unfolded Protein Response (UPR)” and “Endoplasmic Reticulum Stress” pathways consistent with ectopic expression of the truncated SERT mRNA which does not code for transmembrane domain 1 of SERT and likely result in mis‐ or unfolded protein. In addition, IPA identified the “Circadian Rhythm Signaling” pathway in line with previous reports looking at changes in circadian rhythm and REM sleep in SERT‐KO mice. IPA “upstream” analysis also suggests that expression changes of several genes were linked to brain‐derived neurotrophic factor (BDNF) and whereas BDNF expression changes were detected between WT and SERT‐KO samples, the difference did not reach significance. However, this analysis does not discriminate isoforms of BDNF, an area of future focus. Finally, although our study focuses on the HPC, other tissues including those outside the central nervous system could be affected by inappropriate SERT expression.Support or Funding InformationNIH‐NIGMS P20 GM104360

Depression-like behaviour of MOAP-1-/- mice - altered serotonergic mechanisms

Event Abstract Back to Event Depression-like behaviour of MOAP-1-/- mice - altered serotonergic mechanisms Hui Zhao1*, Nur-Ezan Mohamed1, Su Jing Chan1, Victor C. Yu2 and Peter T. Wong1 1 National University of Singapore, Department of Pharmacology, Yong Loo Lin School of Medicine,, Singapore 2 National University of Singapore , Department of Pharmacy, Faculty of Science, Singapore Modulator of apoptosis 1 (MOAP-1), as an upstream protein of Bax, plays a key role in mediating Bax function. Bax KO mice have been reported to show depression-like behaviour. Our studies on MOAP-1-/- (KO) mice show that they exhibited similar depression-like behaviour in the force swimming test (FST) and tail suspension test (TST), when compared to wildtype (WT) controls. Here we report new findings that suggest altered serotonergic mechanisms in the MOAP-1 KO mice may underlie the depression-like behaviour. When the mice were subjected to stress in the form of 3 successive daily exposure to water, the level of expression of the tryptophan hydroxylase 2 (TPH2, the rate-limiting enzyme in the biosynthesis of serotonin) protein was significantly lower in the midbrain region of the KO mice when compared to the WT controls. Consistently, the number of TPH2-positive cells in the dorsal raphe nucleus (DRN) were observed to be significantly less in the KO mice using immunofluorescent staining. These results suggest that the KO mice were less able to respond to stress by increases in their central serotonergic activities. In support, we observed decreased plasma serotonin (5-HT) levels in KO mice when compared with WT mice. On the contrary, it is interesting to note that SERT (serotonin transporter), 5-HT1A, and 5-HT2A receptor mRNA expression levels in the midbrain remained unchanged. However, acute treatment with a single dose of the serotonin selective reuptake inhibitor fluoxetine decreased the immobility time (in the FST) of the KO mice, while that of the WT mice remained unchanged. Taken together, our findings strongly suggest that the absence of MOAP-1 in the KO mice resulted in a reduced capacity for the mice to adapt to stress through increases in central serotonergic activities. Keywords: Depression, behaviour, mouse model, midbrain, serotonergic Conference: 14th Meeting of the Asian-Pacific Society for Neurochemistry, Kuala Lumpur, Malaysia, 27 Aug - 30 Aug, 2016. Presentation Type: Poster Presentation Session Topic: 14th Meeting of the Asian-Pacific Society for Neurochemistry Citation: Zhao H, Mohamed N, Chan S, Yu VC and Wong PT (2016). Depression-like behaviour of MOAP-1-/- mice - altered serotonergic mechanisms. Conference Abstract: 14th Meeting of the Asian-Pacific Society for Neurochemistry. doi: 10.3389/conf.fncel.2016.36.00132 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 04 Aug 2016; Published Online: 11 Aug 2016. * Correspondence: Ms. Hui Zhao, National University of Singapore, Department of Pharmacology, Yong Loo Lin School of Medicine,, Singapore, Singapore, Singapore, a0123672@u.nus.edu Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Hui Zhao Nur-Ezan Mohamed Su Jing Chan Victor C Yu Peter T Wong Google Hui Zhao Nur-Ezan Mohamed Su Jing Chan Victor C Yu Peter T Wong Google Scholar Hui Zhao Nur-Ezan Mohamed Su Jing Chan Victor C Yu Peter T Wong PubMed Hui Zhao Nur-Ezan Mohamed Su Jing Chan Victor C Yu Peter T Wong Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Altered intestinal neuroendocrine gene expression in humans with obesity.

Gastrointestinal hormones are critically involved in the regulation of food intake and body weight. Previous studies support an interplay between gastrointestinal hormones and the serotonergic system. This study explored intestinal neuroendocrine expression patterns in humans with obesity versus nonobese humans. Jejunum samples were collected from 164 humans with obesity (120 women; BMI (mean ± SD): 43.5 ± 6.6 kg/m(2) ) while they underwent Roux-en-Y gastric bypass surgery and from 18 nonobese humans (7 women; BMI: 23.5 ± 3.0 kg/m(2) ) undergoing distinct intestinal surgeries. mRNA expression of cholecystokinin (CCK), peptide YY3-36 (PYY), nesfatin1, ghrelin, ghrelin O-acyltransferase (GOAT), leptin, leptin receptor (leptinR), glucagon-like-peptide 1 receptor (GLP1R), serotonin transporter (SERT), tryptophan hydroxylase 1 (TPH1), and serotonin receptor 3A (5HT3A R) was determined with qRT-PCR. Ghrelin and GOAT protein expression was quantified using immunohistological stainings. Statistical analyses were performed with SPSS. Jejunum samples from humans with obesity showed a higher expression of GOAT (mRNA and protein), TPH1, and SERT mRNA compared with the nonobese humans (all P < 0.05). Positive correlations were observed between TPH1, CCK, PYY, and nesfatin1 in nonobese and GOAT, ghrelin, TPH1, SERT, CCK, and PYY in humans with obesity (all P < 0.01). Our top-down approach substantiates the dysregulation of jejunal neuroendocrine hormones in obesity.

Lactobacillus rhamnosus GG supernatant upregulates serotonin transporter expression in intestinal epithelial cells and mice intestinal tissues.

The role that probiotics play in relieving irritable bowel syndrome (IBS) has been demonstrated; however, the mechanism by which IBS is affected remains unclear. In this study, serotonin transporter (SERT) mRNA and serotonin transporter protein (SERT-P) levels in HT-29, Caco-2 cells, and mice intestinal tissues were examined after treatment with Lactobacillus rhamnosus GG supernatant (LGG-s). HT-29 and Caco-2 cells were treated with different concentrations of LGG-s for 12 and 24h and C57BL/6 mice received supplements of different concentrations for 4weeks. SERT mRNA and SERT-P levels were detected by real-time PCR and Western blotting. SERT mRNA and SERT-P levels in HT-29 and Caco-2 cells were higher than those in the control 24h after treatment. Undiluted LGG-s upregulated SERT mRNA levels by 9.4-fold in the first week, which dropped in the second week. The double-diluted LGG-s upregulated SERT mRNA by 2.07-fold in the first week; levels dropped to 1.75-fold within the second week and under base expression levels by the third week, while they again climbed to 1.56-fold in the fourth week. The triple-diluted LGG-s could not upregulate SERT mRNA expression until the end of the fourth week. The SERT-P levels in the double-diluted LGG-s group were higher than that in the control but fluctuated with time. SERT-P levels in the triple-diluted LGG-s were higher than that in the control in the last 2weeks and increased with time. LGG-s can upregulate SERT mRNA and SERT-P levels in intestinal epithelial cells and mice intestinal tissues.

Methotrexate causes a change in intestinal 5-hydroxytryptamine metabolism in rats

The effects of methotrexate on 5-hydroxytryptamine (5-HT) metabolism in the intestinal tissue of rats were investigated during the delayed phase after a single administration. Rats were i.p. injected with methotrexate or with saline as a control, and kaolin and food intakes were measured by an automatic monitoring apparatus. At 96h after administration, dissected-out ileal tissue was frozen rapidly in liquid nitrogen for further analysis or fixed for immunohistochemical staining. Methotrexate at a dose of 50mg/kg caused a time-dependent increase in kaolin intake lasting up to 72h after administration, which returned to the control level at 96h after administration. This dose of methotrexate caused a gradual decrease in body weight, food intake, and water intake lasting up to 72h, which approached the control level at 96h. Methotrexate caused pathologic changes, including a moderate inflammatory response in the ileal tissue and an increase in the number of l-tryptophan hydroxylase (TPH)-expressing cells in the ileal mucosa. Methotrexate also caused a significant increase in 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) content and in TPH1 mRNA expression in the ileal tissues. It had no significant effects on mRNA expression of serotonin transporter, COX-1, or COX-2 or on myeloperoxidase activity. This study demonstrated, for the first time, that methotrexate caused a change in the ileal 5-HT metabolism associated with hyperplasia of mucosal enterochromaffin cells.

Effects of genetic and early environmental risk factors for depression on serotonin transporter expression and methylation profiles.

The serotonin transporter (SERT) gene-linked polymorphic region (5-HTTLPR) has been implicated in moderating the link between life stress and depression. However, respective molecular pathways of gene–environment (GxE) interaction are largely unknown. Sustained alterations in SERT gene expression profiles, possibly mediated by epigenetic modifications, are a frequent correlate of depression and may thus constitute a putative mediator of GxE interaction. Here, we aimed to investigate joint effects of 5-HTTLPR and self-reported environmental adversity throughout the lifespan (prenatal, early and recent stress/trauma) on in vivo SERT mRNA expression in peripheral blood cells. To further evaluate whether environmentally induced changes in SERT expression are mediated by epigenetic modifications, we analyzed 83 CpG sites within a 799-bp promoter-associated CpG island of the SERT gene using the highly sensitive method of bisulfite pyrosequencing. Participants were 133 healthy young adults. Our findings show that both the 5-HTTLPR S allele and maternal prenatal stress/child maltreatment are associated with reduced in vivo SERT mRNA expression in an additive manner. Remarkably, individuals carrying both the genetic and the environmental risk factors exhibited 32.8% (prenatal stress) and 56.3% (child maltreatment) lower SERT mRNA levels compared with those without any risk factor. Our data further indicated that changes in SERT mRNA levels were unlikely to be mediated by DNA methylation profiles within the SERT CpG island. It is thus conceivable that the persistent changes in SERT expression may in turn relate to altered serotonergic functioning and possibly convey differential disease vulnerability associated with 5-HTTLPR and early adversity.

Distinctive behavioral and cellular responses to fluoxetine in the mouse model for Fragile X syndrome.

Fluoxetine is used as a therapeutic agent for autism spectrum disorder (ASD), including Fragile X syndrome (FXS). The treatment often associates with disruptive behaviors such as agitation and disinhibited behaviors in FXS. To identify mechanisms that increase the risk to poor treatment outcome, we investigated the behavioral and cellular effects of fluoxetine on adult Fmr1 knockout (KO) mice, a mouse model for FXS. We found that fluoxetine reduced anxiety-like behavior of both wild-type and Fmr1 KO mice seen as shortened latency to enter the center area in the open field test. In Fmr1 KO mice, fluoxetine normalized locomotor hyperactivity but abnormally increased exploratory activity. Reduced brain-derived neurotrophic factor (BDNF) and increased TrkB receptor expression levels in the hippocampus of Fmr1 KO mice associated with inappropriate coping responses under stressful condition and abolished antidepressant activity of fluoxetine. Fluoxetine response in the cell proliferation was also missing in the hippocampus of Fmr1 KO mice when compared with wild-type controls. The postnatal mRNA expression of serotonin transporter (SERT) was reduced in the thalamic nuclei of Fmr1 KO mice during the time of transient innervation of somatosensory neurons suggesting that developmental changes of SERT expression were involved in the differential cellular and behavioral responses to fluoxetine in wild-type and Fmr1 mice. The results indicate that changes of BDNF/TrkB signaling contribute to differential behavioral responses to fluoxetine among individuals with ASD.

N-ethylmaleimide-sensitive factor interacts with the serotonin transporter and modulates its trafficking: implications for pathophysiology in autism.

BackgroundChanges in serotonin transporter (SERT) function have been implicated in autism. SERT function is influenced by the number of transporter molecules present at the cell surface, which is regulated by various cellular mechanisms including interactions with other proteins. Thus, we searched for novel SERT-binding proteins and investigated whether the expression of one such protein was affected in subjects with autism.MethodsNovel SERT-binding proteins were examined by a pull-down system. Alterations of SERT function and membrane expression upon knockdown of the novel SERT-binding protein were studied in HEK293-hSERT cells. Endogenous interaction of SERT with the protein was evaluated in mouse brains. Alterations in the mRNA expression of SERT (SLC6A4) and the SERT-binding protein in the post-mortem brains and the lymphocytes of autism patients were compared to nonclinical controls.ResultsN-ethylmaleimide-sensitive factor (NSF) was identified as a novel SERT-binding protein. NSF was co-localized with SERT at the plasma membrane, and NSF knockdown resulted in decreased SERT expression at the cell membranes and decreased SERT uptake function. NSF was endogenously co-localized with SERT and interacted with SERT. While SLC6A4 expression was not significantly changed, NSF expression tended to be reduced in post-mortem brains, and was significantly reduced in lymphocytes of autistic subjects, which correlated with the severity of the clinical symptoms.ConclusionsThese data clearly show that NSF interacts with SERT under physiological conditions and is required for SERT membrane trafficking and uptake function. A possible role for NSF in the pathophysiology of autism through modulation of SERT trafficking, is suggested.