A Conserved Intramolecular Ion-Pair Plays a Critical but Divergent Role in Regulation of Dimerization and Transport Function among the Monoamine Transporters.

The monoamine transporters, including the serotonin transporter (SERT), dopamine transporter (DAT), and norepinephrine transporter (NET), are the therapeutic targets for the treatment of many neuropsychiatric disorders. Despite significant progress in characterizing the structures and transport mechanisms of these transporters, the regulation of their transport functions through dimerization or oligomerization remains to be understood. In the present study, we identified a conserved intramolecular ion-pair at the third extracellular loop (EL3) connecting TM5 and TM6 that plays a critical but divergent role in the modulation of dimerization and transport functions among the monoamine transporters. The disruption of the ion-pair interactions by mutations induced a significant spontaneous cross-linking of a cysteine mutant of SERT and an increase in cell surface expression but with an impaired specific transport activity. On the other hand, similar mutations of the corresponding ion-pair residues in both DAT and NET resulted in an opposite effect on their oxidation-induced dimerization, cell surface expression, and transport function. Reversible biotinylation experiments indicated that the ion-pair mutations slowed down the internalization of SERT but stimulated the internalization of DAT. In addition, cysteine accessibility measurements for monitoring SERT conformational changes indicated that substitution of the ion-pair residues resulted in profound effects on the rate constants for cysteine modification in both the extracellular and cytoplasmatic substrate permeation pathways. Furthermore, molecular dynamics simulations showed that the ion-pair mutations increased the interfacial interactions in a SERT dimer but decreased it in a DAT dimer. Taken together, we propose that the transport function is modulated by the equilibrium between monomers and dimers on the cell surface, which is regulated by a potential compensatory mechanism but with different molecular solutions among the monoamine transporters. The present study provided new insights into the structural elements regulating the transport function of the monoamine transporters through their dimerization.

Serotonin transporter knockdown relieves depression-like behavior and ethanol-induced CPP in mice after chronic social defeat stress.

Patients with stress-triggered major depression disorders (MDD) can often seek comfort or temporary relief through alcohol consumption, as they may turn to it as a means of self-medication or coping with overwhelming emotions. The use of alcohol as a coping mechanism for stressful events can escalate, fostering a cycle where the temporary relief it provides from depression can deepen into alcohol dependence, exacerbating both conditions. Although, the specific mechanisms involved in stress-triggered alcohol dependence and MDD comorbidities are not well understood, a large body of literature suggests that the serotonin transporter (SERT) plays a critical role in these abnormalities. To further investigate this hypothesis, we used a lentiviral-mediated knockdown approach to examine the role of hippocampal SERT knockdown in social defeat stress-elicited depression like behavior and ethanol-induced place preference (CPP). The results showed that social defeat stress-pro depressant effects were reversed following SERT knockdown demonstrated by increased sucrose preference, shorter latency to feed in the novelty suppressed feeding test, and decreased immobility time in the tail suspension and forced swim tests. Moreover, and most importantly, social stress-induced ethanol-CPP acquisition and reinstatement were significantly reduced following hippocampal SERT knockdown using short hairpin RNA shRNA-expressing lentiviral vectors. Finally, we confirmed that SERT hippocampal mRNA expression correlated with measures of depression- and ethanol-related behaviors by Pearson's correlation analysis. Taken together, our data suggest that hippocampal serotoninergic system is involved in social stress-triggered mood disorders as well as in the acquisition and retrieval of ethanol contextual memory and that blockade of this transporter can decrease ethanol rewarding properties.

Alterations in the brain serotonin system and serotonin-regulated behavior during aging in zebrafish males and females.

The brain serotonin (5-HT) system performs a neurotrophic function and supports the plasticity of the nervous system, while its age-related changes can increase the risk of senile neurodegeneration. Zebrafish brain is highly resistant to damage and neurodegeneration due to its high regeneration potential and it is a promising model object in searching for molecular factors preventing age-related neurodegeneration. In the present study alterations in 5-HT-related behavior in the home tank and the novel tank diving test, as well as 5-HT, 5-HIAA levels, tryptophan hydroxylase (TPH), monoamine oxidase (MAO) activity and the expression of genes encoding TPH, MAO, 5-HT transporter and 5-HT receptors in the brain of 6, 12, 24 and 36 month old zebrafish males and females are investigated. Marked sexual dimorphism in the locomotor activity in the novel tank test is revealed: females of all ages move slower than males. No sexual dimorphism in 5-HT-related traits is observed. No changes in 5-HT and 5-HIAA levels in zebrafish brain during aging is observed. At the same time, the aging is accompanied by a decrease in the locomotor activity, TPH activity, tph2 and htr1aa genes expression as well as an increase in the MAO activity and slc6a4a gene expression in their brain. These results indicate that the brain 5-HT system in zebrafish is resistant to age-related alterations.

Gene-Environment Interactions in Irrational Beliefs: The Roles of Childhood Adversity and Multiple Candidate Genes.

Cognitive behavioral therapy is based on the view that maladaptive thinking is the causal mechanism of mental disorders. While this view is supported by extensive evidence, very limited work has addressed the factors that contribute to the development of maladaptive thinking. The present study aimed to uncover interactions between childhood maltreatment and multiple genetic differences in irrational beliefs. Childhood maltreatment and irrational beliefs were assessed using multiple self-report instruments in a sample of healthy volunteers (N = 452). Eighteen single-nucleotide polymorphisms were genotyped in six candidate genes related to neurotransmitter function (COMT; SLC6A4; OXTR), neurotrophic factors (BDNF), and the hypothalamic-pituitary-adrenal axis (NR3C1; CRHR1). Gene-environment interactions (G×E) were first explored in models that employed one measure of childhood maltreatment and one measure of irrational beliefs. These effects were then followed up in models in which either the childhood maltreatment measure, the irrational belief measure, or both were substituted by parallel measures. Consistent results across models indicated that childhood maltreatment was positively associated with irrational beliefs, and these relations were significantly influenced by COMT rs165774 and OXTR rs53576. These results remain preliminary until independent replication, but they represent the best available evidence to date on G×E in a fundamental mechanism of psychopathology.

Serotonin Transporter Deficiency Induces Metabolic Alterations in the Ileal Mucosa.

Serotonin transporter (SERT) deficiency has been implicated in metabolic syndrome, intestinal inflammation, and microbial dysbiosis. Interestingly, changes in microbiome metabolic capacity and several alterations in host gene expression, including lipid metabolism, were previously observed in SERT-/- mice ileal mucosa. However, the precise host or microbial metabolites altered by SERT deficiency that may contribute to the pleiotropic phenotype of SERT KO mice are not yet understood. This study investigated the hypothesis that SERT deficiency impacts lipid and microbial metabolite abundances in the ileal mucosa, where SERT is highly expressed. Ileal mucosal metabolomics was performed by Metabolon on wild-type (WT) and homozygous SERT knockout (KO) mice. Fluorescent-activated cell sorting (FACS) was utilized to measure immune cell populations in ileal lamina propria to assess immunomodulatory effects caused by SERT deficiency. SERT KO mice exhibited a unique ileal mucosal metabolomic signature, with the most differentially altered metabolites being lipids. Such changes included increased diacylglycerols and decreased monoacylglycerols in the ileal mucosa of SERT KO mice compared to WT mice. Further, the ileal mucosa of SERT KO mice exhibited several changes in microbial-related metabolites known to play roles in intestinal inflammation and insulin resistance. SERT KO mice also had a significant reduction in the abundance of ileal group 3 innate lymphoid cells (ILC3). In conclusion, SERT deficiency induces complex alterations in the ileal mucosal environment, indicating potential links between serotonergic signaling, gut microbiota, mucosal immunity, intestinal inflammation, and metabolic syndrome.

Identification of the potassium-binding site in serotonin transporter.

Clearance of serotonin (5-hydroxytryptamine, 5-HT) from the synaptic cleft after neuronal signaling is mediated by serotonin transporter (SERT), which couples this process to the movement of a Na+ ion down its chemical gradient. After release of 5-HT and Na+ into the cytoplasm, the transporter faces a rate-limiting challenge of resetting its conformation to be primed again for 5-HT and Na+ binding. Early studies of vesicles containing native SERT revealed that K+ gradients can provide an additional driving force, via K+ antiport. Moreover, under appropriate conditions, a H+ ion can replace K+. Intracellular K+ accelerates the resetting step. Structural studies of SERT have identified two binding sites for Na+ ions, but the K+ site remains enigmatic. Here, we show that K+ antiport can drive substrate accumulation into vesicles containing SERT extracted from a heterologous expression system, allowing us to study the residues responsible for K+ binding. To identify candidate binding residues, we examine many cation binding configurations using molecular dynamics simulations, predicting that K+ binds to the so-called Na2 site. Site-directed mutagenesis of residues in this site can eliminate the ability of both K+ and H+ to drive 5-HT accumulation into vesicles and, in patch clamp recordings, prevent the acceleration of turnover rates and the formation of a channel-like state by K+ or H+. In conclusion, the Na2 site plays a pivotal role in orchestrating the sequential binding of Na+ and then K+ (or H+) ions to facilitate 5-HT uptake in SERT.

[The association of polymorphisms of the serotonin transporter gene SLC6A4 with depression]. / Svyaz' polimorfizmov gena serotoninovogo transportera SLC6A4 s depressiei.

OBJECTIVE: To study the relationship of polymorphic variants of the SLC6A4 gene with depression among people aged 25-44 years in Novosibirsk. MATERIAL AND METHODS: Under the WHO program «MONICA-psychosocial (MOPSY)¼, a random representative sample of people aged 25-44 years from the population of the Oktyabrsky district of Novosibirsk (men n=725, mean age 43.4±0.4 years, response - 71.3%, women n=710, mean age 44.8±0.4 years, response - 72%). Depression was assessed using the MONICA-MOPSY psychosocial questionnaire. Every fourth respondent was examined for polymorphic variants of 5HTTLPR-VNTR SNP rs25531 A>G of the SLC6A4 gene. The study was carried out within the framework of the budget topic Reg. No. 122031700094-5. RESULTS: The high level of depression among people aged 25-44 was 12.8% (for men 9.1%, for women - 15.92%); the average level of depression occurred in 24.5% of the population (among men in 21.24%, among women in 26.76%) (χ2=17.071, df=2, p<0.001). The most common genotype of the SLC6A4 gene, among people aged 25--4 years old in Novosibirsk, was SLA - 43.29%, LALA - 26.53% - in second place, SS - 17.87% - third, LALG - 6 genotypes were less represented genotypes. 74%, SLG - 4.18%, LGLG - 1.39%. Carrying the SLA genotype (53.3% and 63.6%) increased the chance of developing both the average level of depression by 2.359 (95% CI 1.278-4.355) times, and depression in general by 1.933 (95% CI 1.142-3.271) times, compared with persons carrying the LALA genotype (32.0% and 46.9%), (χ2=7.674, df=1, p<0.01 and χ2=6.095, df=1, p<0.05). Persons carrying the LALG genotype (54.5%) also had a higher chance of developing a mean level of depression RR=2.929 (95% CI 1.039-8.261), compared with carriers of the LALA genotype (32.0%) (χ2=4.326, df =1, p<0.05) (p<0.05). CONCLUSION: Associative links between polymorphic variants of the SLC6A4 gene and depression have been established.

5-HT_FAsTR: a versatile, label-free, high-throughput, fluorescence-based microplate assay to quantify serotonin transport and release.

The neurotransmitter serotonin plays a pivotal role in mood and depression. It also acts as a vasoconstrictor within blood vessels and is the main neurotransmitter in the gastrointestinal system. In neurotransmission, released serotonin is taken up by serotonin transporters, which are principal targets of antidepressants and the psychostimulant, ecstasy. The investigation of serotonin transporters have relied almost exclusively on the use of radiolabeled serotonin in heterogenous end-point assays. Here we adapt the genetically encoded fluorescent biosensor, iSeroSnFR, to establish and validate the Serotonin (5-HT) Fluorescence Assay for Transport and Release (5-HT_FAsTR) for functional and pharmacological studies of serotonin transport and release. We demonstrate the applicability of the method for the study of a neuronal, high-affinity, low-capacity serotonin transporter (SERT) as well as an extraneuronal low-affinity, high-capacity organic cation transporter and mutants thereof. 5HT_FAsTR offers an accessible, versatile and reliable semi-homogenous assay format that only relies on a fluorescence plate reader for repeated, real-time measurements of serotonin influx and efflux. 5HT_FAsTR accelerates and democratizes functional characterization and pharmacological studies of serotonin transporters and genetic variants thereof in disease states such as depression, anxiety and ADHD.

Pharmacogenetic Factors Influence Escitalopram Pharmacokinetics and Adverse Events in Youth with a Family History of Bipolar Disorder: A Preliminary Study.

Introduction: Escitalopram is an effective and generally well-tolerated antidepressant, but children of parents with bipolar disorder (BD) may be at increased risk for adverse events associated with antidepressants, including increased irritability, restlessness, impulsivity, and manic symptoms. This risk may be influenced by polymorphisms in genes encoding cytochrome P450 enzymes (CYP2C19 or CYP2D6), the serotonin transporter (SLC6A4), and the serotonin receptor 2A subtype (HTR2A). We explored whether gene-drug interactions influence the emergence of adverse events in depressed and/or anxious youth with a family history of BD. Materials and Methods: Children and adolescents aged 12-17 years with a first-degree relative with bipolar I disorder were treated with escitalopram and monitored for adverse effects, underwent pharmacogenetic testing, and provided serum escitalopram levels. Emergence of adverse events was determined by study clinicians, and symptoms were tracked using the Treatment-Emergent Activation and Suicidality Assessment Profile (TEASAP) and Pediatric Adverse Events Rating Scale. Clinical Pharmacogenetics Implementation Consortium guidelines were used to determine CYP2C19 and CYP2D6 phenotypes. Results: Slower CYP2C19 metabolizers had greater dose-normalized 24-hour area under the curve (AUC0-24; p = 0.025), trough concentrations (Ctrough; p = 0.013), and elimination half-lives (t1/2; p < 0.001). CYP2D6 phenotype was not significantly associated with any pharmacokinetic parameter. Slower CYP2D6 metabolizers had increased TEASAP akathisia (p = 0.015) scores. HTR2A A/A and A/G genotypes were associated with increased TEASAP "self-injury, suicidality, and harm to others" subscale scores (p = 0.017). Escitalopram maximum concentration, AUC0-24, CYP2C19 phenotype, and SLC6A4 genotype were not associated with adverse events. Conclusions: CYP2C19 phenotype influences escitalopram pharmacokinetics whereas CYP2D6 phenotype does not. Slower CYP2D6 metabolism was associated with increased akathisia, and HTR2A A/A or A/G genotypes were associated with increased risk of self-harm or harm to others. Larger cohorts are needed to identify associations between genetic test results and antidepressant-associated adverse events. Trial Registration: ClinicalTrials.gov identifier: NCT02553161.

SLC6A4 gene variants moderate associations between childhood food insecurity and adolescent mental health.

BACKGROUND: Food insecurity is a persistent concern in the United States and has been shown to affect child mental health and behavior. The SLC6A4 gene has been indicated as a moderator of the effects of chronic stress on anxiety in adolescents aged 14-21. However, it is unclear if SLC6A4 may also play a role in the effects of childhood food insecurity, a form of chronic stress, on adolescent mental health. This study aimed to identify effects of food insecurity on adolescents' mental health and delinquent behavior when both mom and child go hungry in the child's early years, and the potential interaction with SLC6A4 variants (SS/LL). METHODS: The data and sample for this research are from the Future of Families and Child Wellbeing Study. The cohort consists of 4898 children (age 1-15 years, male = 47%, African American = 50%) and their respective caregivers sampled from large cities in the United States from 1998 to 2000. RESULTS: The SLC6A4 serotonin transporter short/short allele emerged statistically significant as a moderator of childhood food insecurity and adolescent mental health. Specifically, the presence of the short/short allele increased anxiety symptoms in adolescents with exposure to food insecurity in childhood. CONCLUSION: The SLC6A4 short/short allele amplifies risk of anxiety-related mental illness when children experience food insecurity. The gene-environment interaction provides insight into the mechanistic pathway of the effects of poverty-related adversity, such as food insecurity, on developmental trajectories of mental health.

Chronic social defeat stress induces anxiety-like behaviors via downregulation of serotonin transporter in the prefrontal serotonergic system in mice.

The serotonergic (5-HTergic) system is closely involved in the pathophysiology of mood and anxiety disorders and the responsibility of this system may differ for each symptom. In this study, we examined the relationship between the dysfunction of the 5-HTergic system and abnormal behaviors in the social defeat stress model, an animal model of mood and anxiety disorders and in mice with knockdown of Slc6a4, the gene encoding SERT. Monoamine content, serotonin (5-HT) release, 5-HT uptake, 5-HT transporter (SERT) protein levels, and behaviors were investigated in mice subjected to chronic social defeat stress and in mice with knockdown of Slc6a4, in 5-HTergic neurons projecting to the prefrontal cortex (PFC). Furthermore, DNA methylation of Slc6a4 was examined in mice subjected to chronic social defeat stress. Increased turnover, increased extracellular basal levels, decreased release and decreased uptake of 5-HT, and decreased SERT protein levels were observed in the PFC of the stressed mice. The decreased 5-HT uptake correlated with anxiety-like behavior characterized by decreased time spent in the open arms of the elevated plus maze. DNA methylation was increased in the CpG island of Slc6a4 in 5-HTergic neurons projecting to the PFC of the stressed mice. Similar to the stressed mice, mice with Slc6a4 knockdown in 5-HTergic neurons projecting to the PFC also showed decreased release and uptake of 5-HT in the PFC and increased anxiety-like behavior. Chronic stress may induce anxiety due to dysfunction in the prefrontal 5-HTergic system via decreased SERT expression in the PFC.

Serotonin and the serotonin transporter in the adrenal gland.

The adrenal glands are key components of the mammalian endocrine system, helping maintain physiological homeostasis and the coordinated response to stress. Each adrenal gland has two morphologically and functionally distinct regions, the outer cortex and inner medulla. The cortex is organized into three concentric zones which secrete steroid hormones, including aldosterone and cortisol. Neural crest-derived chromaffin cells in the medulla are innervated by preganglionic sympathetic neurons and secrete catecholamines (epinephrine, norepinephrine) and neuropeptides into the bloodstream, thereby functioning as the neuroendocrine arm of the sympathetic nervous system. In this article we review serotonin (5-HT) and the serotonin transporter (SERT; SLC6A4) in the adrenal gland. In the adrenal cortex, 5-HT, primarily sourced from resident mast cells, acts as a paracrine signal to stimulate aldosterone and cortisol secretion through 5-HT4/5-HT7 receptors. Medullary chromaffin cells contain a small amount of 5-HT due to SERT-mediated uptake and express 5-HT1A receptors which inhibit secretion. The atypical mechanism of the 5-HT1A receptors and interaction with SERT fine tune this autocrine pathway to control stress-evoked catecholamine secretion. Receptor-independent signaling by SERT/intracellular 5-HT modulates the amount and kinetics of transmitter release from single vesicle fusion events. SERT might also influence stress-evoked upregulation of tyrosine hydroxylase transcription. Transient signaling via 5-HT3 receptors during embryonic development can limit the number of chromaffin cells found in the mature adrenal gland. Together, this emerging evidence suggests that the adrenal medulla is a peripheral hub for serotonergic control of the sympathoadrenal stress response.

Tailoring Type II Diabetes Treatment: Investigating the Effect of 5-HTT Polymorphisms on HbA1c Levels after Metformin Initiation.

Aims: To investigate the effect of serotonin transporter (5-HTT) polymorphisms on change in HbA1c levels six months after metformin initiation in type 2 diabetes patients. Materials and Methods: Participants of PROVALID (PROspective cohort study in patients with type 2 diabetes mellitus for VALidation of biomarkers) within the GIANTT (Groningen Initiative to ANalyse Type 2 Diabetes Treatment) cohort who initiated metformin were genotyped for combined 5-HTTLPR/rs25531 (L∗L∗, L∗S∗, and S∗S∗) and 5-HTT VNTR (STin 2.12, 12/-, and 10/-) polymorphisms, respectively. Multiple linear regression was applied to determine the change in HbA1c level from baseline date to six months across 5-HTTLPR/VNTR genotype groups, adjusted for baseline HbA1c, age, gender, triglyceride level, low-density lipoprotein level, and serum creatinine. Results: 157 participants were included, of which 56.2% were male. The average age was 59.3 ± 9.23 years, and the mean baseline HbA1c was 7.49% ± 1.21%. 5-HTTLPR was characterized in 46 patients as L∗L∗, 70 patients as L∗S∗, and 41 patients as S∗S∗ genotypes. No significant association was found between 5-HTTLPR and 5-HTT VNTR genotypes and change in HbA1c after adjustments. Conclusions: 5-HTT polymorphisms did not affect HbA1c levels six months after the start of metformin. Further long-term studies in large samples would be relevant to determine which polymorphisms can explain the variation in response to metformin treatment.

Serotonin Transporter-dependent Histone Serotonylation in Placenta Contributes to the Neurodevelopmental Transcriptome.

Brain development requires appropriate regulation of serotonin (5-HT) signaling from distinct tissue sources across embryogenesis. At the maternal-fetal interface, the placenta is thought to be an important contributor of offspring brain 5-HT and is critical to overall fetal health. Yet, how placental 5-HT is acquired, and the mechanisms through which 5-HT influences placental functions, are not well understood. Recently, our group identified a novel epigenetic role for 5-HT, in which 5-HT can be added to histone proteins to regulate transcription, a process called H3 serotonylation. Here, we show that H3 serotonylation undergoes dynamic regulation during placental development, corresponding to gene expression changes that are known to influence key metabolic processes. Using transgenic mice, we demonstrate that placental H3 serotonylation is dependent on 5-HT uptake by the serotonin transporter (SERT/SLC6A4). SERT deletion robustly reduces enrichment of H3 serotonylation across the placental genome, and disrupts neurodevelopmental gene networks in early embryonic brain tissues. Thus, these findings suggest a novel role for H3 serotonylation in coordinating placental transcription at the intersection of maternal physiology and offspring brain development.

Methylation of serotonin regulating genes in cord blood cells: association with maternal metabolic parameters and correlation with methylation in peripheral blood cells during childhood and adolescence.

BACKGROUND: Serotonin (5-hydroxytryptamine, 5-HT) signaling is involved in neurodevelopment, mood regulation, energy metabolism, and other physiological processes. DNA methylation plays a significant role in modulating the expression of genes responsible for maintaining 5-HT balance, such as 5-HT transporter (SLC6A4), monoamine oxidase A (MAOA), and 5-HT receptor type 2A (HTR2A). Maternal metabolic health can influence long-term outcomes in offspring, with DNA methylation mediating these effects. We investigated associations between maternal metabolic parameters-pre-pregnancy body mass index (pBMI), gestational weight gain (GWG), and glucose tolerance status (GTS), i.e., gestational diabetes mellitus (GDM) versus normal glucose tolerance (NGT)-and cord blood methylation of SLC6A4, MAOA, and HTR2A in participants from our PlaNS birth cohort. CpG sites (15, 9, and 2 in each gene, respectively) were selected based on literature and in silico data. Methylation levels were quantified by bisulfite pyrosequencing. We also examined the stability of methylation patterns in these genes in circulating blood cells from birth to adolescence using longitudinal DNA methylation data from the ARIES database. RESULTS: None of the 203 PlaNS mothers included in this study had preexisting diabetes, 99 were diagnosed with GDM, and 104 had NGT; all neonates were born at full term by planned Cesarean section. Methylation at most CpG sites differed between male and female newborns. SLC6A4 methylation correlated inversely with maternal pBMI and GWG, while methylation at HTR2A site -1665 correlated positively with GWG. None of the maternal metabolic parameters statistically associated with MAOA methylation. DNA methylation data in cord blood and peripheral blood at ages 7 and 15 years were available for 808 participants from the ARIES database; 4 CpG sites (2 in SLC6A4 and 2 in HTR2A) overlapped between the PlaNS and ARIES cohorts. A positive correlation between methylation levels in cord blood and peripheral blood at 7 and 15 years of age was observed for both SLC6A4 and HTR2A CpG sites. CONCLUSIONS: Methylation of 5-HT regulating genes in cord blood cells is influenced by neonatal sex, with maternal metabolism playing an additional role. Inter-individual variations present in circulating blood cells at birth are still pronounced in childhood and adolescence.

Offspring's own serotonin transporter genotype, independently from the maternal one, increases anxiety- and depression-like behavior and alters neuroplasticity markers in rats.

INTRODUCTION: Developmental changes due to early life variations in the serotonin system affect stress-related behavior and neuroplasticity in adulthood. These outcomes can be caused both by offspring's own and maternal serotonergic genotype. We aimed to dissociate the contribution of the own genotype from the influences of mother genotype. METHODS: Sixty-six male homozygous (5-HTT-/-) and heterozygous (5-HTT+/-) serotonin transporter knockout and wild-type rats from constant 5-HTT genotype mothers crossed with varying 5-HTT genotype fathers were subjected to tests assessing anxiety- and depression-like behaviors. Additionally, we measured plasma corticosterone levels and mRNA levels of BDNF, GABA system and HPA-axis components in the prelimbic and infralimbic cortex. Finally, we assessed the effect of paternal 5-HTT genotype on these measurements in 5-HTT+/- offspring receiving their knockout allele from their mother or father. RESULTS: 5-HTT-/- offspring exhibited increased anxiety- and depression-like behavior in the elevated plus maze and sucrose preference test. Furthermore, Bdnf isoform VI expression was reduced in the prelimbic cortex. Bdnf isoform IV and GABA related gene expression was also altered but did not survive false discovery rate (FDR) correction. Finally, 5-HTT+/- offspring from 5-HTT-/- fathers displayed higher levels of anxiety- and depression-like behavior and changes in GABA, BDNF and HPA-axis related gene expression not surviving FDR correction. LIMITATIONS: Only male offspring was tested. CONCLUSIONS: Offspring's own 5-HTT genotype influences stress-related behaviors and Bdnf isoform VI expression, independently of maternal 5-HTT genotype. Paternal 5-HTT genotype separately influenced these outcomes. These findings advance our understanding of the 5-HTT genotype dependent susceptibility to stress-related disorders.

Chronic exposure to imipramine induces a switch from depression-like to mania-like behavior in female serotonin transporter knockout rats: Role of BDNF signaling in the infralimbic cortex.

BACKGROUND: Bipolar disorder (BD) is a highly burdensome psychiatric disorder characterized by alternating states of mania and depression. A major challenge in the clinic is the switch from depression to mania, which is often observed in female BD patients during antidepressant treatment such as imipramine. However, the underlying neural basis is unclear. METHODS: To investigate the potential neuronal pathways, serotonin transporter knockout (SERT KO) rats, an experimental model of female BD patients, were subjected to a battery of behavioral tests under chronic treatment of the antidepressant imipramine. In addition, the expression of brain-derived neurotrophic factor (BDNF) and its downstream signaling was examined in the prefrontal cortex. RESULTS: Chronic exposure to imipramine reduced anxiety and sociability and problem-solving capacity, and increased thigmotaxis and day/night activity in all animals, but specifically in female SERT KO rats, compared to female wild-type (WT) rats. Further, we found an activation of BDNF-TrkB-Akt pathway signaling in the infralimbic, but not prelimbic, cortex after chronic imipramine treatment in SERT KO, but not WT, rats. LIMITATIONS: Repeated testing behaviors could potentially affect the results. Additionally, the imipramine induced changes in behavior and in the BDNF system were measured in separate animals. CONCLUSIONS: Our study indicates that female SERT KO rats, which mirror the female BD patients with the 5-HTTLPR s-allele, are at higher risk of a switch to mania-like behaviors under imipramine treatment. Activation of the BDNF-TrkB-Akt pathway in the infralimbic cortex might contribute to this phenotype, but causal evidence remains to be provided.

Neuropeptide Y gene variants and Agreeableness: interaction effect with the birth cohort and the serotonin transporter promoter polymorphism.

OBJECTIVE: Neuropeptide Y (NPY) is a powerful regulator of anxious states, including social anxiety, but evidence from human genetic studies is limited. Associations of common gene variants with behaviour have been described as subject to birth cohort effects, especially if the behaviour is socially motivated. This study aimed to examine the association of NPY rs16147 and rs5574 with personality traits in highly representative samples of two birth cohorts of young adults, the samples having been formed during a period of rapid societal transition. METHODS: Both birth cohorts (original n = 1238) of the Estonian Children Personality Behaviour and Health Study (ECPBHS) self-reported personality traits of the five-factor model at 25 years of age. RESULTS: A significant interaction effect of the NPY rs16147 and rs5574 and birth cohort on Agreeableness was found. The T/T genotype of NPY rs16147 resulted in low Agreeableness in the older cohort (born 1983) and in high Agreeableness in the younger cohort (born 1989). The C/C genotype of NPY rs5574 was associated with higher Agreeableness in the younger but not in the older cohort. In the NPY rs16147 T/T homozygotes, the deviations from average in Agreeableness within the birth cohort were dependent on the serotonin transporter promoter polymorphism. CONCLUSIONS: The association between the NPY gene variants and a personality domain reflecting social desirability is subject to change qualitatively in times of rapid societal changes, serving as an example of the relationship between the plasticity genes and environment. The underlying mechanism may involve the development of the serotonergic system.

Most patients with disorders of gut-brain interaction receive pharmacotherapy with major or moderate drug-gene interactions.

BACKGROUND: How variations predicted by pharmacogenomic testing to alter drug metabolism and therapeutic response affect outcomes for patients with disorders of gut- brain interaction is unclear. AIMS: To assess the prevalence of pharmacogenomics-predicted drug-gene interactions and symptom outcomes for patients with disorders of gut-brain interaction. METHODS: Patients who were treated in our clinical practice for functional dyspepsia/bowel disorder underwent pharmacogenomic testing. The change in symptoms from baseline to 6 months was compared for patients with variations in CYP2D6 and CYP2C19, which metabolize neuromodulators, and SLC6A4, which encodes the sodium- dependent serotonin transporter. RESULTS: At baseline, 79 of 94 participants (84%) had at least one predicted major drug- gene interaction, and all 94 (100%) had at least one predicted moderate interaction. For the 44 participants who completed a survey of their symptoms at 6 months, the mean (SD) irritable bowel syndrome-symptom severity score decreased from 284 (71) at baseline to 231 (95) at 6 months (p < 0.001). Among patients taking selective serotonin reuptake inhibitors, the decrease in symptom severity (p = 0.03) and pain (p = 0.002) scores from baseline to 6 months was greater for patients with a homozygous SLC6A4 long/long genotype (n = 30) (ie, increased serotonin transporter activity) than for patients with homozygous short/short or heterozygous long/short genotypes (n = 64). Symptom outcomes were not affected by CYP2D6 or CYP2C19 variations. CONCLUSIONS: The homozygous SLC6A4 long/long genotype confers better symptom resolution for patients with disorders of gut-brain interaction who take selective serotonin reuptake inhibitors than do the homozygous short/short or heterozygous long/short genotypes.

Aberrant colon metabolome and the sudden infant death syndrome.

BACKGROUND: The Sudden Infant Death Syndrome (SIDS) has been associated with increased peripheral serotonin and an abnormal colonic microbiome, suggesting the colonic metabolome may also be abnormal. This study addresses this potential correlation by comparing colonic autopsy tissue from SIDS to age-matched non-SIDS controls. METHODS: Untargeted metabolomic analysis by mass spectrometry is used to assess human colonic metabolomic differences including serotonin. Expression of genes associated with colonic serotonin synthesis and transport (TPH1, TPH2, DDC, SCL6A4) is measured by qRT-PCR. Microbiome analysis is performed to compare the SIDS and non-SIDS colonic microbiome. RESULTS: Unsupervised hierarchical cluster and principal component analyses of metabolomic data shows increased variability in the SIDS cohort and separation of SIDS cases from the non-SIDS controls. There is a trend toward increased serotonin in the SIDS cohort but there is no significant difference in expression of the serotonin synthesis and transport genes between SIDS and non-SIDS control cohorts. Microbiome analysis shows no significant difference between the SIDS and non-SIDS control cohorts. CONCLUSIONS: This study demonstrates increased variability in the colonic metabolome and a trend towards increased colonic serotonin in SIDS. The underlying cause of colon metabolomic variability, and its potential role in SIDS pathogenesis, warrants further investigation. IMPACT STATEMENT: The key message of this article is that SIDS is associated with an aberrant colonic metabolome. This is a novel observation suggesting another component in the pathophysiology underlying SIDS. Investigation of why the colonic metabolome is aberrant may offer new insights to SIDS pathogenesis and new strategies to reduce risk.