Serotonin Transporter Research Articles

Altered Serotonin 1B Receptor Binding After Escitalopram for Depression Is Correlated With Treatment Effect.

Major depressive disorder (MDD) is commonly treated with selective serotonin reuptake inhibitors (SSRIs). SSRIs inhibit the serotonin transporter (5-HTT), but the downstream antidepressant mechanism of action of these drugs is poorly understood. The serotonin 1B (5-HT1B) receptor is functionally linked to 5-HTT and 5-HT1B receptor binding and 5-HT1B receptor mRNA is reduced in the raphe nuclei after SSRI administration in primates and rodents, respectively. The effect of SSRI treatment on 5-HT1B receptor binding in patients with MDD has not been examined previously. This positron emission tomography (PET) study aimed to quantify brain 5-HT1B receptor binding changes in vivo after SSRI treatment for MDD in relation to treatment effect. Eight unmedicated patients with moderate to severe MDD underwent PET with the 5-HT1B receptor radioligand [11C]AZ10419369 before and after 3 to 4 weeks of treatment with the SSRI escitalopram 10mg daily. Depression severity was assessed at time of PET and after 6 to 7 weeks of treatment with the Montgomery-Åsberg Depression Rating Scale. We observed a significant reduction in [11C]AZ10419369 binding in a dorsal brainstem (DBS) region containing the median and dorsal raphe nuclei after escitalopram treatment (P = .036). Change in DBS [11C]AZ10419369 binding correlated with Montgomery-Åsberg Depression Rating Scale reduction after 3-4 (r = 0.78, P = .021) and 6-7 (r = 0.94, P < .001) weeks' treatment. Our findings align with the previously reported reduction of 5-HT1B receptor binding in the raphe nuclei after SSRI administration and support future studies testing change in DBS 5-HT1B receptor binding as an SSRI treatment response marker.

Design, Docking, Characterisation, and Synthesis of Pyrimidine Derivatives for Antidepressant Activity.

According to the report, in 2022, the prevalence rate of depression in India was 4.50%, and the cases stood at 56,675,969. The development of antidepressant agents has reduced the number of depressant and suicidal cases. Many researchers have found that pyrimidine possesses antidepressant activity. With this background, we thought of synthesizing pyrimidine derivatives. The objective of this study is to carry out molecular docking, synthesis, characterization, and evaluation of 2-((4,6-diphenylpyrimidin-2-yl)oxy)-N-phenylacetamide derivatives (17-26) as in vivo antidepressant agent. The designed compounds were checked for their activity using Molegro virtual docker (MVD) and were further synthesized. Benzaldehyde reacted with acetophenone to give compound (3), which gave compound (4) upon reaction with urea. In another reaction, substituted anilines (5) were reacted with chloroacetyl chloride (6) to yield compounds (7-16), which upon further reaction with compound (4) yielded the final derivatives (17-26). The synthesized compounds were characterized by spectral analysis and checked for their antidepressant activity. The MolDock scores of the derivatives ranged from -147.097 to -182.095, whereas of active ligand IXX_801 was -115.566. All the synthesized pyrimidine derivatives showed better affinity towards the Cryo-EM structure of the wild-type human serotonin transporter complexed with vilazodone, imipramine, and 15B8 Fab protein (PDB ID: 7LWD) as compared to standard drug clomipramine (-101.064). All the synthesized derivatives were screened for antidepressant activity at a 100mg/kg dose level compared to the standard clomipramine HCl at a dose level of 20mg/kg. Among all the synthesized derivatives, compound 24 showed the most potent antidepressant activity, and Compound 20 showed moderate antidepressant activity, which reduced the duration of immobility times to 35.42% and 31.97% at 100mg/kg dose level when compared to the control, respectively. Compound 24 showed the highest MolDock score as well as found to be the most potent antidepressant agent.

Theranostics in Neuroendocrine Tumors.

Neuroendocrine tumors (NETs) are rare tumors that develop from cells of the neuroendocrine system and can originate in multiple organs and tissues such as the bowels, pancreas, adrenal glands, ganglia, thyroid, and lungs. This review will focus on gastroenteropancreatic NETs (more commonly called NETs) characterized by frequent somatostatin receptor (SSTR) overexpression and pheochromocytomas/paragangliomas (PPGLs), which typically overexpress norepinephrine transporter. Advancements in SSTR-targeted imaging and treatment have revolutionized the management of patients with NETs. This comprehensive review delves into the current practice, discussing the use of the various Food and Drug Administration-approved SSTR-agonist positron emission tomography tracers and the predictive imaging biomarkers, and elaborating on 177Lu-DOTATATE peptide receptor radionuclide therapy including the evolving areas of posttherapy imaging practices and peptide receptor radionuclide therapy retreatment. SSTR-targeted imaging and therapy can also be used in patients with PPGL; however, this patient population has demonstrated the best outcomes from norepinephrine transporter-targeted therapy with 131I-metaiodobenzylguanidine. Metaiodobenzylguanidine theranostics for PPGL will be discussed, noting that in 2024 it became commercially unavailable in the United States. Therefore, the use and reported success of SSTR theranostics for PPGL will also be explored.

The effect of single nucleotide polymorphisms on depression in combination with coronary diseases: a systematic review and meta-analysis.

Depression and coronary heart disease (CHD) have common risk mechanisms. Common single nucleotide polymorphisms (SNPs) may be associated with the risk of depression combined with coronary heart disease. This study was designed according to the PRISMA-P guidelines. We will include case-control studies and cohort studies investigating the relationship between gene SNPs and depression and coronary heart disease comorbidities. The Newcastle-Ottawa Scale (NOS) will be used to assess the risk of bias. When measuring dichotomous outcomes, we will use the odds ratio (OR) and 95% confidence interval (95%CIs) in a case-control study. Five genetic models (allele model, homozygous model, co-dominant model, dominant model, and recessive model) will be evaluated for each included study. Subgroup analysis by ethnicity will be performed. If necessary, post hoc analysis will be made according to different types. A total of 13 studies were included in this study, and the types of genes included are FKBP5 and SGK1 genes that act on glucocorticoid; miR-146a, IL-4-589, IL-6-174, TNF-α-308, CRP-717 genes that act on inflammatory mechanisms; eNOS genes from endothelial cells; HSP70 genes that act on the autoimmune response; ACE2 and MAS1 genes that act to mediate Ang(1-7) in the RAS system; 5-HTTLPR gene responsible for the transport of serotonin 5-HT and neurotrophic factor BDNF gene. There were three studies on 5-HTTLPR and BDNF genes, respectively, while there was only one study targeting FKBP5, SGK1, miR-146a, IL-4-589, IL-6-174, TNF-alpha-308, CRP-717, eNOS, HSP70, ACE2, and MAS1 genes. We did not perform a meta-analysis for genes reported in a single study, and meta-analysis was performed separately for studies exploring the 5-HTTLPR and BDNF genes. The results showed that for the 5-HTTLPR gene, there was a statistically significant association between 5-HTTLPR gene polymorphisms and depression in combination with coronary diseases (CHD-D) under the co-dominant model (LS vs LL: OR 1.76, 95%CI 1.20-2.59; SS vs LL: OR 2.80, 95%CI 1.45 to 5.41), the dominant model (LS+SS vs LL: OR 2.06, 95%CI 1.44 to 2.96), and the homozygous model (SS vs LL: OR 2.80 95%CI 1.45 to 5.5.41) were statistically significant for CHD-D, demonstrating that polymorphisms in the 5-HTTLPR gene are associated with the development of CHD-D and that the S allele in the 5-HTTLPR gene is likely to be a risk factor for CHD-D. For the BDNF gene, there were no significant differences between one of the co-dominant gene models (AA vs GG: OR 6.63, 95%CI 1.44 to 30.64), the homozygous gene model (AA vs GG: OR 6.63,95% CI 1.44 to 30.64), the dominant gene model (GA+AA vs GG: OR4.29, 95%CI 1.05 to 17.45), recessive gene model (AA vs GG+GA: OR 2.71, 95%CI 1.16 to 6.31), and allele model (A vs G: OR 2.59, 95%CI 1.18 to 5.67) were statistically significant for CHD-D, demonstrating that BDNFrs6265 gene polymorphisms are associated with the CHD-D development and that the A allele in the BDNFrs6265 gene is likely to be a risk factor for CHD-D. We analyzed the allele frequencies of SNPs reported in a single study and found that the SNPs in the microRNA146a gene rs2910164, the SNPs in the ACE2 gene rs2285666 and the SNPs in the SGK1 gene rs1743963 and rs1763509 were risk factors for the development of CHD-D. We performed a subgroup analysis of three studies involving the BDNFrs6265 gene. The results showed that European populations were more at risk of developing CHD-D than Asian populations in both dominant model (GA+AA vs GG: OR 10.47, 95%CI 3.53 to 31.08) and co-dominant model (GA vs GG: OR 6.40, 95%CI 1.98 to 20.73), with statistically significant differences. In contrast, the studies involving the 5-HTTLPR gene were all Asian populations, so subgroup analyses were not performed. We performed sensitivity analyses of studies exploring the 5-HTTLPR and BDNF rs6265 genes. The results showed that the results of the allele model, the dominant model, the recessive model, the homozygous model and the co-dominant model for both 5-HTTLPR and BDNF rs6265 genes were stable. Due to the limited number of studies of the 5-HTTLPR and BDNF genes, it was not possible to determine the symmetry of the funnel plot using Begg's funnel plot and Egger's test. Therefore, we did not assess publication bias. SNPs of the microRNA146a gene at rs2910164, the ACE2 gene at the rs2285666 and the SGK1 gene at rs1743963 and rs1763509, and the SNPs at the 5-HTTLPR and BDNF gene loci are associated with the onset of comorbid depression in coronary heart disease. We recommend that future research focus on studying SNPs' impact on comorbid depression in coronary heart disease, specifically targeting the 5-HTTLPR and BDNF gene at rs6265. https://www.crd.york.ac.uk/prospero/, identifier CRD42021229371.

Serotonin Transporter (SLC6A4) and FK506-Binding Protein 5 (FKBP5) Genotype and Methylation Relationships with Response to Meditation in Veterans with PTSD.

Meditation-based interventions are novel and effective non-pharmacologic treatments for veterans with PTSD. We examined relationships between treatment response, early life trauma exposure, DNA polymorphisms, and methylation in the serotonin transporter (SLC6A4) and FK506-binding protein 5 (FKBP5) genes. DNA samples and clinical outcomes were examined in 72 veterans with PTSD who received meditation-based therapy in two separate studies of mindfulness-based stress reduction (MBSR) and Transcendental Meditation (TM). The PTSD Checklist was administered to assess symptoms at baseline and after 9weeks of meditation intervention. We examined the SLC6A4 promoter (5HTTLPR_L/S insertion/deletion + rs25531_A/G) polymorphisms according to previously defined gene expression groups, and the FKBP5 variant rs1360780 previously associated with PTSD disease risk. Methylation for CpG sites of SLC6A4 (28 sites) and FKBP5 (45 sites) genes was quantified in DNA samples collected before and after treatment. The 5HTTLPR LALA high expression genotype was associated with greater symptom improvement in participants exposed to early life trauma (p = 0.015). Separately, pre to post-treatment change of DNA methylation in a group of nine FKBP5 CpG sites was associated with greater symptom improvement (OR = 2.8, 95% CI 1.1-7.1, p = 0.027). These findings build on a wealth of existing knowledge regarding epigenetic and genetic relationships with PTSD disease risk to highlight the potential importance of SLC6A4 and FKBP5 for treatment mechanisms and as biomarkers of symptom improvement.

Tryptophan-5-hydroxylase, serotonin transporter protein and serotonergic receptors in dilated cardiomyopathy

Tryptophan-5-hydroxylase, serotonin transporter protein and serotonergic receptors in dilated cardiomyopathy

Unraveling the molecular basis of cannabidiolic acid methyl Ester's anti-depressive effects in a rat model of treatment-resistant depression

Major depressive disorder (MDD) stands as a significant cause of disability globally. Cannabidiolic Acid-Methyl Ester (CBDA-ME) (EPM-301, HU-580), a derivative of Cannabidiol, demonstrates immediate antidepressant-like effects, yet it has undergone only minimal evaluation in psychopharmacology. Our goal was to investigate the behavioral and potential molecular mechanisms associated with the chronic oral administration of this compound in the Wistar Kyoto (WKY) genetic model of treatment-resistant depression. Male WKY rats were subjected to behavioral assessments before and after receiving chronic (14-day) oral doses of CBDA-ME (0.5 mg/kg), 15 mg/kg of imipramine or vehicle. At the end of the study, plasma corticosterone levels and mRNA expression of various genes in the medial Prefrontal Cortex and Hippocampus were measured. Behavioral outcomes from CBDA-ME treatment indicated an antidepressant-like effect similar to imipramine, as oral ingestion reduced immobility and increased swimming duration in the Forced Swim Test. Neither treatment influenced locomotion in the Open Field Test nor preference in the Saccharin Preference Test. The behavioral impact in WKY rats coincided with reduced corticosterone serum levels, upregulated mRNA expression of Cannabinoid receptor 1, Fatty Acid Amide Hydrolase, and Corticotropin-Releasing Hormone Receptor 1, alongside downregulation of the Serotonin Transporter in the hippocampus. Additionally, there was an upregulation of CB1 mRNA expression and downregulation of Brain-Derived Neurotrophic Factor in the mPFC. These findings contribute to our limited understanding of the antidepressant effects of CBDA-ME and shed light on its potential psychopharmacological mechanisms. This discovery opens up possibilities for utilizing cannabinoids in the treatment of major depressive disorder and related conditions.

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.

Cannabidiol disrupts tryptophan metabolism in the human term placenta

The increasing use of cannabis during pregnancy raises concerns about its impact on fetal development. While cannabidiol (CBD) shows therapeutic promise, its effects during pregnancy remain uncertain. We investigated CBD's influence on tryptophan (TRP) metabolism in the human placenta. TRP is an essential amino acid that is metabolized via the serotonin and kynurenine (KYN) pathways, which are critical for fetal neurodevelopment. We used human term villous placental explants, an advanced ex vivo model, to study CBD's impact on key TRP metabolic enzymes. In addition, vesicles isolated from the microvillous membrane (MVM) of the human placenta were used to assess CBD’s effect on placental serotonin uptake. Explants were exposed to CBD at therapeutic (0.1, 1, 2.5 μg/ml) and non-therapeutic (20 and 40 μg/ml) concentrations to determine its effects on the gene and protein expression of key enzymes in TRP metabolism and metabolite release. CBD upregulated TRP hydroxylase (TPH) and downregulated monoamine oxidase (MAO-A), resulting in reduced levels of 5-hydroxyindoleacetic acid (HIAA). It also downregulated serotonin transporter expression and inhibited serotonin transport across the MVM by up to 60% while simultaneously enhancing TRP metabolism via the kynurenine pathway by upregulating indoleamine-pyrrole 2,3-dioxygenase (IDO-1). Among kynurenine pathway enzymes, kynurenine 3 monooxygenase (KMO) was upregulated while kynurenine aminotransferase 1 (KAT-1) was downregulated; the former is associated with neurotoxic metabolite production, while the latter is linked to reduced neuroprotective metabolite levels. Overall, these results indicate that CBD modulates TRP catabolism in the human placenta, potentially disrupting the tightly regulated homeostasis of the serotonin and KYN pathways.

Postpartum Depression: Etiology, Treatment, and Consequences for Maternal Care.

Postpartum depression (PPD) is a disabling condition that has recently shown an increase in prevalence, becoming an essential public health problem. This study is a qualitative review summarizing the most frequent risk factors associated with PPD, evaluating molecular aspects of PPD and current approaches to detect and prevent PPD. The most prevalent risk factors were detected in the areas of economic and social factors, obstetrical history, lifestyle, and history of mental illness. Research on the genetic basis for PPD has taken place in recent years to identify the genes responsible for establishing targeted therapeutic methods and understanding its pathogenesis. The most frequently studied candidate gene was the serotonin transporter gene (SERT) associated with PPD. Among biological studies, antidepressants and psychological interventions provided the most evidence of successful intervention. The obstetrician can serve an essential role in screening for and treating PPD. Postpartum women with risk factors should be screened using the Edinburgh Postnatal Depression Scale (EPDS), but, at the moment, there are no prevention programs in Europe. In conclusion, data from this review increase concerns among this vulnerable population and can be used to design a screening tool for high-risk pregnant women and create a prevention program.

0064 Sleep Loss Affects Item and Source Memory Differentially as a Function of Serotonin Transporter (5-HTTLPR) Genotype

0064 Sleep Loss Affects Item and Source Memory Differentially as a Function of Serotonin Transporter (5-HTTLPR) Genotype

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.

Feasibility Study of Single-Photon Emission Computed Tomography with Iodine-123 Labeled Metaiodobenzylguanidine for Preclinical Evaluation of Labetalol as a β-Adrenergic Receptor Blocker.

Among clinically used radiopharmaceuticals, iodine-123 labeled metaiodobenzylguanidine ([123I]mIBG) serves for diagnosing neuroendocrine tumors and obtaining images of myocardial sympathetic innervation. mIBG, a structural analogue of norepinephrine (NE), a neurotransmitter acting in peripheral and central nerves, follows a pathway similar to NE, transmitting signals through the NE transporter (NET) located at synaptic terminals. It moves through the body without decomposing, enabling noninvasive image evaluation. In this study, we aimed to quantify [123I]mIBG uptake in the adrenal glands using small animal single-photon emission computed tomography/computed tomography (SPECT/CT) images post [123I]mIBG administration. We investigated the possibility of assessing the effectiveness of β-adrenergic receptor blockers by quantifying SPECT/CT images and biodistribution results to determine the degree of [123I]mIBG uptake in the adrenal glands treated with labetalol, a known β-adrenergic receptor blocker. Upon intravenous administration of [123I]mIBG to mice, SPECT/CT images were acquired over time to confirm the in vivo distribution pattern, revealing a clear uptake in the adrenal glands. Labetalol inhibited the uptake of [123I]mIBG in cell lines expressing NET. A decrease in [123I]mIBG uptake in the adrenal glands was observed in the labetalol-treated group compared with the normal group through SPECT/CT imaging and biodistribution studies. These results demonstrate that SPECT/CT imaging with [123I]mIBG could be applicable for evaluating the preclinical efficacy of new antihypertensive drug candidates such as labetalol, a β-adrenergic receptor blocker.

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.

Dextromethorphan moderates reward deficiency associated with central serotonin transporter availability in 3,4-methylenedioxy-methamphetamine-treated animals.

The neurotoxicity of 3,4-methylenedioxy-methamphetamine (MDMA) to the serotonergic system is well-documented. Dextromethorphan (DM), an antitussive drug, decreased morphine- or methamphetamine (MA)-induced reward in rats and may prevent MDMA-induced serotonergic deficiency in primates, as indicated by increased serotonin transporter (SERT) availability. We aimed to investigate the effects of DM on reward, behavioral sensitization, and neurotoxicity associated with loss of SERT induced by chronic MDMA administration in rats. Conditioned place preference (CPP) and locomotor activity tests were used to evaluate drug-induced reward and behavioral sensitization; 4-[ 18 F]-ADAM/animal-PET and immunohistochemistry were used to explore the effects of DM on MDMA-induced loss of SERT. MDMA significantly reduced SERT binding in the rat brain; however, co-administration of DM significantly restored SERT, enhancing the recovery rate at day 14 by an average of ~23% compared to the MDMA group. In confirmation of the PET findings, immunochemistry revealed MDMA reduced SERT immunoactivity in all brain regions, whereas DM markedly increased the serotonergic fiber density after MDMA induction. Behavioral tests and in vivo longitudinal PET imaging demonstrated the CPP indexes and locomotor activities of the reward system correlate negatively with PET 4-[ 18 F]ADAM SERT activity in the reward system. Our findings suggest MDMA induces functional abnormalities in a network of brain regions important to decision-making processes and the motivation circuit. DM may exert neuroprotective effects to reverse MDMA-induced neurotoxicity.

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.

Vesicular monoamine transporter-2 inhibitor JPC-141 prevents methamphetamine-induced dopamine toxicity and blocks methamphetamine self-administration in rats

Previous research has demonstrated therapeutic potential for VMAT2 inhibitors in rat models of methamphetamine use disorder. Here, we report on the neurochemical and behavioral effects of 1-(2-methoxyphenethyl)-4-phenethypiperazine (JPC-141), a novel analog of lobelane. JPC-141 potently inhibited (Ki = 52 nM) [3H]dopamine uptake by VMAT2 in striatal vesicles with 50 to 250-fold greater selectivity for VMAT2 over dopamine, norepinephrine and serotonin plasmalemma transporters. Also, JPC-141 was 57-fold more selective for inhibiting VMAT2 over [3H]dofetilide binding to hERG channels expressed by HEK293, suggesting relatively low potential for cardiotoxicity. When administered in vivo to rats, JPC-141 prevented the METH-induced reduction in striatal dopamine content when given either prior to or after a high dose of METH, suggesting a reduction in METH-induced dopaminergic neurotoxicity. In behavioral assays, JPC-141 decreased METH-stimulated locomotor activity in METH-sensitized rats at doses of JPC-141 which did not alter locomotor activity in the saline control group. Moreover, JPC-141 specifically decreased iv METH self-administration at doses that had no effect on food-maintained responding. These findings support the further development of VMAT2 inhibitors as pharmacotherapies for individuals with methamphetamine use disorder.

Differential DNA methylation in iPSC-derived dopaminergic neurons: a step forward on the role of SNORD116 microdeletion in the pathophysiology of addictive behavior in Prader-Willi syndrome.

A microdeletion including the SNORD116 gene (SNORD116 MD) has been shown to drive the Prader-Willi syndrome (PWS) features. PWS is a neurodevelopmental disorder clinically characterized by endocrine impairment, intellectual disability and psychiatric symptoms such as a lack of emotional regulation, impulsivity, and intense temper tantrums with outbursts. In addition, this syndrome is associated with a nutritional trajectory characterized by addiction-like behavior around food in adulthood. PWS is related to the genetic loss of expression of a minimal region that plays a potential role in epigenetic regulation. Nevertheless, the role of the SNORD116 MD in DNA methylation, as well as the impact of the oxytocin (OXT) on it, have never been investigated in human neurons. We studied the methylation marks in induced pluripotent stem-derived dopaminergic neurons carrying a SNORD116 MD in comparison with those from an age-matched adult healthy control. We also performed identical neuron differentiation in the presence of OXT. We performed a genome-wide DNA methylation analysis from the iPSC-derived dopaminergic neurons by reduced-representation bisulfite sequencing. In addition, we performed RNA sequencing analysis in these iPSC-derived dopaminergic neurons differentiated with or without OXT. The analysis revealed that 153,826 cytosines were differentially methylated between SNORD116 MD neurons and control neurons. Among the differentially methylated genes, we determined a list of genes also differentially expressed. Enrichment analysis of this list encompassed the dopaminergic system with COMT and SLC6A3. COMT displayed hypermethylation and under-expression in SNORD116 MD, and SLC6A3 displayed hypomethylation and over-expression in SNORD116 MD. RT-qPCR confirmed significant over-expression of SLC6A3 in SNORD116 MD neurons. Moreover, the expression of this gene was significantly decreased in the case of OXT adjunction during the differentiation. SNORD116 MD dopaminergic neurons displayed differential methylation and expression in the COMT and SLC6A3 genes, which are related to dopaminergic clearance.

Molecular Design of SERTlight: A Fluorescent Serotonin Probe for Neuronal Labeling in the Brain.

The serotonergic transmitter system plays fundamental roles in the nervous system in neurotransmission, synaptic plasticity, pathological processes, and therapeutic effects of antidepressants and psychedelics, as well as in the gastrointestinal and circulatory systems. We introduce a novel small molecule fluorescent agent, termed SERTlight, that specifically labels serotonergic neuronal cell bodies, dendrites, and axonal projections as a serotonin transporter (SERT) fluorescent substrate. SERTlight was developed by an iterative molecular design process, based on an aminoethyl-quinolone system, to integrate structural elements that impart SERT substrate activity, sufficient fluorescent brightness, and a broad absence of pharmacological activity, including at serotonin (5-hydroxytryptamine, 5HT) receptors, other G protein-coupled receptors (GPCRs), ion channels, and monoamine transporters. The high labeling selectivity is not achieved by high affinity binding to SERT itself but rather by a sufficient rate of SERT-mediated transport of SERTlight, resulting in accumulation of these molecules in 5HT neurons and yielding a robust and selective optical signal in the mammalian brain. SERTlight provides a stable signal, as it is not released via exocytosis nor by reverse SERT transport induced by 5HT releasers such as MDMA. SERTlight is optically, pharmacologically, and operationally orthogonal to a wide range of genetically encoded sensors, enabling multiplexed imaging. SERTlight enables labeling of distal 5HT axonal projections and simultaneous imaging of the release of endogenous 5HT using the GRAB5HT sensor, providing a new versatile molecular tool for the study of the serotonergic system.

Long COVID-19 and Peripheral Serotonin: A Commentary and Reconsideration.

We believe there are serious problems with a recently published and highly publicized paper entitled "Serotonin reduction in post-acute sequelae of viral infection." The blood centrifugation procedure reportedly used by Wong et al would produce plasma that is substantially (over 95%) depleted of platelets. Given this, their published mean plasma serotonin values of 1.2 uM and 2.4 uM for the control/contrast groups appear to be at least 30 to 60 times too high and should be disregarded. The plasma serotonin values reported for the long COVID and viremia patients also should be disregarded, as should any comparisons to the control/contrast groups. We also note that the plasma serotonin means for the two control/contrast groups are not in good agreement. In the "Discussion" section, Wong et al state that their results tend to support the use of selective serotonin reuptake inhibitors (SSRIs) for the treatment of COVID-19, and they encourage further clinical trials of SSRIs. While they state that, "Our animal models demonstrate that serotonin levels can be restored and memory impairment reversed by precursor supplementation or SSRI treatment", it should be noted that no data are presented showing an increase or restoration in circulating serotonin with SSRI administration. In fact, one would expect a marked decline in platelet serotonin due to SSRIs' effective inhibition of the platelet serotonin transporter. Wong et al hypothesize that problems of long COVID arise from too little peripheral serotonin. However, given the frequent presence of a hyperaggregation state in long COVID, and the known augmenting effects of platelet serotonin on platelet aggregation, it is plausible to suggest that reductions in platelet serotonin might be associated with a lessening of the cardiovascular sequelae of COVID-19.