Identifying human diamine sensors for death related putrescine and cadaverine molecules

Pungent chemical compounds originating from decaying tissue are strong drivers of animal behavior. Two of the best-characterized death smell components are putrescine (PUT) and cadaverine (CAD), foul-smelling molecules produced by decarboxylation of amino acids during decomposition. These volatile polyamines act as 'necromones', triggering avoidance or attractive responses, which are fundamental for the survival of a wide range of species. The few studies that have attempted to identify the cognate receptors for these molecules have suggested the involvement of the seven-helix trace amine-associated receptors (TAARs), localized in the olfactory epithelium. However, very little is known about the precise chemosensory receptors that sense these compounds in the majority of organisms and the molecular basis of their interactions. In this work, we have used computational strategies to characterize the binding between PUT and CAD with the TAAR6 and TAAR8 human receptors. Sequence analysis, homology modeling, docking and molecular dynamics studies suggest a tandem of negatively charged aspartates in the binding pocket of these receptors which are likely to be involved in the recognition of these small biogenic diamines

Novel Psychoactive Substances: : the pharmacology of stimulants and hallucinogens

This is an Accepted Manuscript of an article published by Taylor & Francis in Expert Review of Clinical Pharmacology, on March 2016, available online at doi: : http://www.tandfonline.com/doi/full/10.1586/17512433.2016.1167597.There are increasing levels of concern relating to the rapidly evolving novel psychoactive substances/NPS and web markets’ scenarios. The paper aims at providing an overview of the clinical pharmacological issues related to some of the most popular NPS categories, e.g. stimulants and hallucinogens. NPS intake is typically associated with the imbalance of a complex range of neurotransmitter pathways/receptors, namely: dopamine; cannabinoid/CB1; and 5-HT2A. The intake is almost invariably undetectable with standard screening tests. Hence, it may frequently occur that the acute management of NPS misusers will need to focus on decreasing levels of both self/outward-directed aggression and agitation. Benzodiazepines may be considered as first line treatment. Alternatively, propofol and/or antipsychotics can be administered. Focus will be as well on treatment of possible rhabdomyolysis and hyperthermia. Indeed, future studies should inform better tailored management/treatment strategies.Peer reviewedFinal Accepted Versio

Epigenetic Alterations and an Increased Frequency of Micronuclei in Women with Fibromyalgia

Fibromyalgia (FM), characterized by chronic widespread pain, fatigue, and cognitive/mood disturbances, leads to reduced workplace productivity and increased healthcare expenses. To determine if acquired epigenetic/genetic changes are associated with FM, we compared the frequency of spontaneously occurring micronuclei (MN) and genome-wide methylation patterns in women with FM () to those seen in comparably aged healthy controls ( (MN); (methylation)). The mean (sd) MN frequency of women with FM (51.4 (21.9)) was significantly higher than that of controls (15.8 (8.5)) (; df = 1; ). Significant differences ( sites) in methylation patterns were observed between cases and controls considering a 5% false discovery rate. The majority of differentially methylated (DM) sites (91%) were attributable to increased values in the women with FM. The DM sites included significant biological clusters involved in neuron differentiation/nervous system development, skeletal/organ system development, and chromatin compaction. Genes associated with DM sites whose function has particular relevance to FM included BDNF, NAT15, HDAC4, PRKCA, RTN1, and PRKG1. Results support the need for future research to further examine the potential role of epigenetic and acquired chromosomal alterations as a possible biological mechanism underlying FM

Analisi della dinamica in-vitro della 3-Iodiotironamina (T1AM) utilizzando modellistica compartimentale

La 3-Iodiotironamina (T1AM) è un composto endogeno, derivato della tiroide, ottenuto da una serie di degradazioni successive della Tiroxina. L’interesse per le iodotironamine si è acceso a partire dal 2004, in seguito alla dimostrazione che il composto T1AM (3-iodotironamina) deve essere considerato come un ormone vero e proprio. Questa conclusione deriva dalle seguenti considerazioni: • T1AM è un composto endogeno; • T1AM interagisce con specifici recettori, detti TAARs (TraceAmine Associated Receptors), che sono totalmente distinti dai TAR ed appartengono alla classe dei recettori di membrana accoppiati a proteine G (GPCR = G Protein Coupled Receptors); • T1AM produce effetti funzionali su diversi organi ed apparati, in particolare i principali effetti sono una riduzione della temperatura corpore, un aumento del metabolismo dei lipidi a spese di quello dei carboidrati, una modulazione della secrezione di insulina, una risposta cardiaca inotropa e cronotropa negativa ( cioè una riduzione della contrattilità e della frequenza cardiaca). • La sostanza viene degradata, da parte dell’azione in cascata di enzimi quali monoaminossidasi (MAO) e aldeide deidrogenasi, in acido 3-iodiotiroacetico, chiamato comunemente TA1. In particolare, però, i meccanismi che regolano la dinamica di assorbimento- conversione- rilascio della sostanza non sono stati ancora chiariti o semplicemente data una stima quantitativa, sia in-vivo che ex-vivo che in-vitro. • In questo lavoro di tesi mi sono occupato di analizzare la dinamica in-vitro del T1AM, in particolare in colture cellulari statiche di cellule cardiomiocitiche di ratto della linea immortalizzata H9C2. Gli esperimenti eseguiti sono stati campionamenti di T1AM e TA1 sia nel mezzo di coltura che nell’interno delle cellule, per 1440 minuti. Vi sono stati vari esperimenti anche con inibitori della conversione e del trasporto della sostanza. Le concentrazioni sono state misurate utilizzando il tandem HPLC (High Performance Liquid Chromatography) e MS (Mass Spectrometry). In particolare il mio lavoro è stato lo sviluppo di modelli matematici atti a descrivere e correlare i dati sperimentali ottenuti. Nel primo capitolo di questa tesi vi è riassunta la letteratura riguardante il T1AM, così da fornire un quadro completo sulla fisiologia fino ad ora scoperta. Il capitolo due prevede un analisi statistica sui dati sperimentali, in particolare dell’incertezza dovuta al fatto che la misura di concentrazione non è diretta, ma deriva da un modello matematico di misura. Per il calcolo dell’incertezza finale (e quindi del peso) da applicare ad ogni dato sono state usate tecniche derivanti dalla teoria degli errori. Questa parte è stata implementata utilizzando fogli di calcolo in ambiente Microsoft Excel. Sempre nel capitolo due è stato provato ad approssimare i dati tramite modelli parametrici non-compartimentali (principalmente serie di esponenziali), ed ho provato a vedere se da queste semplici descrizioni era possibile trovare parametri macroscopici che descrivevano sia la dinamica della sostanza, che l’effetto indotto su di essi da sostanze aggiuntive quali ad esempio gli inibitori della conversione, e tentare una prima stima delle funzioni di trasferimento e dei tempi caratteristici del fenomeno. Il confronto di questi parametri ha poi avuto atto. Il tutto è stato eseguito con degli script in ambiente MATLAB, utilizzando funzioni provenienti principalmente dalle toolbox: Curve Fitting, Control System, Symbolic Math. Il capitolo 3 raccoglie invece i modelli compartimentali che simulano il sistema e la sua fisiologia. In particolare inizialmente sono stati provati modelli compartimentali di tipo LTI (Lineari Tempo Invarianti), per via del ricco arsenale di strumenti atti alla loro analisi, ed in seguito modelli non-lineari più accurati che più fedelmente simulano la realtà fisiologica. Il software utilizzati in questo caso sono MATLAB e SAAM II. Alla fine del capitolo vengono esposti i risultati ottenuti e vengono proposti sviluppi futuri e proseguimenti naturali del lavoro

Inhibition of the dorsomedial hypothalamus, but not the medullary raphe pallidus, decreases hyperthermia and mortality from MDMA given in a warm environment.

The central mechanisms through which MDMA mediates life-threatening hyperthermia when taken in a warm environment are not well described. It is assumed that MDMA alters normal thermoregulatory circuits resulting in increased heat production through interscapular brown adipose tissue (iBAT) and decreased heat dissipation through cutaneous vasoconstriction. We studied the role of the dorsomedial hypothalamus (DMH) and medullary raphe pallidus (mRPa) in mediating iBAT, tail blood flow, and locomotor effects produced by MDMA. Rats were instrumented with guide cannulas targeting either the DMH or the mRPa-brain regions involved in regulating iBAT and cutaneous vascular beds. In all animals, core temperature and locomotion were recorded with surgically implanted telemetric transmitters; and additionally either iBAT temperature (via telemetric transmitter) or tail artery blood flow (via tail artery Doppler cuff) were also recorded. Animals were placed in an environmental chamber at 32°C and microinjected with either control or the GABA agonist muscimol (80pmol) followed by an intravenous injection of saline or MDMA (7.5 mg kg-1). To prevent undue suffering, a core temperature of 41°C was chosen as the surrogate marker of mortality. Inhibition of the DMH, but not the mRPa, prevented mortality and attenuated hyperthermia and locomotion. Inhibition of either the DMH or the mRPa did not affect iBAT temperature increases or tail blood flow decreases. While MDMA increases iBAT thermogenesis and decreases heat dissipation through cutaneous vasoconstriction, thermoregulatory brain regions known to mediate these effects are not involved. Rather, the finding that inhibiting the DMH decreases both locomotion and body temperature suggests that locomotion may be a key central contributor to MDMA-evoked hyperthermia

A Transcriptome-Led Exploration of Molecular Mechanisms Regulating Somatostatin-Producing D-Cells in the Gastric Epithelium.

The stomach epithelium contains a myriad of enteroendocrine cells that modulate a range of physiological functions, including postprandial secretion of regulatory peptides, gastric motility, and nutrient absorption. Somatostatin (SST)-producing D-cells are present in the oxyntic and pyloric regions of the stomach, and provide a tonic inhibitory tone that regulates activity of neighboring enteroendocrine cells and gastric acid secretion. Cellular mechanisms underlying the effects of regulatory factors on gastric D-cells are poorly defined due to problems in identifying primary D-cells, and uncertainty remains about which stimuli influence D-cells directly. In this study, we introduce a transgenic mouse line, SST-Cre, which upon crossing with Cre reporter strains, facilitates the identification and purification of gastric D-cells, or cell-specific expression of genetically encoded calcium indicators. Populations of D-cells from the gastric antrum and corpus were isolated and analyzed by RNA sequencing and quantitative RT-PCR. The expression of hormones, hormone receptors, neurotransmitter receptors, and nutrient receptors was quantified. Pyy, Gipr, Chrm4, Calcrl, Taar1, and Casr were identified as genes that are highly enriched in D-cells compared with SST-negative cells. Hormone secretion assays performed in mixed gastric epithelial cultures confirmed that SST secretion is regulated by incretin hormones, cholecystokinin, acetylcholine, vasoactive intestinal polypeptide, calcitonin gene-related polypeptide, oligopetides, and trace amines. Cholecystokinin and oligopeptides elicited increases in intracellular calcium in single-cell imaging experiments performed using cultured D-cells. Our data provide the first transcriptomic analysis and functional characterization of gastric D-cells, and identify regulatory pathways that underlie the direct detection of stimuli by this cell type.This work was supported by the Wellcome Trust (WT088357/Z/09/Z and WT084210/Z/07/Z) and the MRC Metabolic Diseases Unit (MRC_MC_UU_12012/3).This is the final version of the article. It first appeared from Endocrine Society via http://dx.doi.org/10.1210/en.2015-1301

Trace amine receptor in GtoPdb v.2025.3

Trace amine-associated receptors were discovered from a search for novel 5-HT receptors [9], where 15 mammalian orthologues were identified and divided into two families. The TA1 receptor (nomenclature as agreed by the NC-IUPHAR Subcommittee for the Trace amine receptor [59]) has affinity for the endogenous trace amines tyramine, β-phenylethylamine and octopamine in addition to the classical amine dopamine [9]. Emerging evidence suggests that TA1 is a modulator of monoaminergic activity in the brain [95] with TA1 and dopamine D2 receptors shown to form constitutive heterodimers when co-expressed [30]. In addition to trace amines, receptors can be activated by amphetamine-like psychostimulants, and endogenous thyronamines

Trace amine receptor in GtoPdb v.2023.1

Trace amine-associated receptors were discovered from a search for novel 5-HT receptors [9], where 15 mammalian orthologues were identified and divided into two families. The TA1 receptor (nomenclature as agreed by the NC-IUPHAR Subcommittee for the Trace amine receptor [58]) has affinity for the endogenous trace amines tyramine, β-phenylethylamine and octopamine in addition to the classical amine dopamine [9]. Emerging evidence suggests that TA1 is a modulator of monoaminergic activity in the brain [94] with TA1 and dopamine D2 receptors shown to form constitutive heterodimers when co-expressed [30]. In addition to trace amines, receptors can be activated by amphetamine-like psychostimulants, and endogenous thyronamines

Pharmacology of novel psychoactive substances

This PhD work consists of an in vitro and in vivo part. In the in vivo part, we investigated the role of dopamine in the acute clinical effects of 3,4-methylenedioxymethamphetamine (MDMA, “ecstasy”) in healthy human subjects. The role of dopamine in the addictive effects of drug of abuse is well established, but whether it contributes to the acute psychotropic effects of MDMA is unclear. In this pharmacological interaction study, we used the dopamine and weak norepinephrine transporter inhibitor bupropion (Stahl et al. 2004) as a pharmacological tool to block the MDMA-induced dopamine release and to study the role of dopamine in the effects of MDMA. We hypothesized that bupropion would decrease the subjective effects of MDMA to the extent that they depend on MDMA-induced release of dopamine. We included 16 healthy human subjects in this double-blind, placebo-controlled, crossover study. Bupropion pretreatment slightly increased MDMA plasma concentration and prolonged but not reduced the subjective effects contrary to our hypothesis. Additionally, bupropion reduced the MDMA-induced elevations in plasma norepinephrine concentrations and the heart rate response to MDMA. These findings support a role for norepinephrine in the MDMA-induced cardiostimulant effects but no role for MDMA-induced transporter-mediated dopamine release in the elevated mood effects after MDMA administration. Possibly, most of the acute psychotropic effects of MDMA are mediated via transporter-mediated release of serotonin and norepinephrine as previously shown (Hysek et al. 2011, Hysek et al. 2012). In the second and main part of this work we characterized the pharmacological profiles of novel psychoactive substances (NPS). Specifically, we studied whether and how potently NPS interacted with the human transporters for norepinephrine, dopamine, and serotonin, stably expressed in human embryonic kidney (HEK293) cells. Additionally, we assessed binding affinity to the serotonin 5-HT1A, 5-HT2A, 5-HT2C-receptors and the activation potency and activation efficacy at 5-HT2A and 5-HT2B receptors. Furthermore, binding to alpha1A/2A-adrenergic, dopamine D1-3, histamine H1 receptors, as well as trace amine-associated receptor 1 (TAAR1) was also assessed. The NPS studied in this project included para-4-halogenated amphetamine derivatives, which were shown to be relatively more serotonergic than their non-4-halogenated counterparts and pyrovaleronering-substituted cathinones, which were highly potent dopamine transporter inhibitors with a high risk for abuse. Para-halogenated drugs (4-fluoroephedrine, 4-fluoroamphetamine, 4-fluoromethamphetamine, 4-fluoromethcathinone, and 4-bromomethcathinone) also released monoamines, similar to MDMA, whereas pyrovalerones were found to be pure uptake inhibitors. Most benzofurans were similar to MDMA but slightly more serotoninergic than MDMA and additionally activated the serotonin 5-HT2B receptor. The last big group of NPS studied in this project, were novel hallucinogens, which predominantly interacted with the 5-HT2A receptor. This serotonin receptor subtype mediates the hallucinogenic and hallucinogenic-like visual effects of classic serotonergic hallucinogens (Vollenweider et al. 1998, Nichols 2004, Halberstadt et al. 2013, Halberstadt et al. 2014, Halberstadt 2015). Compounds tested in this project included the benzodifuran 8-Bromo-2,3,6,7-benzo-dihydro-difuran-ethylamine (2C-B-FLY), 2C-drugs with their highly potent N-(2-methoxy)benzyl (NBOMe)-derivatives, and lysergic acid diethylamide (LSD). Interestingly, NBOMe derivatives displayed higher affinities at the 5-HT2A receptor than LSD, together with a high selectivity for 5-HT2A over the 5-HT1A receptor, contrary to LSD. NBOMes were partial 5-HT2A receptor agonists, similar to LSD. These novel drugs likely carry a high hallucinogenic potential when used recreationally by humans and the high binding to α1A-receptor (Ki < 1µM) may result in additional vasocontrictive and cardiovascular stimulant effects. Taken together, this PhD contributed to the understanding of the role of dopamine in the effects of MDMA, an important recreational substances. Additionally, we characterized the in vitro pharmacology of many novel designer drugs, which will be helpful in the prediction of the clinical toxicological effects of these newly used recreational drugs

Trace amine receptor (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

Trace amine-associated receptors were discovered from a search for novel 5-HT receptors [9], where 15 mammalian orthologues were identified and divided into two families. The TA1 receptor (nomenclature as agreed by the NC-IUPHAR Subcommittee for the Trace amine receptor [53]) has affinity for the endogenous trace amines tyramine, &#946;-phenylethylamine and octopamine in addition to the classical amine dopamine [9]. Emerging evidence suggests that TA1 is a modulator of monoaminergic activity in the brain [90] with TA1 and dopamine D2 receptors shown to form constitutive heterodimers when co-expressed [28]. In addition to trace amines, receptors can be activated by amphetamine-like psychostimulants, and endogenous thyronamines