Bath Salts Research Articles

Active vaccination reduces reinforcing effects of MDPV in male Sprague-Dawley rats trained to self-administer cocaine.

3,4-Methylenedioxypyrovalerone (MDPV) is a synthetic cathinone abused for its cocaine-like psychostimulant effects in "bath salts" products. While there are currently no pharmacotherapies for MDPV abuse, rodent studies suggest immunotherapy may offer a feasible treatment option. These studies tested the capacity of active vaccination to reduce the reinforcing effects of MDPV in Sprague-Dawley rats. Rats acquired cocaine self-administration (0.32 mg/kg/inf) on an FR1 schedule. Dose-effect functions for cocaine (0.032-1.0 mg/kg/inf) and MDPV (0.001-0.32 mg/kg/inf) were determined under an FR5 schedule. Rats in the vaccine group were immunized during cocaine self-administration. All rats transitioned to a progressive-ratio (PR) schedule to establish breakpoints for cocaine (0.1-1.0 mg/kg/inf) and MDPV (0.01-0.32 mg/kg/inf). Responding was extinguished, and cue-induced and MDPV-primed reinstatement (0.56 mg/kg, IP) were evaluated. No endpoints of cocaine self-administration differed between groups, but the ED50 for MDPV self-administration was significantly lower in control relative to vaccinated rats. Under the PR schedule, MDPV was ~ 2.5-fold more potent in maintaining responding in control than vaccinated rats, but Emax was not different between groups. Vaccination did not reduce MDPV-primed reinstatement, perhaps due to a decrease in antibody titer. Vaccination did not alter acquisition of cocaine self-administration, demonstrating pharmacological selectivity and suggesting that the vaccine did not affect learning or motivation, while effectively reducing the potency of MDPV as a reinforcer. The protective effects of the vaccine were surmounted by large unit doses of MDPV, suggesting maximal efficacy of drug-conjugate vaccines in substance abuse disorders will likely require concurrent behavior modification therapy.

Combined effects of mephedrone and cocaine on locomotor activity and conditioned place preference in male Sprague-Dawley rats.

Mephedrone (MEPH) is one of several synthetic cathinone derivatives and a common constituent of illicit 'bath salts'. Concomitant use of MEPH with other psychostimulants is common among recreational users, but their combined effects have not been evaluated rigorously. In experiment 1, 56 male Sprague-Dawley rats were administered saline, MEPH (1 or 5 mg/kg), COC (5 mg/kg), or a mixture of MEPH (1 or 5 mg/kg) + COC (5 mg/kg) for seven consecutive days. Following a 10-day drug washout, rats were given a challenge injection of COC (5 mg/kg). Locomotor activity was recorded for 60 minutes immediately before and for 60 minutes immediately after injections on days 1, 7, and 17. In experiment 2, an unbiased conditioned place preference procedure was implemented over a 10-day period with a separate group of 66 male Sprague-Dawley rats randomly assigned to similar drug treatments used in experiment. Results of experiment 1 indicated significant increases in horizontal activity after repeated treatment with MEPH+COC mixtures, but not with either drug alone. Additionally, rats pretreated with MEPH + COC mixtures exhibited an augmented response to cocaine following drug abstinence. Evidence for CPP was established in rats treated with 5 mg/kg MEPH, 5 mg/kg COC and the 5 mg/kg MEPH + 5 mg/kg COC mixture. In conclusion, cocaine and mephedrone may have additive locomotor stimulant effects, although further assessment with a wider range of dose combinations must be evaluated. As a precautionary note, concurrent use of these substances may pose an enhanced risk for abuse.

3,4-Methylenedioxypyrovalerone: Neuropharmacological Impact of a Designer Stimulant of Abuse on Monoamine Transporters.

Methylenedioxypyrovalerone (MDPV) is an abused synthetic cathinone, commonly referred to as a "bath salt." Because the dopamine (DA) transporter (DAT) and vesicular monoamine transporter-2 (VMAT-2) are key regulators of both the abuse and neurotoxic potential of structurally and behaviorally related agents, the impact of MDPV on these transporters was investigated. Results revealed that a single in vivo MDPV administration rapidly (within 1 hour) and reversibly increased both rat striatal DAT and VMAT-2 activity, as assessed via [3H]DA uptake in synaptosomes and synaptic vesicles, respectively, prepared from treated rats. There was no evidence of an MDPV-induced increase in plasmalemmal membrane DAT surface expression. Plasma concentrations of MDPV increased dose-dependently as assessed 1 hour after 2.5 and 5.0 mg/kg (s.c.) administration and returned to levels less than 10 ng/ml by 18 hours after 2.5 mg/kg (s.c.). Neither pretreatment with a D1 receptor (SCH23390), a D2 receptor (eticlopride), nor a nicotinic receptor (mecamylamine) antagonist attenuated the MDPV-induced increase in DAT activity. In contrast, eticlopride pretreatment attenuated both the MDPV-induced increase in VMAT-2-mediated DA uptake and an associated increase in cytoplasmic-associated vesicle VMAT-2 immunoreactivity. SCH23390 did not attenuate the MDPV-induced increase in VMAT-2 activity. Repeated MDPV injections did not cause persistent DAergic deficits, as assessed 7 to 8 days later. The impact of MDPV on striatal and hippocampal serotonergic assessments was minimal. Taken together, these data contribute to a growing pharmacological rubric for evaluating the ever-growing list of designer cathinone-related stimulants. The profile of MDPV compared with related psychostimulants is discussed. SIGNIFICANCE STATEMENT: Pharmacological characterization of the synthetic cathinone, 3,4-methylenedioxypyrovalerone (MDPV; commonly referred to as a "bath salt"), is critical for understanding the abuse liability and neurotoxic potential of this and related agents. Accordingly, the impact of MDPV on monoaminergic neurons is described and compared with that of related psychostimulants.

Fragmentation pathways of α-pyrrolidinophenone synthetic cathinones and their application to the identification of emerging synthetic cathinone derivatives

The expanding use of emerging synthetic drugs is creating a growing problem for both seized drug analysts and toxicologists because the clandestine suppliers continually tweak the chemical structures to keep one step ahead of the law. Synthetic cathinones, commonly referred to as bath salts, are a specific class of emerging synthetic drugs. These substances are derivatives of cathinone, which is the psychoactive component of the Catha edulis plant, commonly referred to as khat. Of the synthetic cathinone class of compounds, the α-pyrrolidinophenone synthetic cathinone derivatives stand out as one of the most abused designer drugs.The fragmentation behavior of a series of α-pyrrolidinophenone synthetic cathinones was studied with three different ionization and fragmentation techniques to enhance the current understanding of α-pyrrolidinophenone synthetic cathinones in mass spectrometers. Gas chromatography-electron ionization-mass spectrometry (GC-EI-MS) fragmentation is commonly used by seized drug analysts, whereas liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) is more commonly used in toxicological analyses. Direct analysis in real time mass spectrometry (DART-MS) is becoming more popular as a screening technique, especially in national laboratories. Each ionization and activation method encourage particular pathways of fragmentation, and whereas some pathways are conserved across all platforms, other pathways are unique to a particular instrument. This study combines isotope-labeling, multi-stage mass spectrometry (MSn) and accurate mass measurements with high-resolution mass spectrometry (HRMS) to enhance the current understanding about α-pyrrolidinophenone synthetic cathinones. This manuscript provides characteristic protonated tandem mass spectrometry fragmentation pathways and the mechanistic origins of the EI-MS fragmentation observed for this class of synthetic cathinones and provides examples of how this knowledge can be applied to the identification of novel synthetic cathinones.

The Psychiatric Characteristics of People on a Mephedrone (“bath salts”) Binge

Background: Despite solutions presented by the European Union and national regulations introduced by many countries, the problem of mephedrone (4-MMC) is growing. Objectives: The aim of this study was to investigate the effect of regular mephedrone intake with other psychoactive substances on the clinical picture of patients, including self-harms and suicide attempts. Methods: The study involved a group of 601 patients addicted to mephedrone who were admitted to a psychiatric hospital between 2010 and 2018 due to regular mephedrone intake. Results: There was a statistically significant relationship between sleep disorders and mephedrone combined with alcohol (p < .05) or cannabinols (p < .05). However, the highest number of statistically significant correlations was reported when mephedrone was combined with opioids. There was a growing year-on-year percentage of people who attempted suicide because of regular mephedrone intake (p < .001). The more psychoactive substances were combined with mephedrone, the greater the risk of attempted suicide (p < .01). 20% of the examined group were hospitalized several times. Among those hospitalized several times, significantly more people took more than one additional psychoactive substance (p < .01). Conclusions: The more psychoactive substances combined with mephedrone, the more clinical symptoms are associated with it.

Identification of novel fragmentation pathways and fragment ion structures in the tandem mass spectra of protonated synthetic cathinones

The expanding use of emerging synthetic drugs such as synthetic cathinones, or “bath salts”, is a growing public health concern and a continual challenge for drug analysts. In the tandem mass spectra of protonated α-pyrrolidinophenone cathinones, the tropylium ion at m/z 91 is often among the most abundant product ions, but its mechanistic origin is currently unexplained. This project combined electrospray ionization multi-stage mass spectrometry (ESI-MSn), high-resolution mass spectrometry (HRMS), isotopic labeling and ion spectroscopy to enhance our understanding of the fragmentation pathways and mechanisms of a variety of α-pyrrolidinophenone cathinones. The fragmentation trends derived from these ESI-MS/MS studies are: 1) unlike N-alkylated cathinones, abundant radical cations are not observed from even-electron precursors of α-pyrrolidinophenones; 2) the loss of a 71 Da pyrrolidine neutral to form an alkylphenone cation is always observed; 3) a series of neutral alkenes are lost from the alkylphenone cation to form intermediate cations with phthalane-like structures. The phthalane intermediates then eliminate the carbonyl carbon as CO or C2H2O to form a tropylium ion at m/z 91. The α-carbon of the original cathinone is almost exclusively retained in the tropylium ion. If the original cathinone is substituted on the aromatic ring, the observed tropylium ion will be shifted by the mass of the substitution. These findings explain the characteristic ions in ESI-MS/MS spectra of synthetic cathinones and will help analysts better employ mass spectral observations in future casework.

Psychoactive Bath Salts and Neurotoxicity Risk.

Synthetic cathinones are new designer drugs that possess hallucinogenic and psychostimulant properties, and are designed to mimic the effects of illegal substances such as cocaine, amphetamines, and 3.4-methylenedioxymethamphetamine (ecstasy) and to produce rewarding effects, circumventing existing laws and penalties. Synthetic cathinones, also referred to as 'bath salts', have become popular particularly among young people since the mid-2000s. Similar to other psychomotor stimulants, synthetic cathinones have the potential to increase monoamine concentration in the synaptic cleft by targeting the plasma membrane transporters of dopamine, norepinephrine, and serotonin. Because of their structural similarities to amphetamines, it has been suggested that synthetic cathinones may have a neurotoxicity profile similar to that of their amphetamine congeners. Therefore, it has been hypothesized that synthetic cathinones may induce neurotoxicity on monoamine nerve endings in the striatum, hippocampus, and cortex. To date, with regard to synthetic cathinone neurotoxicity, parameters such as monoamine depletion, biosynthetic enzyme inhibition, cytotoxicity, generation of reactive oxygen species, pro-oxidation status, and the ability to induce neuroinflammation were investigated in both in vitro and in vivo experimental studies. Compared with amphetamines, synthetic cathinones appear to have more moderate effects than their amphetamine congeners in terms of neurotoxic effects. However, many synthetic cathinone users take these substances simultaneously with other substances such as benzodiazepines, amphetamines, ecstasy, tetrahydrocannabinol, and ethanol and this abuse can modify their neurotoxic effects. Hence, it is important to understand the underlying mechanism of early neurotoxic effects in case of polysubstance use. In this review, we aimed to present up-to-date information on the abuse potential of synthetic cathinones, their legal status, mechanism of action, and particularly their neurotoxic effects.

Interactions between reinforcement history and drug-primed reinstatement: Studies with MDPV and mixtures of MDPV and caffeine.

Many drugs of abuse are mixed with other psychoactive substances (e.g., caffeine) prior to their sale or use. Synthetic cathinones (e.g., 3,4-methylenedioxypyrovalerone [MDPV]) are commonly mixed with caffeine or other cathinones (e.g., 3,4-methylenedioxy-N-methylcathinone [methylone]), and these "bath salts" mixtures (e.g., MDPV + caffeine) can exhibit supra-additive interactions with regard to their reinforcing and discriminative stimulus properties. However, little is known about relapse-related effects of drug mixtures. In these studies, male Sprague-Dawley rats self-administered 0.032 mg/kg/inf MDPV or a mixture of MDPV + caffeine (0.029 + 0.66 mg/kg/inf, respectively) and then underwent multiple rounds of extinction and reinstatement testing to evaluate the influence of reinforcement history and drug-associated stimuli on the effectiveness of saline (drug-paired stimuli alone), MDPV (0.032-1.0 mg/kg), caffeine (1.0-32 mg/kg), and mixtures of MDPV:caffeine (in 3:1, 1:1, and 1:3 ratios, relative to each drug's ED50 ) to reinstate responding. Dose-addition analyses were used to determine the nature of the drug-drug interaction for each mixture. MDPV and caffeine dose-dependently reinstated responding and were equally effective, regardless of reinforcement history. Most fixed ratio mixtures of MDPV + caffeine exhibited supra-additive interactions, reinstating responding to levels greater than was observed with caffeine and/or MDPV alone. Drug-associated stimuli also played a key role in reinstating responding, especially for caffeine. Together, these results demonstrate that the composition of drug mixtures can impact relapse-related effects of drug mixtures, and "bath salts" mixtures (MDPV + caffeine) may be more effective at promoting relapse-related behaviors than the constituents alone. Further research is needed to determine how other polysubstance reinforcement histories can impact relapse-related behaviors.

Effects of SCH‐23390 and Alpha‐Methyl‐Tyrosine on the Discriminative Stimulus Effects of Mephedrone in Male Rats

The illicit synthetic cathinone mephedrone (4‐methylmethcathinone) often termed a “bath salt” produces discriminative stimulus effects that are similar, but not identical, to those for central nervous system stimulants such as cocaine and methamphetamine. This may be due to its capacity to bind atypically at both dopamine and serotonin transporters compared to those drugs. The purpose of the present study was to further elucidate the mechanism of action underlying mephedrone’s discriminative stimulus effects by administering it with the dopamine type‐1 receptor antagonist SCH‐23390 (n=11) or the catecholamine synthesis inhibitor alpha‐methyl‐tyrosine (alpha‐MT; n=8). Male Long‐Evans rats were trained to discriminate intraperitoneal injections of mephedrone (3.2 mg/kg) from saline under a fixed‐ratio 20 (FR‐20) schedule of food presentation. Mephedrone dose‐effect curves (0.32 – 10 mg/kg) were then established by administering increasing cumulative doses of mephedrone on test days, and comparing those effects with multiple (4–5) injections of saline. The interaction of SCH‐23390 (0.032 – 0.1 mg/kg) and alpha‐MT (200 mg/kg) with mephedrone was examined by administering an acute injection of these drugs prior to increasing cumulative doses of mephedrone. As shown previously by our laboratory, saline administration produced almost exclusively saline‐lever responding while increasing cumulative doses of mephedrone dose‐dependently increased mephedrone‐lever responding. When SCH‐23390 preceded cumulative doses of mephedrone, neither dose alone produced mephedrone‐lever responding and only the larger of the two doses tested (0.1 mg/kg) shifted the mephedrone dose‐effect curve rightward; however, this shift was still less than 2‐fold and produced rate‐decreasing effects when administered alone. Acute administration of alpha‐MT alone also did not produce mephedronelever responding nor decrease rate, and in combination did not produce a marked rightward shift in the mephedrone dose‐effect curve. Taken together, these data indicate that neither SCH‐23390 nor alpha‐MT effectively antagonize the discriminative stimulus effects of mephedrone, and the alpha‐MT data in particular suggest that mephedrone’s discriminative stimulus effects do not depend on newly synthesized catecholamines.

Effects of Ambient Temperature and Social Housing on Locomotor Activity, Thermoregulation, and Weight Loss Following “Binge” Administration of Structurally‐Related Abused Psychostimulants

3,4‐methylenedioxypyrovalerone (MDPV) and α‐pyrrolidinopentiophenone (α‐PVP) are common active constituents of abused “bath salts” products. The structures of these synthetic cathinone analogues differ only in that MDPV contains a methylenedioxy ring. Additionally, MDPV is structurally similar to the amphetamine analogue 3,4‐methylenedioxymethamphetamine (MDMA), which differs fromthe structure of methamphetamine (METH) only by the addition of a methylenedioxy ring. MDPV, α‐PVP, MDMA, and METH are four psychostimulants often used interchangeably by humans in crowded, hot environments. The effects of MDMA are known to be dramatically impacted by crowding and ambient temperature and because of the structural similarities and use patterns in humans, it was of interest to investigate the modulatory effects of such environmental variables on the in vivo pharmacology of the four compounds in mice. In the present studies, adult male NIH Swiss mice were housed singly or in groups of 3 or 6, at 20°C, then administered a “binge” regimen of 1.0, 3.0, or 10.0 mg/kg MDPV, α‐PVP, MDMA or METH, or repeated saline, via intraperitoneal injection, every 2 hours for a total of 4 injections. Locomotor activity and core temperature were continuously collected throughout drug administration, continuing 2 hours after the 4th injection. The lowest dose per social housing condition different from saline was repeated at an elevated temperature of 28°C. Doses were limited by lethal effects which were potentiated both by the increased ambient temperature and social housing conditions. Increased ambient temperature resulted in increased locomotor activity for all compounds, and an increased hyperthermic effect occurred in animals administered MDPV or MDMA, both methylenedioxy ring containing compounds. No pronounced differences regarding body weight and core temperature as a function of social housing were observed. Brain regions (including hippocampus and striatum) were collected for neurochemical analysis 10 days after the last injection. Regions from one hemisphere will be processed and analyzed for tissue content of monoamines and their primary metabolites by HPLC, while samples from the other hemisphere will be utilized to determine GLUT2 concentration via western blotting. These studies suggest that the addition of specific moieties, in this case a methylenedioxy ring, to structurally similiar cathinones and amphetamines can result in same effect compared to the parent compound.Support or Funding InformationThese studies supported by DA039195, DA022981 and the UAMS Center for Translational Neuroscience.

Enantioselectivity on the absorption of methylone and pentedrone using Caco-2 cell line: Development and validation of an UHPLC method for cathinones quantification

Synthetic cathinones, such as methylone and pentedrone, are psychoactive derivatives of cathinone, sold in the internet as “plant food” or “bath salts”.However, the level at which these compounds and their enantiomers cross the intestinal barrier has not been yet determined. Thus, the present study aimed to analyze the enantioselectivity on the permeability of these drugs through the intestinal barrier by using the Caco-2 cell line, a widely used in vitro model for drug permeability studies. To achieve this goal, an UHPLC-UV method was developed and validated to quantify both synthetic cathinones.The developed UHPLC-UV method revealed high selectivity and a linearity from 1 to 500 μM with correlation coefficients always higher than 0.999. The method has an accuracy that ranged between 89 and 107%, inter-day and intra-day precisions with coefficients of variation below 10%, limits of detection and quantification of 0.31 μM and 0.93 μM for methylone and 0.17 μM and 0.52 μM for pentedrone, respectively. In Caco-2 cells, a differentiated passage of the enantiomers across monolayer was observed for both cathinones. For pentedrone, the difference was observed after the first hour, being R-(−)-pentedrone the most permeable compound. Regarding methylone, the difference was noted after one hour and 30 min, with S-(−)-methylone being the most absorbed enantiomer.In conclusion, a fully validated method was successfully applied for studying the permeability of methylone and pentedrone enantiomers in an in vitro model of human intestine, which allowed to discover, for the first time, the enantioselectivity in drug permeability of this class of drugs.

Differential effects of synthetic psychoactive cathinones and amphetamine stimulants on the gut microbiome in mice.

The list of pharmacological agents that can modify the gut microbiome or be modified by it continues to grow at a high rate. The greatest amount of attention on drug-gut microbiome interactions has been directed primarily at pharmaceuticals used to treat infection, diabetes, cardiovascular conditions and cancer. By comparison, drugs of abuse and addiction, which can powerfully and chronically worsen human health, have received relatively little attention in this regard. Therefore, the main objective of this study was to characterize how selected synthetic psychoactive cathinones (aka “Bath Salts”) and amphetamine stimulants modify the gut microbiome. Mice were treated with mephedrone (40 mg/kg), methcathinone (80 mg/kg), methamphetamine (5 mg/kg) or 4-methyl-methamphetamine (40 mg/kg), following a binge regimen consisting of 4 injections at 2h intervals. These drugs were selected for study because they are structural analogs that contain a β-keto substituent (methcathinone), a 4-methyl group (4-methyl-methamphetamine), both substituents (mephedrone) or neither (methamphetamine). Mice were sacrificed 1, 2 or 7 days after treatment and DNA from caecum contents was subjected to 16S rRNA sequencing. We found that all drugs caused significant time- and structure-dependent alterations in the diversity and taxonomic structure of the gut microbiome. The two phyla most changed by drug treatments were Firmicutes (methcathinone, 4-methyl-methamphetamine) and Bacteriodetes (methcathinone, 4-methyl-methamphetamine, methamphetamine, mephedrone). Across time, broad microbiome changes from the phylum to genus levels were characteristic of all drugs. The present results signify that these selected psychoactive drugs, which are thought to exert their primary effects within the CNS, can have profound effects on the gut microbiome. They also suggest new avenues of investigation into the possibility that gut-derived signals could modulate drug abuse and addiction via altered communication along the gut-brain axis.

Void the Analogue Act

This article defends Justice Neil Gorsuch’s suggestion that the Federal Analogue Act (“Analog Act”), a statute criminalizing all substances “substantially similar” to Schedule I and II narcotics, should be voided for vagueness using the same rationale employed by Johnson v. United States. This article also explores alternative rationale sfor voiding the statute as unconstitutionally vague. Whether under Johnson or traditional void for vagueness doctrine, the Analog Act fails to satisfy criminal due process requirements. In its attempt to draft a law broad enough to prospectively criminalize all possible future narcotics, Congress created a statute so vague as to prevent any advance knowledge of the substances criminalized while also leaving every substance in a superposition of legal states as both legal and illegal. “Substantially similar” possesses no statutory or scientific definition, leading juries to routinely reach opposite conclusions on the same substance’s legality and depriving defendants of advance notice. No scienter requirement cures this vagueness. In McFadden v. United States, the Supreme Court’s attempt to resolve the decades’ old scienter circuit split effectively opened the permissible range of scienter so wide as to permit prosecutions for mundane substances: possessing chocolate while knowing it contains a controlled substance analog permits prosecution for methamphetamine possession. The Analog Act’s vagueness and narrow scope also frustrate prosecutions for even the most dangerous analogs, a failure hobbling law enforcement and directly contributing to the proliferation of synthetic cannabis, the MDMA-like “bath salts,” fentanyl analogs, and the coming crisis of unregulated benzodiazepines. Yet at the same time the Analog Act’s great breadth threatens the emerging hemp industry by criminalizing CBD and other cannabinoids intended by Congress to be exempt from criminal restrictions. As illustrated by Congress’s synthetic cannabis legislation and the DEA’s categorical ban on fentanyl analogs, the Analog Act has long outlived its marginal value as flexible modern substance-specific prohibitions protect against emerging psychoactive substances without trampling constitutional protections.

Influence on the Sweating Function by the Successive Application of Artificial CO2 Bath Salts

Influence on the Sweating Function by the Successive Application of Artificial CO₂ Bath Salts

Amphetamine and the Smart Drug 3,4-Methylenedioxypyrovalerone (MDPV) Induce Generalization of Fear Memory in Rats.

Human studies have consistently shown that drugs of abuse affect memory function. The psychostimulants amphetamine and the “bath salt” 3,4-methylenedioxypyrovalerone (MDPV) increase brain monoamine levels through a similar, yet not identical, mechanism of action. Findings indicate that amphetamine enhances the consolidation of memory for emotional experiences, but still MDPV effects on memory function are underinvestigated. Here, we tested the effects induced by these two drugs on generalization of fear memory and their relative neurobiological underpinnings. To this aim, we used a modified version of the classical inhibitory avoidance task, termed inhibitory avoidance discrimination task. According to such procedure, adult male Sprague–Dawley rats were first exposed to one inhibitory avoidance apparatus and, with a 1-min delay, to a second apparatus where they received an inescapable footshock. Forty-eight hours later, retention latencies were tested, in a randomized order, in the two training apparatuses as well as in a novel contextually modified apparatus to assess both strength and generalization of memory. Our results indicated that both amphetamine and MDPV induced generalization of fear memory, whereas only amphetamine enhanced memory strength. Co-administration of the β-adrenoceptor antagonist propranolol prevented the effects of both amphetamine and MDPV on the strength and generalization of memory. The dopaminergic receptor blocker cis-flupenthixol selectively reversed the amphetamine effect on memory generalization. These findings indicate that amphetamine and MDPV induce generalization of fear memory through different modulations of noradrenergic and dopaminergic neurotransmission.

Solid-phase extraction based on cation-exchange sorbents followed by liquid chromatography high-resolution mass spectrometry to determine synthetic cathinones in urine

Cathinone derivates have become very popular as legal highs and are easily obtained on the market as “bath salts”, getting around the legislation due to their modified structure based on banned substances. The aim of this project is to evaluate two mixed-mode cation-exchange sorbents in solid-phase extraction (SPE) and their efficacy in pretreating urine samples for the determination of synthetic cathinones by liquid chromatography high-resolution mass spectrometry (LC-HRMS). A method for determining a group of eleven trending synthetic cathinones in urine using SPE followed by LC-HRMS using Orbitrap as analyser was developed and validated. In the extraction step, two different cation-exchange sorbents were evaluated and compared, a weak one (Oasis WCX) and a strong one (Oasis MCX). Better results were obtained for Oasis MCX in terms of recoveries and matrix effects (lower, if a clean-up step was applied). Method quantification limits were set at 0.2 ng mL−1 and method detection limits were between 0.04 and 0.16 ng mL−1. The results obtained enabled the studied compounds to be quantified at the usual levels at which they are present in urine samples. Moreover, these limits were lower than the ones found in the literature using similar extraction strategies, which shows that the reported strategy can be a useful tool in forensic and toxicological analyses.

Successful use of a novel lux® i-Amylose-1 chiral column for enantioseparation of "legal highs" by HPLC.

Bath salts, fumigations, cleaners and air fresheners, behind these terms substances are hidden, which count as "Legal Highs". These fancy names are used to pretend Legal Highs as harmless compounds, to circumvent legal regulations for marketing as well as to increase the sales. Besides classic illicit drugs of synthetic origin such as amphetamines, cocaine and MDMA, the trade of these compounds, also known as new psychoactive substances (NPS), is not uncommon today. In many countries, NPS are still not subject to drug control. Among them, there are stimulants such as new amphetamine derivatives or cathinones, which possess a chiral centre. Little is known about the fact that the two possible enantiomers may differ in their pharmacological effect. The aim of this study was to test a novel HPLC column for the enantioseparation of a set of 112 NPS coming from different chemical groups and collected by internet purchases during the years 2010-2018. The CSP, namely Lux® 5μm i-Amylose-1, LC Column 250 x 4.6mm, was run in normal phase mode under isocratic conditions, UV detection was performed at 245nm and 230nm, injection volume was 10μl and flow rate was 1ml/min. With a mobile phase consisting of n-hexane/isopropanol/diethylamine (90:10:0.1), herein, 79 NPS were resolved into their enantiomers successfully, for 37 of them baseline resolution was achieved. After increase of lipophily of the mobile phase to 99:1:0.1, another 27 compounds were baseline separated. It was found that all separated NPS are traded as racemic compounds.

Charge regulation radically modifies electrostatics in membrane stacks.

Motivated by biological membrane-containing organelles in plants and photosynthetic bacteria, we study charge regulation in a model membrane stack. Considering (de)protonation as the simplest mechanism of charge equilibration between the membranes and with the bathing environment, we uncover a symmetry-broken charge state in the stack with a quasiperiodic effective charge sequence. In the case of a monovalent bathing salt solution our model predicts complex, inhomogeneous charge equilibria depending on the strength of the (de)protonation reaction, salt concentration, and membrane size. Our results shed light on the basic reorganization mechanism of thylakoid membrane stacks.

The psychoactive cathinone derivative pyrovalerone alters locomotor activity and decreases dopamine receptor expression in zebrafish (Danio rerio).

IntroductionPyrovalerone (4‐methyl‐β‐keto‐prolintane) is a synthetic cathinone (beta‐keto‐amphetamine) derivative. Cathinones are a concern as drugs of abuse, as related street drugs such as methylenedioxypyrovalerone have garnered significant attention. The primary mechanism of action of cathinones is to inhibit reuptake transporters (dopamine and norepinephrine) in reward centers of the central nervous system.MethodsWe measured bioenergetic, behavioral, and molecular responses to pyrovalerone (nM‐µM) in zebrafish to evaluate its potential for neurotoxicity and neurological impairment.ResultsPyrovalerone did not induce any mortality in zebrafish larvae over a 3‐ and 24‐hr period; however, seizures were prevalent at the highest dose tested (100 µM). Oxidative phosphorylation was not affected in the embryos, and there was no change in superoxide dismutase 1 expression. Following a 3‐hr treatment to pyrovalerone (1–100 µM), larval zebrafish (6d) showed a dose‐dependent decrease (70%–90%) in total distance moved in a visual motor response (VMR) test. We interrogated potential mechanisms related to the hypoactivity, focusing on the expression of dopamine‐related transcripts as cathinones can modulate the dopamine system. Pyrovalerone decreased the expression levels of dopamine receptor D1 (~60%) in larval zebrafish but did not affect the expression of tyrosine hydroxylase, dopamine active transporter, or any other dopamine receptor subunit examined, suggesting that pyrovalerone may regulate the expression of dopamine receptors in a specific manner.DiscussionFurther studies using zebrafish are expected to reveal new insight into molecular mechanisms and behavioral responses to cathinone derivates, and zebrafish may be a useful model for understanding the relationship between the dopamine system and bath salts.

Evaluation of the effect of aroma oil as a bath salt on cognitive function.

BackgroundAromatherapy is a non‐pharmacological therapy for the improvement of dementia symptoms. This study aimed to assess the effects of aroma oil as a bath salt on cognitive function, olfactory function and sleep quality.MethodsThis was a randomised controlled trial. Overall, 49 patients were able to provide consent, and 35 were finally analysed (Alzheimer's disease: 10, mild cognitive impairment: 25). The patients were randomly assigned to use 0.1%, 0.5% or 1% aroma bath salt. During daily bathing, bath salt was added to the bath water, and the subjects remained in the bathroom for ≥10 min. The intervention period was 24 weeks, and the observation periods were 4 weeks before and after using the aroma bath salt. We performed the Touch Panel‐type Dementia Assessment Scale (TDAS), the Odour Stick Identification Test for Japanese (OSIT‐J) and the Japanese version of the Pittsburgh Sleep Quality Index (PSQI‐J) five times during the before and after observation periods and after the 12‐week intervention.ResultsThere were no significant changes in the TDAS, OSIT‐J and PSQI‐J scores before and after the intervention in all groups. Moreover, there were no significant differences in the TDAS, OSIT‐J and PSQI‐J scores between the groups before and after the intervention. In the correlation analysis of changes in the TDAS and other tests during the intervention period, significant associations between TDAS and sleep latency and sleep disturbances, which are sub‐items of PSQI‐J, were observed in the use of 0.1% aroma bath salt group.ConclusionsThe use of aroma bath salt was not associated with improvement in cognitive function, olfactory function or sleep quality. However, sleep‐related aspects were associated with changes in cognitive function before and after use of aroma bath salt, which suggested that there is a link between improvements in sleep and that in cognitive function.