Analytical characterization and pharmacological evaluation of the new psychoactive substance 4-fluoromethylphenidate (4F-MPH) and differentiation between the (±)-threo- and (±)-erythro- diastereomers

Misuse of (±)-threo-methylphenidate (methyl-2-phenyl-2-(piperidin-2-yl)acetate; Ritalin®, MPH) has long been acknowledged, but the appearance of MPH analogs in the form of ‘research chemicals’ has only emerged in more recent years. 4-Fluoromethylphenidate (4F-MPH) is one of these recent examples and this study presents the identification and analytical characterization of two powdered 4F-MPH products that were obtained from an online vendor in 2015. Interestingly, the products appeared to have originated from two distinct batches given that one product consisted of (±)-threo-4F-MPH isomers whereas the second sample consisted of a mixture of (±)-threo and (±)-erythro 4F-MPH. Monoamine transporter studies using rat brain synaptosomes revealed that the biological activity of the 4F-MPH mixture resided with the (±)-threo- and not the (±)-erythro isomers based on higher potencies determined for blockage of dopamine uptake (IC50 4F-MPHmixture = 66 nM vs. IC50 (±)-threo = 61 nM vs. IC50 (±)-erythro = 8,528 nM) and norepinephrine uptake (IC50 4F-MPHmixture = 45 nM vs. (±)-threo = 31 nM vs. IC50 (±)-erythro = 3,779 nM). In comparison, MPH was three times less potent than (±)-threo-4F-MPH at the dopamine transporter (IC50 = 131 nM) and around 2.5-times less potent at the norepinephrine transporter (IC50 = 83 nM). Both substances were catecholamine selective with IC50 values of 8,805 nM and >10,000 nM for (±)-threo-4F-MPH and MPH at the serotonin transporter. These findings suggest that the psychostimulant properties of (±)-threo-4F-MPH might be more potent in humans than MPH

Fluorinated phenmetrazine “legal highs” act as substrates for high-affinity monoamine transporters of the SLC6 family

A variety of new psychoactive substances (NPS) are appearing in recreational drug markets worldwide. NPS are compounds that target various receptors and transporters in the central nervous system to achieve their psychoactive effects. Chemical modifications of existing drugs can generate NPS that are not controlled by current legislation, thereby providing legal alternatives to controlled substances such as cocaine or amphetamine. Recently, 3-fluorophenmetrazine (3-FPM), a derivative of the anorectic compound phenmetrazine, appeared on the recreational drug market and adverse clinical effects of the drug have been reported. Phenmetrazine is known to elevate extracellular monoamine concentrations by an amphetamine-like mechanism. Here we tested 3-FPM and its positional isomers, 2-FPM and 4-FPM, for their abilities to interact with plasma membrane monoamine transporters for dopamine (DAT), norepinephrine (NET) and serotonin (SERT). We found that 2-, 3- and 4-FPM inhibit uptake mediated by DAT and NET in HEK293 cells with potencies comparable to cocaine (IC50 values 50 µM). Experiments directed at identifying transporter-mediated reverse transport revealed that FPM isomers induce efflux via DAT, NET and SERT in HEK293 cells, and this effect is augmented by the Na+/H+ ionophore monensin. Each FPM evoked concentration-dependent release of monoamines from rat brain synaptosomes. Hence, this study reports for the first time the mode of action for 2-, 3- and 4-FPM and identifies these NPS as monoamine releasers with marked potency at catecholamine transporters implicated in abuse and addiction

3,4-Methylenedioxymethamphetamine (MDMA) neurotoxicity in rats: a reappraisal of past and present findings

RATIONALE: 3,4-Methylenedioxymethamphetamine (MDMA) is a widely abused illicit drug. In animals, high-dose administration of MDMA produces deficits in serotonin (5-HT) neurons (e.g., depletion of forebrain 5-HT) that have been interpreted as neurotoxicity. Whether such 5-HT deficits reflect neuronal damage is a matter of ongoing debate. OBJECTIVE: The present paper reviews four specific issues related to the hypothesis of MDMA neurotoxicity in rats: (1) the effects of MDMA on monoamine neurons, (2) the use of “interspecies scaling” to adjust MDMA doses across species, (3) the effects of MDMA on established markers of neuronal damage, and (4) functional impairments associated with MDMA-induced 5-HT depletions. RESULTS: MDMA is a substrate for monoamine transporters, and stimulated release of 5-HT, NE, and DA mediates effects of the drug. MDMA produces neurochemical, endocrine, and behavioral actions in rats and humans at equivalent doses (e.g., 1–2 mg/kg), suggesting that there is no reason to adjust doses between these species. Typical doses of MDMA causing long-term 5-HT depletions in rats (e.g., 10–20 mg/kg) do not reliably increase markers of neurotoxic damage such as cell death, silver staining, or reactive gliosis. MDMA-induced 5-HT depletions are accompanied by a number of functional consequences including reductions in evoked 5-HT release and changes in hormone secretion. Perhaps more importantly, administration of MDMA to rats induces persistent anxiety-like behaviors in the absence of measurable 5-HT deficits. CONCLUSIONS: MDMA-induced 5-HT depletions are not necessarily synonymous with neurotoxic damage. However, doses of MDMA which do not cause long-term 5-HT depletions can have protracted effects on behavior, suggesting even moderate doses of the drug may pose risks

The psychoactive aminoalkylbenzofuran derivatives, 5-APB and 6-APB, mimic the effects of 3,4-methylenedioxyamphetamine (MDA) on monoamine transmission in male rats

Rationale: The non-medical use of new psychoactive substances (NPS) is a worldwide public health concern. The so-called “benzofury” compounds, 5-(2-aminopropyl)benzofuran (5-APB) and 6-(2-aminopropyl)benzofuran (6-APB), are NPS with stimulant-like properties in human users. These substances are known to interact with monoamine transporters and 5-HT receptors in transfected cells, but less is known about their effects in animal models. Methods: Here, we used in vitro monoamine transporter assays in rat brain synaptosomes to characterize the effects of 5-APB and 6-APB, together with their N-methyl derivatives 5-MAPB and 6-MAPB, in comparison to 3,4-methylenedioxyamphetamine (MDA) and 3,4-methylenedioxymethamphetamine (MDMA). In vivo neurochemical and behavioral effects of 5-APB (0.3 and 1.0 mg/kg, i.v.) and 6-APB (0.3 and 1.0 mg/kg, i.v.) were assessed in comparison to MDA (1.0 and 3.0 mg/kg, i.v.) using microdialysis sampling in the nucleus accumbens of conscious male rats. Results: All four benzofuran derivatives were substrate-type releasers at dopamine transporters (DAT), norepinephrine transporters (NET) and serotonin transporters (SERT) with nanomolar potencies, similar to the profile of effects produced by MDA and MDMA. However, the benzofurans were at least 3-fold more potent than MDA and MDMA at evoking transporter-mediated release. Like MDA, both benzofurans induced dose-related elevations in extracellular dopamine and serotonin in the brain, but benzofurans were more potent than MDA. The benzofuran derivatives also induced profound behavioral activation characterized by forward locomotion which lasted for at least 2 h post-injection. Conclusions: Overall, benzofurans are more potent than MDA in vitro and in vivo, producing sustained stimulant-like effects in rats. These data suggest that benzofuran-type compounds may have abuse liability, and could pose risks for adverse effects, especially if used in conjunction with abused drugs or medications which enhance monoamine transmission in the brain