Dopaminergic Effects of Major Bath Salt Constituents 3,4-Methylenedioxypyrovalerone (MDPV), Mephedrone, and Methylone Are Enhanced Following Co-exposure.

Designer drug mixtures popularized as "bath salts" often contain the synthetic cathinones 3,4 methylenedioxypyrovalerone (MDPV), mephedrone, and methylone in various combinations. However, most preclinical investigations have only assessed the effects of individual bath salt constituents, and little is known about whether co-exposure to MDPV, mephedrone, and methylone produces significant neuropharmacological interactions. This study evaluated and compared how MDPV, mephedrone, and methylone influence discrete brain tissue dopamine (DA) levels and motor stimulant responses in mice when administered alone and as a ternary mixture. Male adolescent Swiss-Webster mice received intraperitoneal injections of saline or 1 or 10 mg/kg doses of MDPV, mephedrone, or methylone, or a cocktail of all three cathinones at doses of 1, 3.3, or 10 mg/kg each. The effect of each treatment on DA and DA metabolite levels in mesolimbic and nigrostriatal brain tissue was quantified 15 min after a single exposure using HPLC-ECD. Additionally, locomotor activity was recorded in mice after acute (day 1) and chronic intermittent (day 7) dosing. MDPV, mephedrone, and methylone produced dose-related increases in mesolimbic and nigrostriatal DA levels that were significantly enhanced following their co-administration. In addition, mice treated with the cathinone cocktail displayed decreased locomotor activity on day 1 that was exacerbated by day 7 and not observed with any of the drugs alone. Our findings demonstrate a significant enhanced effect of MDPV, mephedrone, and methylone on both DA, and these effects on DA result in significant alterations in locomotor activity.

Neurogenesis within the hippocampus after chronic methylphenidate exposure.

Methylphenidate is a psychostimulant used to treat attention deficit hyperactivity disorder. Neurogenesis occurs throughout adulthood within the dentate gyrus of the hippocampus and can be altered by psychoactive medications; however, the impact of methylphenidate on neurogenesis is not fully understood. We investigated the effects of chronic low (1 mg/kg) and high (10 mg/kg) intraperitoneal doses of methylphenidate on neurogenesis in mouse hippocampus following 28 days and 56 days of treatment. Interestingly, methylphenidate, at both doses, increased neurogenesis. However, if methylphenidate treatment was not continued, the newly generated cells did not survive after 28 days. If treatment was continued, the newly generated neurons survived only in the mice receiving low-dose methylphenidate. To investigate the mechanism for this effect, we examined levels of proteins linked to cell proliferation in the hippocampus, including brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), vascular endothelial growth factor (VEGF), tropomyosin receptor kinase B (TrkB), and beta-catenin. BDNF or GDNF levels were not significantly different between groups. However, hippocampal VEGF, TrkB, and beta-catenin were significantly increased in mice receiving low-dose methylphenidate for 28 days compared to controls. Interestingly, high-dose methylphenidate significantly decreased beta-catenin after 28 days and decreased VEGF, beta-catenin, and TrkB after 56 days compared to controls. Thus, low-dose methylphenidate appears to increase cell proliferation and cell survival in the hippocampus, and these effects may be mediated by increase in VEGF, TrkB, and beta-catenin. While high dose methylphenidate may initially increase neuronal proliferation, newly generated neurons are unable to survive long-term, possibly due to decrease in VEGF, TrkB and beta-catenin.

A simple and sensitive high-performance liquid chromatography-electrochemical detection assay for the quantitative determination of monoamines and respective metabolites in six discrete brain regions of mice.

A rapid, sensitive, and reproducible assay is described for the quantitative determination of the monoamine neurotransmitters dopamine, norepinephrine and serotonin, their metabolites, and the internal standard 3,4-dihydroxybenzlyamine hydro-bromide in mouse brain homogenate using high-performance liquid chromatography with electrochemical detection. The method was validated in the following brain areas: frontal cortex, striatum, nucleus accumbens, hippocampus, substantia nigra pars compacta and ventral tegmental area. Biogenic amines and relevant metabolites were extracted from discrete brain regions using a simple protein precipitation procedure, and the chromatography was achieved using a C18 column. The method was accurate over the linear range of 0.300-30 ng/mL (r = 0.999) for dopamine and 0.300-15 ng/mL (r = 0.999) for norepinephrine, 3,4-dihydroxybenzlyamine hydro-bromide, homovanillic acid and 5-hydroxyindolacetic acid, with detection limits of ~0.125 ng/mL (5 pg on column) for each of these analytes. Accuracy and linearity for serotonin were observed throughout the concentration range of 0.625-30 ng/mL (r = 0.998) with an analytical detection limit of ~0.300 ng/mL (12 pg on column). Relative recoveries for all analytes were approximately ≥90% and the analytical run time was <10 min. The described method utilized minimal sample preparation procedures and was optimized to provide the sensitivity limits required for simultaneous monoamine and metabolite analysis in small, discrete brain tissue samples.

Chromatographic and electrophoretic strategies for the chiral separation and quantification of d- and l-threo methylphenidate in biological matrices.

Commercially available methylphenidate (MPH) exists as a racemic mixture composed of the d- and l-threo enantiomers. Various pharmacokinetic studies of MPH have shown a greater pharmacological potency of the d-threo enantiomer. Furthermore, it was deduced that the stereoselective cleavage of MPH to produce ritalinic acid (RA) by human carboxylesterase results in a higher oral bioavailability of the d-threo enantiomer. As a requirement for pharmaceutical regulation authorities, efforts have been made to determine the differential biological distribution of d- and l-threo MPH and RA enantiomers. In support of these efforts, numerous analytical procedures have been developed for the chiral separation and quantification of MPH enantiomers in a variety of biological matrices. The available methodologies accomplish the enantioseparation and quantification of MPH using gas chromatography, liquid chromatography or capillary electrophoretic techniques coupled with a variety of detectors. The current review discusses the technical procedures involved, and the sensitivity and selectivity of these assays.