Class of 2012 Abstract / Specific Aims: The purpose of this study is to identify the incidence of symptoms associated after exposure to “bath salts,” a term for synthetic cathinones in Arizona. Methods: This is a retrospective chart review of reported exposures to synthetic cathinones to the Arizona Poison and Drug Information Center and the Banner Good Samaritan Poison and Drug Information Center. Main Results: There were 306 cases of synthetic cathinone exposures reviewed and 76.5% were males (n=234) and 23.5% were females (n=72). They were ingested, inhaled, snorted, or injected. The mean age of exposure to synthetic cathinones was 29 years old. The most common symptoms included agitation (48.7%), hallucinations (27.1%), confusion (17.6%), hypertension (21.9%), tachycardia (50.6%), CK elevation (17.3%) and chest pain (9.5%). Less frequent symptoms exhibited in synthetic cathinone abuse included other CNS effects, gastrointestinal symptoms, muscular dysfunction, visual disturbances, and respiratory issues. Conclusions: The symptoms exhibited after exposure to synthetic cathinones were mainly neurologic and cardiovascular. In most cases, symptoms were effectively resolved within 24 to 48 hours after treatment with intravenous fluids and benzodiazepines. In some reports, patients were also given oxygen, anti-emetics, sedatives and anti-psychotic medications. Medical outcomes included major (1.6%), moderate (42.2%) and minor effects (26.1%) while 92 patients were lost to follow-up.

Synthetic cathinones represent threatening and high abuse-potential designer drugs. These are analogs of cathinone (the b-keto analog of amphetamine (AMPH)) a naturally occurring stimulant found in the plant Catha Edulis. Methcathinone (MCAT) was the first synthetic analog of cathinone to be identified in 1987 by Glennon and co-workers and it exerted its action primarily through the dopamine transporter (DAT). Most central stimulants exert their action via monomaine transporters by causing either the release (e.g. cathinone analogs such as MCAT) or by preventing the reuptake (e.g. cocaine) of the neurotransmitter dopamine (DA) thus increasing the extracellular synaptosomal concentration of this neurotransmitter. In 2010, a new class of designer cathinone-like drugs called ‘bath salts’, initially a combination of methylenedioxypyrovalerone (MDPV), methylone (methylenedioxymethcathione, MDMC) and mephedrone (MEPH), soared to popularity. It caused extremely detrimental side effects; it was exceedingly popular for its recreational use and posed a threat to public health. At the time, their mechanisms of action were unknown. Our group identified that MDPV produced actions distinct from other cathinone analogs (i.e., it was identified as the first cathinone-like compound to act as a reuptake inhibitor at the dopamine transporter (DAT)). These findings suggested that not all cathinone-like compounds act uniformly and this insinuated that unique structural features on the cathinone scaffold might contribute to different effects at the transporter level. The overall goal of this project was to study the mechanisms of action of synthetic cathinones (including ‘bath salts’) at the monoamine transporters. We investigated the contribution of each of various structural features on the cathinone scaffold (i.e, the terminal amine, a and b positions, and the phenyl ring). We also constructed homology models of the human dopamine and serotonin transporters (hDAT and hSERT respectively) to help explain differences in selectivity between the neurochemical and behavioral aspects of DAT and SERT. Overall we found that structural features contributed to similar or distinctive mechanisms of action and also contributed to selectivity at monoamine transporters. Our studies provide information that can be useful to drug and health regulatory agencies to help prevent, treat, or curb the future abuse of such drugs.

abstract: Globally, addiction to stimulants such as methamphetamine (METH) remains a significant public health problem. Despite decades of research, no approved anti-relapse medications for METH or any illicit stimulant exist, and current treatment approaches suffer from high relapse rates. Recently, synthetic cathinones have also emerged as popular abused stimulants, leading to numerous incidences of toxicity and death. However, contrary to traditional illicit stimulants, very little is known about their addiction potential. Given the high relapse rates and lack of approved medications for METH addiction, chapters 2 and 3 of this dissertation assessed three different glutamate receptor ligands as potential anti-relapse medications following METH intravenous self-administration (IVSA) in rats. In chapters 4 through 7, using both IVSA and intracranial self-stimulation (ICSS) procedures, experiments assessed abuse liability of the popular synthetic cathinones 3,4-Methylenedioxypyrovalerone (MDPV) , methylone, α-pyrrolidinovalerophenone (α-PVP) and 4-methylethylcathinone (4-MEC). Results from these seminal studies suggest that these drugs possess similar abuse potential to traditional illicit stimulants such as METH, cocaine, and 3,4-methylenedioxymethamphetamine (MDMA). Finally, studies outlined in chapter 8 assessed the potential neurotoxic or adverse cognitive effects of METH and MDPV following IVSA procedures for the purpose of identifying potential novel pharmacotherapeutic targets. However, results of these final studies did not reveal neurotoxic or adverse cognitive effects when using similar IVSA procedural parameters that were sufficient for establishing addiction potential, suggesting that these parameters do not allow for sufficient drug intake to produce similar neurotoxicity or cognitive deficits reported in humans. Thus, these models may be inadequate for fully modeling the adverse neural and psychological consequences of stimulant addiction. Together, these studies support the notion for continued research into the abuse liability and toxicity of METH and synthetic cathinones and suggest that refinements to traditional IVSA models are needed for both more effective assessment of potential cognitive and neural deficits induced by these drugs and screening of potentially clinically efficacious pharmacotherapeutics. / Dissertation/Thesis / Doctoral Dissertation Psychology 2014

Earlier this decade, “bath salts” were popularized as legal alternatives to the pyschostimulants cocaine and the amphetamines. These products contained synthetic cathinones including 3,4-methylenedioxypyrovalerone (MDPV), 4-methylmethcathinone (mephedrone), and 3,4-methylenedioxymethcathinone (methylone). Studies indicate that the cathinones have similar pharmacology to controlled psychostimulants, increasing levels of dopamine (DA) in the synaptic cleft. Most preclinical investigations have only assessed the effect of these synthetic cathinones independently; however, case reports and DEA studies indicate that “bath salts” often contain mixtures of these substances. Therefore, in a recent study by our laboratory, we examined effects of individual versus combined exposure to MDPV, mephedrone, and methylone. Interestingly, an enhanced effect on the levels of DA was observed, as well as significant alterations in locomotor activity following co-exposure to the cathinones. In this study, we examine whether the enhanced effects of the drug combination were due to pharmacokinetic (PK) interactions. It is known that many of the same cytochrome P450 (CYP) isoenzymes metabolize each of these three drugs. Therefore, it is probable that the drugs’ PK would differ when administered individually as compared to in combination. We hypothesize that combined exposure to MDPV, mephedrone, and methylone will result in increased drug concentrations and enhanced total drug concentrations when compared to individual administration. The pharmacokinetics of MDPV, mephedrone, and methylone in the brain and plasma were examined following intraperitoneal injection in mice. Briefly, adolescent male Swiss-Webster mice were injected intraperitoneally with either 10 mg/kg MDPV, 10 mg/kg mephedrone, 10 mg/kg methylone, or 10 mg/kg combined MDPV, mephedrone, and methylone. Following injection, brains and plasma were collected at the following time points: 1, 10, 15, 30, 60, and 120 minutes. Samples were then flash-frozen and stored at -70°C until analysis. Drugs were extracted via solid-phase extraction and concentrations were determined using a previously validated and published high pressure-liquid chromatography tandem mass spectrometry (LC-MS/MS) method. Following intraperitoneal administration, all drugs quickly crossed the blood-brain barrier and entered the brain. Peak drug concentrations, time to peak concentration, drug half-lives, and total drug exposure (as measured by area under the curve) are compared when drugs were given individually versus in combination. These data provide insight into the consequences of co-exposure to popular “bath salt” products.

No description available.

Two deaths involving 3, 4-methylenedioxypyrovalerone (MDPV) are reported. MDPV is a synthetic cathinone stimulant found in "bath salts" with neurological and cardiovascular toxicity. Biological specimens were analyzed for MDPV by GC/MS and LC/MS. A White man was found dead with signs of nausea and vomiting after repeatedly abusing bath salts during a weekend binge. Femoral venous blood and urine had MDPV concentrations of 39 ng/mL and 760 ng/mL. The second fatality was a White man with a history of drug and bath salt abuse found dead at a scene in total disarray after exhibiting fits of anger and psychotic behavior. Femoral venous blood and urine had MDPV concentrations of 130 ng/mL and 3800 ng/mL. The blood and urine MDPV concentrations are within the reported recreational concentration ranges (blood 24-241 ng/mL and urine 34-3900 ng/mL). Both decedents' deaths were attributed to relevant natural causes in a setting of MDPV abuse.

Two deaths involving 3, 4-methylenedioxypyrovalerone (MDPV) are reported. MDPV is a synthetic cathinone stimulant found in "bath salts" with neurological and cardiovascular toxicity. Biological specimens were analyzed for MDPV by GC/MS and LC/MS. A White man was found dead with signs of nausea and vomiting after repeatedly abusing bath salts during a weekend binge. Femoral venous blood and urine had MDPV concentrations of 39 ng/mL and 760 ng/mL. The second fatality was a White man with a history of drug and bath salt abuse found dead at a scene in total disarray after exhibiting fits of anger and psychotic behavior. Femoral venous blood and urine had MDPV concentrations of 130 ng/mL and 3800 ng/mL. The blood and urine MDPV concentrations are within the reported recreational concentration ranges (blood 24-241 ng/mL and urine 34-3900 ng/mL). Both decedents' deaths were attributed to relevant natural causes in a setting of MDPV abuse.

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.

New designer drugs are constantly emerging onto the illicit drug market and it is often difficult to validate and maintain comprehensive analytical methods for accurate detection of these compounds. Generally, toxicology laboratories utilize a screening method, such as immunoassay, for the presumptive identification of drugs of abuse. When a positive result occurs, confirmatory methods, such as gas chromatography (GC) or liquid chromatography (LC) coupled with mass spectrometry (MS), are required for more sensitive and specific analyses. In recent years, the need to study the activities of these compounds in screening assays as well as to develop confirmatory techniques to detect them in biological specimens has been recognized. Severe intoxications and fatalities have been encountered with emerging designer drugs, presenting analytical challenges for detection and identification of such novel compounds. The first major task of this research was to evaluate the performance of commercially available immunoassays to determine if designer drugs were cross-reactive. The second major task was to develop and validate a confirmatory method, using LC-MS, to identify and quantify these designer drugs in biological specimens. Cross-reactivity towards the cathinone derivatives was found to be minimal. Several other phenethylamines demonstrated cross-reactivity at low concentrations, but results were consistent with those published by the assay manufacturer or as reported in the literature. Current immunoassay-based screening methods may not be ideal for presumptively identifying most designer drugs, including the “bath salts.” For this reason, an LC-MS based confirmatory method was developed for 32 compounds, including eight cathinone derivatives, with limits of quantification in the range of 1-10 ng/mL. The method was fully validated for selectivity, matrix effects, stability, recovery, precision, and accuracy. In order to compare the screening and confirmatory techniques, several human specimens were analyzed to demonstrate the importance of using a specific analytical method, such as LC-MS, to detect designer drugs in serum as immunoassays lack cross-reactivity with the novel compounds. Overall, minimal cross-reactivity was observed, highlighting the conclusion that these presumptive screens cannot detect many of the designer drugs and that a confirmatory technique, such as the LC-MS, is required for the comprehensive forensic toxicological analysis of designer drugs.

In recent years, the abuse of synthetic cathinones or ‘bath salts’ has become a major public health concern. Although these compounds were initially sold legally and labeled “not for human consumption”, the ‘bath salts’ are psychostimulants, with similar structures and pharmacologic mechanisms to cocaine, the amphetamines, and 3,4 methylendioxymethamphetamine (MDMA, Molly, or Ecstasy). The reported use of these substances by women of child-bearing age highlights the necessity of studies seeking to delineate risks of prenatal exposure. Three popular drugs of this type are methylone, mephedrone, and 3, 4-methylenedioxypyrovalerone (MDPV). Unfortunately, there is currently no information available on the teratogenicity of these compounds, or of the extent to which they cross the placenta. As such, the purpose of this study was to examine the pharmacokinetic profile of the ‘bath salts’ in a pregnancy model. Pregnant mice (E17.5 gestation) were injected intraperitoneally with a cocktail of 5mg/kg methylone, 10mg/kg mephedrone, and 3mg/kg (MDPV) dissolved in sterile saline. Maternal brain, maternal plasma, placenta, and fetal brain were collected at 30s, 1min, 5min, 10min, 15min, 30min, 1h, 2h, 4h, and 8h following injection. Methylone, mephedrone, and MDPV were extracted from tissue by solid phase extraction, and concentrations were determined using a previously validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. Interestingly, all 3 cathinones reached measurable concentrations in the placenta, as well as the fetal brain; in fact, for MDPV, the maximal concentration (Cmax) was highest in fetal brain, while mephedrone's highest Cmax value was achieved in placenta. Additionally, the total drug exposure for all 3 compounds (as represented by area under the curve, AUC) was higher in fetal matrices (placenta and fetal brain) than in maternal matrices (maternal brain and plasma), and the half-lives for the drugs were longer. Given the extensive presence of methylone, mephedrone, and MDPV in the fetal brain following prenatal exposure, fetal risk is definitely a concern. As there are currently no prenatal studies available on the teratogenicity of these agents, pregnant patients should be informed about the potential risks that these substances may have.