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Flakka Drug: Symptoms, Side Effects & Complications

We used an effect-scaling procedure, wherein the unit dose was increased until greater than 10% lethality was observed, to achieve near maximal toxicity (Fantegrossi et al. 2008; Wang et al. 2004). Group two underwent dosing with α-PPP and was assessed in the NOR test four days later and sacrificed for neurochemical analysis the following day. Many studies measure the neurochemical effects of stimulants of abuse during direct drug effects (Ahmed et al., 2004; Mantsch et al., 2004), which does not provide information on lasting brain changes. In contrast, other studies measured neurochemical effects several days after the last drug exposure (Briand et al., 2008; Hadlock et al., 2011; Schwendt et al., 2009), which may be during withdrawal.

The term “man on flakka” does not refer to a specific individual but rather describes the behavior typically exhibited by individuals under the influence of flakka (alpha-PVP), a synthetic cathinone known for inducing extreme and unpredictable actions. Exposure to α-PPP significantly impaired performance in the Y-maze, decreased overall exploratory activity in the novel object recognition test, and resulted in regionally specific depletions in monoamine neurochemistry. In contrast, it had no significant effect on elevated plus maze performance or object discrimination in the novel object recognition test. The locomotor-stimulant effects of α-PPP were comparable to cocaine (30mg/kg), and α-PPP (80mg/kg) did not induce hyperthermia. On January 22, 2013, the US DEA published a request for information specifically regarding 8 additional synthetic cathinones, 2 of the most prominent being 4-methyl-N-ethcathinone (4-MEC) and α‐pyrrolidinopentiophenone (α-PVP) (DEA, 2013). Their similarity in chemical structure suggests that α-PVP and 4-MEC likely emerged as replacements for MDPV and mephedrone, respectively (Figure 1).

The dosing regimen was administered on day 1, and its persistent effects on anxiety, memory, and tissue monoamine neurochemistry were assessed as presented. Group one underwent dosing with α-PPP (80 mg/kg, QID, q2h, IP) and was assessed in the Y-maze and elevated plus maze (EPM) four days later, with EPM assessments occurring at least three hours after the Y-maze assessments. Group two underwent dosing with α-PPP (80 mg/kg, QID, q2h, IP) and was assessed in the NOR test four days later and sacrificed for neurochemical analysis the following day. Amphetamine derivatives induce persistent neurochemical depletions as well as disruption of learning and memory under this dosing regimen (Murnane et al. 2012).

Cell viability and mitochondrial function were measured by assessment of 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) reduction by mitochondrial dehydrogenases after 24- and 72-h exposure to the drugs. A solution of MTT (0.5 mg/ml) was added to the cells, and the culture was incubated for a further 3-h at 37 °C. After aspiration of culture medium, formazan crystals were dissolved in DMSO, and its absorbance was measured at 570 nm using Bio-Rad microplate reader model 680; this value being proportional to the number of cells with intact mitochondria. The mean values for each group were obtained by subtracting the mean OD of the positive control (1% (v/v) Triton-X100, added 30 min before MTT) from the value. The results are expressed as percentages of the negative control group values, these values being considered 100% viable. In the current study, we assessed the persistent behavioral and neurochemical effects of exposure to the second-generation synthetic cathinone α-pyrrolidinopropiophenone (α-PPP).

  • Its molecular pharmacology was recently characterized, demonstrating that α-PPP has high affinity for the dopamine and norepinephrine transporters (approximately 1–2 μM), where it functions as a reuptake inhibitor.
  • Given the paucity of literature on the neurotoxic effects of synthetic cathinones, it is difficult to overly generalize or compare our findings to the literature.
  • Sex differences in neurochemistry in LgA groups were largely confined to 5-HIAA levels (Fig. 3).
  • However, neither PVP, PV8, or PV9 lowered the fluorescence anisotropy of the TMA-DPH probe, which reflects the fluidity of the polar head-group portion of the cell membrane (Fig. 8).
  • Statistical analyses were conducted using NCSS (Number Cruncher Statistical Systems, Kaysville, Utah, USA).

Significant reductions were also seen in H9c2(2-1) cells between 10 and 300 μM, and in SH-SY5Y, Hep G2, and RPMI 2650 cells between 25 and 300 μM after 72-h incubation (Fig. 6b). Similarly, after 24-h incubation, 4-MeO-PV8 decreased the viability of all tested cell lines (25–300 μM for SH-SY5Y and RPMI 2650 cells, 200 and 300 μM for Hep G2 and H9c2(2-1) cells), with the greatest effect being 59% reduction of the survival for SH-SY5Y, 68% for Hep G2, 87% for RPMI 2650, and 33% for H9c2(2-1). Extending incubation time to 72 h increased the cytotoxicity at 300 μM, leading to the decrease of the viability by 91% for SH-SY5Y, 97% for Hep G2, 98% for RPMI 2650, and 63% for H9c2(2-1). Moreover, a broader concentration range was found to elicit a significant drop in the viability for the Hep G2 (25–300 μM) and H9c2(2-1) (10–300 μM) cell lines, compared to 24-h exposure (Fg. 4c).

Figure 1.

Pyrovalerone derivatives (α-pyrrolidinophenones) form a branch of synthetic cathinones, a second most prominent group of novel psychoactive substances. Although the toxicity of 3,4-MDPV, a progenitor of the α-pyrrolidinophenones, is well described, little is known of the potential cytotoxicity of the new members of this group entering the recreational drug market each year. Additionally, an impact of pyrovalerones on the fluidity of the plasma membrane, as the potential mechanism of their cytotoxicity, was examined. The longer side-chain α-pyrrolidinophenones and their fluoro- and methoxy-analogs produce more pronounced maximal cytotoxicity, with regard to mitochondrial activity and cell membrane integrity, than the five-carbon α-PVP and its substituted derivatives.

What are the symptoms of flakka abuse?

Neurotransmitter levels were measured to investigate if neuronal signaling changed as a function of duration of synthetic cathinone exposure and modeled different stages of drug abuse (Koob and Volkow, 2010). 4MMC shows escalation of self-administration during LgA conditions (Nguyen et al., 2017b; Watterson et al., 2014). In H9c2(2-1) cells, the viability was reduced by 76–78% at 200 μM and by approximately 96% at 300 μM, irrespective of the incubation time. In the concentration range of 10–300 μM, significant cytotoxic effects were observed in SH-SY5Y, Hep G2, and RPMI 2650 cells after 24-h incubation, and in SH-SY5Y and RPMI 2650 cells after 72-h incubation. In H9c2(2-1) cells, significant effects were observed at 100–300 μM, irrespective of the incubation time (Fig. 6c). Substituted analogs differ from native PV8 as they affect H9c2(2-1) cell viability even after 24 h.

Despite the DEA’s efforts to ban flakka, the drug is still being manufactured and sold illegally, often through online vendors. The core chemical compound used to make flakka, alpha-PVP, is still legal in many parts of the world. This section collects any data citations, data availability statements, or supplementary materials included in this article. Standards for GLU analysis were prepared by weighing approximately 25 mg of GLU (Alfa Aesar, Ward Hill, MA).

Materials and Methods

Importantly, the more lipophilic PV8 and PV9 evoked changes in the membrane fluidity across a broader concentration range than PVP, an observation that is in line with the fact that disturbances were found in the alpha-pyrrolidinopentiophenone function internal, highly lipophilic part of the membrane but not in the external polar head-groups. PVP and its substituted counterparts produced only benign damage to the cell membranes after 48 h incubation (Fig. 3). In SH-SY5Y neuroblasts, only 4-F-PVP and 4-MeO-PVP used at 300 μM caused a slight elevation of extracellular LDH activity. In H9c2(2-1) cells the effect was more pronounced and significant at 200 and 300 μM for PVP and 100–300 μM for 4-F-PVP and 4-MeO-PVP. The strongest effect was observed at 300 μM of 4-F-PVP and 4-MeO-PVP, being 43 and 45% of positive control cytotoxicity, respectively (Fig. 3).

Effects of PVP, 4-F-PVP, and 4-MeO-PVP on the Survival of SH-SY5Y, Hep G2, RPMI 2650, and H9c2(2- Cells

The role of serotonin in spontaneous alternations is even more poorly understood than the role of catecholamines. For example, previous studies have shown that depletion of serotonin through provision of a tryptophan-deficient diet to rats results in impaired spontaneous alternations (González-Burgos et al. 1995). In rhesus macaques, provision of a tryptophan deficient diet resulted in significant reductions in cerebrospinal fluid biomarkers of serotonin tone, yet no significant change in recognition memory and significant improvement in spatial working memory (Taffe et al. 2003).

  • For α-PVP, 4-MEC, and METH, higher doses (5, 100, and 3mg/kg, respectively) produced elevations in ICSS threshold values, with only METH producing significant elevations (mean±95% CI; α-PVP, 19.83±38.64; 4-MEC, 28.00±31.72; METH, 84.39±69.48%).
  • This study sought to examine effects of long access (LgA) to α-PVP and 4MMC, using procedures similar to those employed for ShA exposure (Marusich et al., 2019a; Marusich et al., 2019b).
  • There were a few sex differences in GLU levels, but the affected brain region and direction of the effects varied by synthetic cathinone and by self-administration condition (Fig. 3–4).
  • Α-PPP exposure results in persistent changes in exploratory behavior, spatial working memory, and monoamine neurochemistry.
  • For both synthetic cathinones, LgA self-administration enhanced DOPAC/DA compared to ShA synthetic cathinone or ShA saline self-administration in both amygdala and striatum.

LgA females showed lower NE levels than males in hypothalamus, whereas ShA rats showed sex differences in NE levels in the other brain regions. Females had lower NE levels than males in amygdala and thalamus for ShA rats, the latter of which only occurred for 4MMC groups, and higher NE levels than ShA males in hippocampus, PFC, and striatum. There were a few sex differences in GLU levels, but the affected brain region and direction of the effects varied by synthetic cathinone and by self-administration condition (Fig. 3–4). Neurotransmitter and metabolite concentrations for all brain regions and all LgA and naïve rat groups are shown in Fig. There were differences between condition (LgA vs naïve), between males and females, and there were interactions between condition and sex.

Coulometric detection was accomplished with a Thermo Scientific Dionex 6011RS electrode cell, and the signal analyzed on a Thermo Scientific Dionex Chromeleon CDS processing platform. Absolute tissue concentrations (ng/mg) for the monoamine neurotransmitters dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC), serotonin, and norepinephrine, were determined by comparison with external standard curves and corrected for tissue weight, as we have described previously (Murnane et al. 2012). Α-PPP exposure results in persistent changes in exploratory behavior, spatial working memory, and monoamine neurochemistry. This research highlights potential dangers of α-PPP, including potential neurotoxicity, and suggests that the mechanisms underlying the persistent untoward effects of the cathinones may be distinct from those of the amphetamines. Statistical analyses were conducted using NCSS (Number Cruncher Statistical Systems, Kaysville, Utah, USA). In most instances, data from naïve and LgA groups (present study) were statistically analyzed separately from ShA groups (Marusich et al., 2019a; Marusich et al., 2019b) because the neurotransmitter assays were conducted at different times for these studies.

In the present study, to elucidate the persistent effects of exposure to α-PPP, we examined the effects of administration of α-PPP on neurochemistry and behavior. Four days after the dosing regimen, we examined working memory, recognition memory, and anxiety. The following day, we performed an extensive neurochemical profiling across the striatum and prefrontal cortex for monoamine neurochemistry. We hypothesized that exposure to α-PPP would deplete brain levels of dopamine and norepinephrine as well as induce memory deficits and increase anxiety. Rats acquired self-administration of α-PVP and 4MMC, consistent with past studies (Aarde et al., 2015; Nguyen et al., 2017a; Nguyen et al., 2016; Vandewater et al., 2015).

Changes in DA levels in striatum and thalamus are hypothesized to occur during the binge and intoxication stage because activation of the mesolimbic DA system produces acute reinforcing properties of psychostimulants (Koob and Volkow, 2010). Furthermore, the dorsal striatum is involved in escalation of drug taking and compulsive behaviors (Clark et al., 2013; Willuhn et al., 2012). LgA, but not ShA, synthetic cathinone self-administration elevated DOPAC and HVA levels in striatum compared to saline and synthetic cathinone ShA levels (Fig. 5), encompassing the binge and intoxication stage of Koob and Volkow’s model. Interestingly, while the metabolites were altered, DA levels only changed in striatum and thalamus for 4MMC and α-PVP LgA groups, respectively, compared to saline ShA. These findings are consistent with a past study showing that ShA 4MMC self-administration did not alter DA, DOPAC, or HVA in striatum (Motbey et al., 2013).

The present study suggests that 21 days of LgA self-administration models the neurochemistry of the beginning stages of the Koob and Volkow model of dysregulated drug intake (Koob and Le Moal, 2005, 2008; Koob and Volkow, 2010). Most past research using the escalation model of self-administration has used 21 sessions (Anker et al., 2010; Gipson et al., 2011; Kitamura et al., 2006; Marusich et al., 2010; Wee et al., 2007a), or fewer. The identified studies that used more than 21 sessions of self-administration (Ahmed and Koob, 1999; Belin et al., 2009; Greenwell et al., 2009a; Greenwell et al., 2009b) provided very little information on the neurochemical changes that occur beyond 21 sessions. Thus, future research should extend the duration of self-administration past 21 sessions to examine what neurochemical changes transpire and if increasing neurochemical dysregulation occurs with continued drug use as hypothesized. 4-Methylmethcathinone (4MMC; mephedrone) releases dopamine (DA), norepinephrine (NE), and serotonin (5-HT) (Baumann et al., 2012; Cameron et al., 2013; Simmler et al., 2013).

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