Role of Organic Cation Transporter 3 and Plasma Membrane Monoamine Transporter in the Rewarding Properties and Locomotor Sensitizing Effects of Amphetamine in Male andFemale Mice

A lack of effective treatment and sex-based disparities in psychostimulant addiction and overdose warrant further investigation into mechanisms underlying the abuse-related effects of amphetamine-like stimulants. Uptake-2 transporters such as organic cation transporter 3 (OCT3) and plasma membrane monoamine transporter (PMAT), lesser studied potential targets for the actions of stimulant drugs, are known to play a role in monoaminergic neurotransmission. Our goal was to examine the roles of OCT3 and PMAT in mediating amphetamine (1 mg/kg)-induced conditioned place preference (CPP) and sensitization to its locomotor stimulant effects, in males and females, using pharmacological, decynium-22 (D22; 0.1 mg/kg, a blocker of OCT3 and PMAT) and genetic (constitutive OCT3 and PMAT knockout (−/−) mice) approaches. Our results show that OCT3 is necessary for the development of CPP to amphetamine in males, whereas in females, PMAT is necessary for the ability of D22 to prevent the development of CPP to amphetamine. Both OCT3 and PMAT appear to be important for development of sensitization to the locomotor stimulant effect of amphetamine in females, and PMAT in males. Taken together, these findings support an important, sex-dependent role of OCT3 and PMAT in the rewarding and locomotor stimulant effects of amphetamine.

(+)-9-Trifluoroethoxy-α-Dihydrotetrabenazine as a Highly Potent Vesicular Monoamine Transporter 2 Inhibitor for Tardive Dyskinesia

Valbenazine and deutetrabenazine are the only two therapeutic drugs approved for tardive dyskinesia based on blocking the action of vesicular monoamine transporter 2 (VMAT2). But there exist demethylated inactive metabolism at the nine position for both them resulting in low availability, and CYP2D6 plays a major role in this metabolism resulting in the genetic polymorphism issue. 9-trifluoroethoxy-dihydrotetrabenazine (13e) was identified as a promising lead compound for treating tardive dyskinesia. In this study, we separated 13e via chiral chromatography and acquired R,R,R-13e [(+)-13e] and S,S,S-13e [(−)-13e], and we investigated their VMAT2-inhibitory activity and examined the related pharmacodynamics and pharmacokinetics properties using in vitro and in vivo models (+)-13e displayed high affinity for VMAT2 (Ki = 1.48 nM) and strongly inhibited [3H]DA uptake (IC50 = 6.11 nM) in striatal synaptosomes. Conversely, its enantiomer was inactive. In vivo, (+)-13e decreased locomotion in rats in a dose-dependent manner. The treatment had faster, stronger, and longer-lasting effects than valbenazine at an equivalent dose. Mono-oxidation was the main metabolic pathway in the liver microsomes and in dog plasma after oral administration, and glucuronide conjugation of mono-oxidized and/or demethylated products and direct glucuronide conjugation were also major metabolic pathways in dog plasma. O-detrifluoroethylation of (+)-13e did not occur. Furthermore, CYP3A4 was identified as the primary isoenzyme responsible for mono-oxidation and demethylation metabolism, and CYP2C8 was a secondary isoenzyme (+)-13e displayed high permeability across the Caco-2 cell monolayer, and it was not a P-glycoprotein substrate as demonstrated by its high oral absolute bioavailability (75.9%) in dogs. Thus, our study findings highlighted the potential efficacy and safety of (+)-13e in the treatment of tardive dyskinesia. These results should promote its clinical development.

Interaction Profiles of Central Nervous System Active Drugs at Human Organic Cation Transporters 1–3 and Human Plasma Membrane Monoamine Transporter

Many psychoactive compounds have been shown to primarily interact with high-affinity and low-capacity solute carrier 6 (SLC6) monoamine transporters for norepinephrine (NET; norepinephrine transporter), dopamine (DAT; dopamine transporter) and serotonin (SERT; serotonin transporter). Previous studies indicate an overlap between the inhibitory capacities of substances at SLC6 and SLC22 human organic cation transporters (SLC22A1–3; hOCT1–3) and the human plasma membrane monoamine transporter (SLC29A4; hPMAT), which can be classified as high-capacity, low-affinity monoamine transporters. However, interactions between central nervous system active substances, the OCTs, and the functionally-related PMAT have largely been understudied. Herein, we report data from 17 psychoactive substances interacting with the SLC6 monoamine transporters, concerning their potential to interact with the human OCT isoforms and hPMAT by utilizing radiotracer-based in vitro uptake inhibition assays at stably expressing human embryonic kidney 293 cells (HEK293) cells. Many compounds inhibit substrate uptake by hOCT1 and hOCT2 in the low micromolar range, whereas only a few substances interact with hOCT3 and hPMAT. Interestingly, methylphenidate and ketamine selectively interact with hOCT1 or hOCT2, respectively. Additionally, 3,4-methylenedioxymethamphetamine (MDMA) is a potent inhibitor of hOCT1 and 2 and hPMAT. Enantiospecific differences of R- and S-α-pyrrolidinovalerophenone (R- and S-α-PVP) and R- and S-citalopram and the effects of aromatic substituents are explored. Our results highlight the significance of investigating drug interactions with hOCTs and hPMAT, due to their role in regulating monoamine concentrations and xenobiotic clearance.

The Cellular Effects of Nicotine and Tobacco Particulate Matter on Monoamine Transporters

<p>Cigarette smoking causes nearly 6 million deaths worldwide every year (WHO, 2011). Current smoking cessation therapies available to the public are only marginally effective (Jorenby, 2006; Balfour et al., 2000), partly due to our incomplete understanding of the molecular biology of smoking addiction. The majority of studies examining the molecular biology of smoking addiction have focused on nicotine alone. However, there is a growing body of evidence that non-nicotinic components of cigarette smoke contribute to smoking addiction. Nicotine has previously been shown to modulate the function of the monoamine transporters, but studies in the literature are often contradictory and this effect is not completely understood (see Danielson et al., 2011 for review). Furthermore, very few studies have examined the effects of non-nicotinic components of tobacco smoke on the monoamine transporters. This thesis has examined the effects of nicotine and a tobacco extract (TPM) on the dopamine, serotonin, and norepinephrine transporters (DAT, NET, and SERT). Changes in monoamine transporter function, protein expression, and mRNA expression were measured ex vivo in discrete regions of the rat brain following chronic and acute in vivo nicotine and TPM treatment, and in vitro nicotine and TPM treatment. We found that nicotine and TPM affect monoamine transporter function, in a time- and dose-dependent manner, and that intact whole brain circuitry is required for these effects to be seen. In particular, nicotine (0.35 mg/kg) and TPM (containing 0.35 mg/kg nicotine) significantly decreased DAT function in the NAc at 30 min. This effect did not result in a corresponding decrease in DAT protein expression and was mediated by nicotinic receptors containing β2 subunits. Furthermore, TPM caused some changes in monoamine transporter function and mRNA expression that were not observed with nicotine alone. In functional studies this effect was particularly seen in the striatum of rats treated with nicotine (0.35 mg/kg) or TPM (containing 0.35 mg/kg nicotine). Overall these data demonstrate that nicotine affects monoamine transporter function in a nicotinic receptor-dependent manner, and that nicotine and TPM have different effects on monoamine transporter function and expression. This is the first study to examine the effects of TPM on monoamine transporter function, and supports previous evidence of a contribution of non-nicotinic components of cigarette smoke to neuroadaptations related to smoking. Findings from this study contribute to knowledge on the molecular biology of smoking addiction, which could in future lead to the development of more effective smoking cessation therapies.</p>

The Cellular Effects of Nicotine and Tobacco Particulate Matter on Monoamine Transporters

<p>Cigarette smoking causes nearly 6 million deaths worldwide every year (WHO, 2011). Current smoking cessation therapies available to the public are only marginally effective (Jorenby, 2006; Balfour et al., 2000), partly due to our incomplete understanding of the molecular biology of smoking addiction. The majority of studies examining the molecular biology of smoking addiction have focused on nicotine alone. However, there is a growing body of evidence that non-nicotinic components of cigarette smoke contribute to smoking addiction. Nicotine has previously been shown to modulate the function of the monoamine transporters, but studies in the literature are often contradictory and this effect is not completely understood (see Danielson et al., 2011 for review). Furthermore, very few studies have examined the effects of non-nicotinic components of tobacco smoke on the monoamine transporters. This thesis has examined the effects of nicotine and a tobacco extract (TPM) on the dopamine, serotonin, and norepinephrine transporters (DAT, NET, and SERT). Changes in monoamine transporter function, protein expression, and mRNA expression were measured ex vivo in discrete regions of the rat brain following chronic and acute in vivo nicotine and TPM treatment, and in vitro nicotine and TPM treatment. We found that nicotine and TPM affect monoamine transporter function, in a time- and dose-dependent manner, and that intact whole brain circuitry is required for these effects to be seen. In particular, nicotine (0.35 mg/kg) and TPM (containing 0.35 mg/kg nicotine) significantly decreased DAT function in the NAc at 30 min. This effect did not result in a corresponding decrease in DAT protein expression and was mediated by nicotinic receptors containing β2 subunits. Furthermore, TPM caused some changes in monoamine transporter function and mRNA expression that were not observed with nicotine alone. In functional studies this effect was particularly seen in the striatum of rats treated with nicotine (0.35 mg/kg) or TPM (containing 0.35 mg/kg nicotine). Overall these data demonstrate that nicotine affects monoamine transporter function in a nicotinic receptor-dependent manner, and that nicotine and TPM have different effects on monoamine transporter function and expression. This is the first study to examine the effects of TPM on monoamine transporter function, and supports previous evidence of a contribution of non-nicotinic components of cigarette smoke to neuroadaptations related to smoking. Findings from this study contribute to knowledge on the molecular biology of smoking addiction, which could in future lead to the development of more effective smoking cessation therapies.</p>