Selective Dopamine Transporter Research Articles

GMP production of [18F]FE-PE2I on a TRACERLab FX2 N synthesis module, a radiotracer for in vivo PET imaging of the dopamine transport

BackgroundParkinson's disease is a neurodegenerative disorder that is characterized by a degeneration of the dopaminergic system. Dopamine transporter (DAT) positron emission tomography (PET) imaging has emerged as a powerful and non-invasive method to quantify dopaminergic function in the living brain. The PET radioligand, [18F]FE-PE2I, a cocaine chemical derivative, has shown promising properties for in vivo PET imaging of DAT, including high affinity and selectivity for DAT, excellent brain permeability, and favorable metabolism. The aim of the current study was to scale up the production of [18F]FE-PE2I to fulfil the increasing clinical demand for this tracer.ResultsThus, a fully automated and GMP-compliant production procedure has been developed using a commercially available radiosynthesis module GE TRACERLab FX2 N. [18F]FE-PE2I was produced with a radiochemical yield of 39 ± 8% (n = 4, relative [18F]F− delivered to the module). The synthesis time was 70 min, and the molar activity was 925.3 ± 763 GBq/µmol (250 ± 20 Ci/µmol). The produced [18F]FE-PE2I was stable over 6 h at room temperature.ConclusionThe protocol reliably provides a sterile and pyrogen–free GMP-compliant product.

Structure-Activity Relationships for a Recently Controlled Synthetic Cathinone Dopamine Transporter Reuptake Inhibitor: α-Pyrrolidinohexiophenone (α-PHP).

α-Pyrrolidinohexiophenone (α-PHP) is the one-carbon unit α-extended homolog of the better-known and widely abused synthetic cathinone central stimulant α-PVP ("flakka"); both are now U.S. Schedule I controlled substances. Structurally, α-PVP and α-PHP possess a common terminal N-pyrrolidine moiety and differ only with respect to the length of their α-alkyl chain. Using a synaptosomal assay, we previously reported that α-PHP is at least as potent as α-PVP as a dopamine transporter (DAT) reuptake inhibitor. A systematic structure-activity study of synthetic cathinones (e.g., α-PHP) as DAT reuptake inhibitors (i.e., transport blockers), a mechanism thought responsible for their abuse liability, has yet to be conducted. Here, we examined a series of 4-substituted α-PHP analogues and found that, with one exception, all behaved as relatively (28- to >300-fold) selective DAT versus serotonin transporter (SERT) reuptake inhibitors with DAT inhibition potencies of most falling within a very narrow (i.e., <3-fold) range. The 4-CF3 analogue of α-PHP was a confirmed "outlier" in that it was at least 80-fold less potent than the other analogues and displayed reduced (i.e., no) DAT vs SERT selectivity. Consideration of various physicochemical properties of the CF3 group, relative to that of the other substituents involved here, provided relatively little insight. Unlike with DAT-releasing agents, as previously reported by us, a QSAR study was precluded because of the limited range of empirical results (with the exception of the 4-CF3 analogue) for DAT reuptake inhibition.

Behavioral profile of adult zebrafish acutely exposed to a selective dopamine uptake inhibitor, GBR 12909.

The dopamine transporter (DAT) is the main regulator of dopamine concentration in the extrasynaptic space. The pharmacological inhibition of the DAT results in a wide spectrum of behavioral manifestations, which have been identified so far in a limited number of species, mostly in rodents. Here, we used another well-recognized model organism, the zebrafish (Danio rerio), to explore the behavioral effects of GBR 12909, a highly-affine selective DAT blocker. We evaluated zebrafish locomotion, novelty-related exploration, spatial cognition, and social phenotypes in the novel tank, habituation and shoaling tests, following acute 20-min water immersion in GBR 12909. Our findings show hypolocomotion, anxiety-like state, and impaired spatial cognition in fish acutely treated with GBR 12909. This behavioral profile generally parallels that of the DAT knockout rodents and zebrafish, and it overlaps with behavioral effects of other DAT-inhibiting drugs of abuse, such as cocaine and D-amphetamine. Collectively, our data support the utility of zebrafish in translational studies on DAT targeting neuropharmacology and strongly implicate DAT aberration as an important mechanisms involved in neurological and psychiatric diseases.

Longitudinal Positron Emission Tomography Imaging of Presynaptic Terminals in Early Parkinson's Disease.

Imaging tools that allow quantification of Parkinson's disease (PD) progression could facilitate the development of disease-modifying therapies. Cross-sectional studies have shown presynaptic terminal damage in PD patients, but longitudinal data are limited. The aim of this study was to longitudinally assess loss of presynaptic terminals in general and dopaminergic presynaptic terminals in particular as measures of disease progression in early PD. A total of 27 patients with early PD and 18 age- and sex-matched healthy controls underwent positron emission tomography (PET) with 11 C-UCB-J, a ligand for the brain-wide presynaptic terminal marker SV2A, and with 18 F-FE-PE2I, a highly selective dopamine transporter ligand, in combination with a comprehensive motor and non-motor clinical assessment at baseline (BL) and after 26.5± 2.1 months (Y2). SUVR-1 images were calculated and volumes of interest were delineated based on individual 3D T1 magnetic resonance imaging (MRI). PD patients showed significant 2-year worsening of Movement Disorder Society-sponsored revision of the Unified Parkinson's Disease Rating Scale Part III (MDS-UPDRS-III) (off medication) scores, but not of non-motor scores. Motor and non-motor scores in controls did not change significantly over 2 years. 18 F-FE-PE2I binding in caudate and putamen showed significant 2-year decline in the PD group and remained unchanged in controls. Longitudinal decline of striatal 18 F-FE-PE2I binding in PD did not correlate with longitudinal changes in MDS-UPDRS-III scores. 11 C-UCB-J PET did not show any region with significant 2-year change in PD or controls. 18 F-FE-PE2I PET showed robust 2-year decline in early PD, but 11 C-UCB-J PET did not. Longitudinal changes in 18 F-FE-PE2I binding did not correlate with clinical motor progression. © 2022 International Parkinson and Movement Disorder Society.

A Novel and Selective Dopamine Transporter Inhibitor, (S)-MK-26, Promotes Hippocampal Synaptic Plasticity and Restores Effort-Related Motivational Dysfunctions.

Dopamine (DA), the most abundant human brain catecholaminergic neurotransmitter, modulates key behavioral and neurological processes in young and senescent brains, including motricity, sleep, attention, emotion, learning and memory, and social and reward-seeking behaviors. The DA transporter (DAT) regulates transsynaptic DA levels, influencing all these processes. Compounds targeting DAT (e.g., cocaine and amphetamines) were historically used to shape mood and cognition, but these substances typically lead to severe negative side effects (tolerance, abuse, addiction, and dependence). DA/DAT signaling dysfunctions are associated with neuropsychiatric and progressive brain disorders, including Parkinson’s and Alzheimer diseases, drug addiction and dementia, resulting in devastating personal and familial concerns and high socioeconomic costs worldwide. The development of low-side-effect, new/selective medicaments with reduced abuse-liability and which ameliorate DA/DAT-related dysfunctions is therefore crucial in the fields of medicine and healthcare. Using the rat as experimental animal model, the present work describes the synthesis and pharmacological profile of (S)-MK-26, a new modafinil analogue with markedly improved potency and selectivity for DAT over parent drug. Ex vivo electrophysiology revealed significantly augmented hippocampal long-term synaptic potentiation upon acute, intraperitoneally delivered (S)-MK-26 treatment, whereas in vivo experiments in the hole-board test showed only lesser effects on reference memory performance in aged rats. However, in effort-related FR5/chow and PROG/chow feeding choice experiments, (S)-MK-26 treatment reversed the depression-like behavior induced by the dopamine-depleting drug tetrabenazine (TBZ) and increased the selection of high-effort alternatives. Moreover, in in vivo microdialysis experiments, (S)-MK-26 significantly increased extracellular DA levels in the prefrontal cortex and in nucleus accumbens core and shell. These studies highlight (S)-MK-26 as a potent enhancer of transsynaptic DA and promoter of synaptic plasticity, with predominant beneficial effects on effort-related behaviors, thus proposing therapeutic potentials for (S)-MK-26 in the treatment of low-effort exertion and motivational dysfunctions characteristic of depression and aging-related disorders.

Low dopamine transporter binding in the nucleus accumbens in geriatric patients with severe depression.

Dysfunction of dopaminergic neurons in the central nervous system is considered to be related to major depressive disorder (MDD). Especially, MDD in geriatric patients is characterized by anhedonia, which is assumed to be associated with reduced dopamine neurotransmission in the reward system. Dopamine transporter (DAT) is considered to reflect the function of the dopamine nerve system. However, previous DAT imaging studies using single photon emission computed tomography or positron emission tomography (PET) have shown inconsistent results. The radioligand [18 F]FE-PE2I for PET enables more precise evaluation of DAT availability. Hence, we aimed to evaluate the DAT availability in geriatric patients with MDD using [18 F]FE-PE2I. Eleven geriatric patients with severe MDD and 27 healthy controls underwent PET with [18 F]FE-PE2I, which has high affinity and selectivity for DAT. Binding potentials (BPND ) in the striatum (caudate and putamen), nucleus accumbens (NAc), and substantia nigra were calculated. BPND values were compared between MDD patients and healthy controls. MDD patients showed significantly lower DAT BPND in the NAc (P = 0.009), and there was a trend of lower BPND in the putamen (P = 0.032) compared to controls. We found low DAT in the NAc and putamen in geriatric patients with severe MDD, which could be related to dysregulation of the reward system.

Regional Effects of Monoamine Transporter Blockers on Extracellular Monoamine Levels

Monoamine transporter inhibitors are used in the treatment of a number of psychiatric disorders, including major depression and ADHD, and have high abuse potential. These drugs increase extracellular levels of monoamines by blocking the reuptake of monoamines through their respective transporters. Although selective norepinephrine (NE), dopamine (DA) and serotonin (5‐HT) transporter inhibitors increase NE, DA and 5‐HT, respectively, through blockade of their cognate transporter, there is substantial evidence showing that these selective inhibitors can also block the reuptake of monoamines from non‐cognate transporters. For example, studies have shown that norepinephrine transporter (NET) inhibitors can increase extracellular NE and DA levels in the PFC and hippocampus. Studies have also shown increased extracellular NE, DA and 5‐HT levels in the PFC by administration of a selective serotonin reuptake inhibitor (SSRI). Moreover, studies from dopamine transporter (DAT) and serotonin transporter (SERT) KO mice have also shown that selective NET and SERT blockers can increase extracellular DA levels in both the PFC and striatum. Although these non‐selective effects have been reported, there has not been a systematic investigation of brain‐region specific effects of selective monoamine transporter inhibitors on extracellular monoamine levels. We therefore sought to determine the effects of selective inhibitors of monoamine transporters on extracellular monoamine levels in the prefrontal cortex (PFC), nucleus accumbens (NAcc) and dorsal striatum (dStr). We used an ex vivo neurotransmitter release assay to determine the dose‐dependent effects of a NET inhibitor, desipramine, a DAT inhibitor, GRB 12909 and a SERT inhibitor, fluoxetine, on extracellular NE, DA and 5‐HT from PFC, NAcc, and dStr slices of wild‐type (WT) mice. We also examined the dose‐dependent effects of amphetamine on extracellular monoamine levels in the brain regions of interest of WT mice. The findings from WT mice were subsequently compared to those obtained from DAT and NET knockout mice. The results show that selective blockade of NET, DAT and SERT increase extracellular levels of NE, DA, and 5‐HT respectively, as well as alter the extracellular levels of non‐selective monoamines in a brain‐region and genotype specific manner. These findings will guide future studies exploring how region‐specific changes in monoamine levels affect the activity of neuronal circuits, physiology and behavior.Support or Funding InformationBBRF/NARSAD Young Investigator award to Nikhil Urs

Alpha-synuclein/synapsin III pathological interplay boosts the motor response to methylphenidate

Loss of dopaminergic nigrostriatal neurons and fibrillary α-synuclein (α-syn) aggregation in Lewy bodies (LB) characterize Parkinson's disease (PD). We recently found that Synapsin III (Syn III), a phosphoprotein regulating dopamine (DA) release with α-syn, is another key component of LB fibrils in the brain of PD patients and acts as a crucial mediator of α-syn aggregation and toxicity.Methylphenidate (MPH), a monoamine reuptake inhibitor (MRI) efficiently counteracting freezing of gait in advanced PD patients, can bind α-syn and controls α-syn-mediated DA overflow and presynaptic compartmentalization. Interestingly, MPH results also efficient for the treatment of attention deficits and hyperactivity disorder (ADHD), a neurodevelopmental psychiatric syndrome associated with Syn III and α-syn polymorphisms and constituting a risk factor for the development of LB disorders.Here, we studied α-syn/Syn III co-deposition and longitudinal changes of α-syn, Syn III and DA transporter (DAT) striatal levels in nigrostriatal neurons of a PD model, the human C-terminally truncated (1−120) α-syn transgenic (SYN120 tg) mouse, in comparison with C57BL/6J wild type (wt) and C57BL/6JOlaHsd α-syn null littermates. Then, we analyzed the locomotor response of these animals to an acute administration of MPH (d-threo) and other MRIs: cocaine, that we previously found to stimulate Syn III-reliant DA release in the absence of α-syn, or the selective DAT blocker GBR-12935, along aging. Finally, we assessed whether these drugs modulate α-syn/Syn III interaction by fluorescence resonance energy transfer (FRET) and performed in silico studies engendering a heuristic model of the α-syn conformations stabilized upon MPH binding.We found that only MPH was able to over-stimulate a Syn III-dependent/DAT-independent locomotor activity in the aged SYN120 tg mice showing α-syn/Syn III co-aggregates. MPH enhanced full length (fl) α-syn/Syn III and even more (1–120) α-syn/Syn III interaction in cells exhibiting α-syn/Syn III inclusions. Moreover, in silico studies confirmed that MPH may reduce α-syn fibrillation by stabilizing a protein conformation with increased lipid binding predisposition.Our observations indicate that the motor-stimulating effect of MPH can be positively fostered in the presence of α-syn/Syn III co-aggregation. This evidence holds significant implications for PD and ADHD therapeutic management.

Ventral striatum supports Methylphenidate therapeutic effects on impulsive choices expressed in temporal discounting task

Methylphenidate (MPH) is a dopamine transporter (DAT) inhibitor used to treat attention-deficit/hyperactivity-disorder (ADHD). ADHD patients make impulsive choices in delay discounting tasks (DDT) and MPH reduces such impulsivity, but its therapeutic site of action remains unknown. Based on the high density of DAT in the striatum, we hypothesized that the striatum, especially the ventral striatum (VS) and caudate nucleus which both encode temporal discounting, can be preferential MPH action sites. To determine whether one of these striatal territories is predominantly involved in the effect of MPH, we trained monkeys to make choices during DDT. First, consistent with clinical observations, we found an overall reduction of impulsive choices with a low dose of MPH administered via intramuscular injections, whereas we reported sedative-like effects with a higher dose. Then, using PET-imaging, we found that the therapeutic reduction of impulsive choices was associated with selective DAT occupancy of MPH in the VS. Finally, we confirmed the selective involvement of the VS in the effect of MPH by testing the animals’ impulsivity with microinjections of the drug in distinct striatal territories. Together, these results show that the therapeutic effect of MPH on impulsive decisions is mainly restricted to its action in the VS.

Structure-Activity Relationships of Novel Thiazole-Based Modafinil Analogues Acting at Monoamine Transporters.

Atypical dopamine reuptake inhibitors, such as modafinil, are used for the treatment of sleeping disorders and investigated as potential therapeutics against cocaine addiction and for cognitive enhancement. Our continuous effort to find modafinil analogues with higher inhibitory activity on and selectivity toward the dopamine transporter (DAT) has previously led to the promising thiazole-containing derivatives CE-103, CE-111, CE-123, and CE-125. Here, we describe the synthesis and activity of a series of compounds based on these scaffolds, which resulted in several new selective DAT inhibitors and gave valuable insights into the structure-activity relationships. Introduction of the second chiral center and subsequent chiral separations provided all four stereoisomers, whereby the S-configuration on both generally exerted the highest activity and selectivity on DAT. The representative compound of this series was further characterized by in silico, in vitro, and in vivo studies that have demonstrated both safety and efficacy profile of this compound class.

Assessment of Striatal Dopamine Transporter Binding in Individuals With Major Depressive Disorder

Major depressive disorder (MDD) might involve dopamine (DA) reductions. The DA transporter (DAT) regulates DA clearance and neurotransmission and is sensitive to DA levels, with preclinical studies (including those involving inescapable stressors) showing that DAT density decreases when DA signaling is reduced. Despite preclinical data, evidence of reduced DAT in MDD is inconclusive. Using a highly selective DAT positron emission tomography (PET) tracer ([11C] altropane), DAT availability was probed in individuals with MDD who were not taking medication. Levels of DAT expression were also evaluated in postmortem tissues from donors with MDD who died by suicide. This cross-sectional PET study was conducted at McLean Hospital (Belmont, Massachusetts) and Massachusetts General Hospital (Boston) and enrolled consecutive individuals with MDD who were not taking medication and demographically matched healthy controls between January 2012 and March 2014. Brain tissues were obtained from the Douglas-Bell Canada Brain Bank. For the PET component, 25 individuals with current MDD who were not taking medication and 23 healthy controls recruited from McLean Hospital were included (all provided usable data). For the postmortem component, 15 individuals with depression and 14 healthy controls were considered. PET scan. Striatal and midbrain DAT binding potential was assessed. For the postmortem component, tyrosine hydroxylase and DAT levels were evaluated using Western blots. Compared with 23 healthy controls (13 women [56.5%]; mean [SD] age, 26.49 [7.26] years), 25 individuals with MDD (19 women [76.0%]; mean [SD] age, 26.52 [5.92] years) showed significantly lower in vivo DAT availability in the bilateral putamen and ventral tegmental area (Cohen d range, -0.62 to -0.71), and both reductions were exacerbated with increasing numbers of depressive episodes. Unlike healthy controls, the MDD group failed to show an age-associated reduction in striatal DAT availability, with young individuals with MDD being indistinguishable from older healthy controls. Moreover, DAT availability in the ventral tegmental area was lowest in individuals with MDD who reported feeling trapped in stressful circumstances. Lower DAT levels (and tyrosine hydroxylase) in the putamen of MDD compared with healthy controls were replicated in postmortem analyses (Cohen d range, -0.92 to -1.15). Major depressive disorder, particularly with recurring episodes, is characterized by decreased striatal DAT expression, which might reflect a compensatory downregulation due to low DA signaling within mesolimbic pathways.

Serotonin transporter inhibition and 5-HT2C receptor activation drive loss of cocaine-induced locomotor activation in DAT Val559 mice

Dopamine (DA) signaling dysfunction is believed to contribute to multiple neuropsychiatric disorders including attention-deficit/hyperactivity disorder (ADHD). The rare DA transporter (DAT) coding substitution Ala559Val found in subjects with ADHD, bipolar disorder and autism, promotes anomalous DA efflux in vitro and, in DAT Val559 mice, leads to increased reactivity to imminent handling, waiting impulsivity, and enhanced motivation for reward. Here, we report that, in contrast to amphetamine and methylphenidate, which induce significant locomotor activation, cocaine administration to these mice elicits no locomotor effects, despite retention of conditioned place preference (CPP). Additionally, cocaine fails to elevate extracellular DA. Given that amphetamine and methylphenidate, unlike cocaine, lack high-affinity interactions with the serotonin (5-HT) transporter (SERT), we hypothesized that the lack of cocaine-induced hyperlocomotion in DAT Val559 mice arises from SERT blockade and augmented 5-HT signaling relative to cocaine actions on wildtype animals. Consistent with this idea, the SERT blocker fluoxetine abolished methylphenidate-induced locomotor activity in DAT Val559 mice, mimicking the effects seen with cocaine. Additionally, a cocaine analog (RTI-113) with greater selectivity for DAT over SERT retains locomotor activation in DAT Val559 mice. Furthermore, genetic elimination of high-affinity cocaine interactions at SERT in DAT Val559 mice, or specific inhibition of 5-HT2C receptors in these animals, restored cocaine-induced locomotion, but did not restore cocaine-induced elevations of extracellular DA. Our findings reveal a significant serotonergic plasticity arising in the DAT Val559 model that involves enhanced 5-HT2C signaling, acting independently of striatal DA release, capable of suppressing the activity of cocaine-sensitive motor circuits.

Involvement of catecholaminergic and GABAAergic mediations in the anxiety-related behavior in long-term powdered diet-fed mice

Dietary habits are important factors which affect metabolic homeostasis and the development of emotion. We have previously shown that long-term powdered diet feeding in mice increases spontaneous locomotor activity and social interaction (SI) time. Moreover, that diet causes changes in the dopaminergic system, especially increased dopamine turnover and decreased dopamine D4 receptor signals in the frontal cortex. Although the increased SI time indicates low anxiety, the elevated plus maze (EPM) test shows anxiety-related behavior and impulsive behavior. In this study, we investigated whether the powdered diet feeding causes changes in anxiety-related behavior. Mice fed a powdered diet for 17 weeks from weaning were compared with mice fed a standard diet (control). The percentage (%) of open arm time and total number of arm entries were increased in powdered diet-fed mice in the EPM test. We also examined the effects of diazepam, benzodiazepine anti-anxiety drug, bicuculline, GABA-A receptor antagonist, methylphenidate, dopamine transporter (DAT) and noradrenaline transporter (NAT) inhibitor, atomoxetine, selective NAT inhibitor, GBR12909, selective DAT inhibitor, and PD168077, selective dopamine D4 receptor agonist, on the changes of the EPM in powdered diet-fed mice. Methylphenidate and atomoxetine are clinically used to treat attention deficit/hyperactivity disorder (ADHD) symptoms. The % of open arm time in powdered diet-fed mice was decreased by treatments of atomoxetine, methylphenidate and PD168077. Diazepam increased the % of open arm time in control diet-fed mice, but not in powdered diet-fed mice. The powdered diet feeding induced a decrease in GABA transaminase, GABA metabolic enzymes, in the frontal cortex. Moreover, the powdered diet feeding induced an increase in NAT expression, but not DAT expression, in the frontal cortex. These results suggest that the long-term powdered diet feeding may cause low anxiety or impulsivity, possibly via noradrenergic and/or dopaminergic, and GABAAergic mediations and increase the risk for onset of ADHD-like behaviors.

Interactions between Cocaine and the Putative Allosteric Dopamine Transporter Ligand SRI-31142.

Drugs that inhibit the dopamine (DA) transporter (DAT) include both therapeutic agents and abused drugs. Recent studies identified a novel series of putative allosteric DAT inhibitors, but the in vivo effects of these compounds are unknown. This study examined the abuse-related behavioral and neurochemical effects produced in rats by SRI-31142 [2-(7-methylimidazo[1,2-a]pyridin-6-yl)-N-(2-phenyl-2-(pyridin-4-yl)ethyl)quinazolin-4-amine], one compound from this series. In behavioral studies, intracranial self-stimulation (ICSS) was used to compare the effects produced by SRI-31142, the abused and nonselective DAT inhibitor cocaine, and the selective DAT inhibitor GBR-12935 [1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)piperazine]. In neurochemical studies, in vivo microdialysis was used to compare the effects of SRI-31142 and cocaine on levels of DA and serotonin in nucleus accumbens (NAc). The effects of SRI-31142 in combination with cocaine were also examined in both procedures. In contrast to cocaine and GBR-12935, SRI-31142 failed to produce abuse-related increases in ICSS or NAc DA; instead, SRI-31142 only decreased ICSS and NAc DA at a dose that was also sufficient to block cocaine-induced increases in ICSS and NAc DA. Pharmacokinetic studies suggested low but adequate brain penetration of SRI-31142, in vitro binding studies failed to identify likely non-DAT targets, and in vitro functional assays failed to confirm DA uptake inhibition in an assay of DAT-mediated fluorescent signals in live cells. These results indicate that SRI-31142 does not produce cocaine-like abuse-related effects in rats. SRI-31142 may have utility to block cocaine effects and may warrant further study as a candidate pharmacotherapy; however, the role of DAT in mediating these effects is unclear, and side effects may be a limiting factor.

Reinforcing Potency and Effectiveness of Synthetic Cathinones: Potency versus Selectivity for DAT

Synthetic cathinones found in “bath salts” preparations interact with monoamine transporters and function as either cocaine‐like monoamine uptake inhibitors or amphetamine‐like substrates. Abuse of these synthetic cathinones has resulted in regulation of some cathinones internationally. Modifications to the chemical structure of regulated cathinones (e.g., MDPV) produce a series of new cathinones (e.g., α‐PVP, α‐PPP, MDPBP, and MDPPP) which may not be subject to existing regulations. Although MDPV is a more potent and effective reinforcer than cocaine, little is known about the reinforcing effects of other synthetic cathinones. Thus, this study examined intravenous self‐administration of cocaine, MDPV, α‐PVP, α‐PPP, MDPBP, and MDPPP to test the hypotheses that 1) stimulants with greater potency to inhibit the dopamine transporter (DAT) are more potent reinforcers, and 2) stimulants with greater selectivity for DAT relative to the serotonin transporter (SERT) are more effective reinforcers. Potencies to inhibit monoamine uptake at DAT and SERT were determined using synaptosomes prepared from rat brains. Separate groups of male Sprague‐Dawley rats (n=12 per group) were used to generate full dose‐response curves under a progressive ratio (PR) schedule of reinforcement for α‐PVP, α‐PPP, MDPBP, and MDPPP; full dose‐response curves for MDPV and cocaine were also generated in each rat (n=48). Rank order potency to inhibit DAT was MDPV &gt; α‐PVP ≈ MDPBP &gt; MDPPP ≈ α‐PPP ≈ cocaine, while rank order DAT/SERT selectivity was α‐PVP &gt; α‐PPP &gt; MDPV &gt; MDPBP &gt; MDPPP &gt; cocaine. The potency to inhibit DAT correlated with the potency of each drug to maintain responding (MDPV &gt; α‐PVP ≈ MDPBP &gt; MDPPP ≈ α‐PPP ≈ cocaine). Relative reinforcing effectiveness as determined by comparing the maximum number of infusions obtained for each drug under a PR schedule of reinforcement was α‐PVP &gt; α‐PPP ≈ MDPV &gt; MDPBP ≈ MDPPP &gt; cocaine, which correlated with DAT/SERT selectivity. These data suggest that (1) potency at DAT is an important determinant of the potency of each drug to function as a reinforcer, (2) each of these synthetic cathinones functions as a more effective reinforcer than cocaine, and (3) DAT/SERT selectivity is related to reinforcing effectiveness and may predict the abuse‐related effects of novel synthetic cathinones in humans.Support or Funding InformationThese experiments were supported by NIH grants R01DA039146 (GTC) and T32DA031115 (BMG) and the Intramural Research Programs of NIDA (MHB, KCR) and NIAAA (KCR).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Simultaneous Multiple MS Binding Assays for the Dopamine, Norepinephrine, and Serotonin Transporters.

In this work, we present label-free, mass-spectrometry-based binding assays (MS Binding Assays), targeting the human dopamine, norepinephrine, and serotonin transporters (hDAT, hNET, and hSERT) in simultaneous binding experiments. Using a validated LC-ESI-MS/MS method for quantification of the selective dopamine transporter inhibitor (R,R)-4-(2-benzhydryloxyethyl)-1-(4-fluorobenzyl)piperidin-3-ol ((R,R)-D-84), the selective norepinephrine transporter inhibitor (S,S)-reboxetine, and the selective serotonin reuptake inhibitor (S)-citalopram, binding affinities at the three monoamine transporters could be characterized simultaneously in a single binding experiment. The performed simultaneous saturation and competition experiments yielded results that are in good accordance with those determined in MS Binding Assays addressing the monoamine transporters individually. The results obtained from this study underscore the potential of MS Binding Assays for simultaneous affinity determination at different targets, which is difficult to accomplish with conventional radioligand binding assays.

11 C-PE2I and 18 F-Dopa PET for assessing progression rate in Parkinson's: A longitudinal study.

18 F-dopa PET measuring aromatic l-amino acid decarboxylase activity is regarded as the gold standard for evaluating dopaminergic function in Parkinson's disease. Radioligands for dopamine transporters are also used in clinical trials and for confirming PD diagnosis. Currently, it is not clear which imaging marker is more reliable for assessing clinical severity and rate of progression. The objective of this study was to directly compare 18 F-dopa with the highly selective dopamine transporter radioligand 11 C-PE2I for the assessment of motor severity and rate of progression in PD. Thirty-three mild-moderate PD patients underwent 18 F-dopa and 11 C-PE2I PET at baseline. Twenty-three were followed up for 18.8 ± 3.4 months. Standard multiple regression at baseline indicated that 11 C-PE2I BPND predicted UPDRS-III and bradykinesia-rigidity scores (P < 0.05), whereas 18 F-dopa Ki did not make significant unique explanatory contributions. Voxel-wise analysis showed negative correlations between 11 C-PE2I BPND and motor severity across the whole striatum bilaterally. 18 F-Dopa Ki clusters were restricted to the most affected putamen and caudate. Longitudinally, negative correlations were found between striatal Δ11 C-PE2I BPND , ΔUPDRS-III, and Δbradykinesia-rigidity, whereas no significant associations were found for Δ18 F-dopa Ki . One cluster in the most affected putamen was identified in the longitudinal voxel-wise analysis showing a negative relationship between Δ11 C-PE2I BPND and Δbradykinesia-rigidity. Striatal 11 C-PE2I appears to show greater sensitivity for detecting differences in motor severity than 18 F-dopa. Furthermore, dopamine transporter decline is closely associated with motor progression over time, whereas no such relationship was found with aromatic l-amino acid decarboxylase. 11 C-PE2I may be more effective for evaluating the efficacy of neuroprotective treatments in PD. © 2017 International Parkinson and Movement Disorder Society.

S-phenylpiracetam, a selective DAT inhibitor, reduces body weight gain without influencing locomotor activity

S-phenylpiracetam is an optical isomer of phenotropil, which is a clinically used nootropic drug that improves physical condition and cognition. Recently, it was shown that S-phenylpiracetam is a selective dopamine transporter (DAT) inhibitor that does not influence norepinephrine (NE) or serotonin (5-HT) receptors. The aim of the present study was to study the effects of S-phenylpiracetam treatment on body weight gain, blood glucose and leptin levels, and locomotor activity. Western diet (WD)-fed mice and obese Zucker rats were treated daily with peroral administration of S-phenylpiracetam for 8 and 12weeks, respectively. Weight gain and plasma metabolites reflecting glucose metabolism were measured. Locomotor activity was detected in an open-field test. S-phenylpiracetam treatment significantly decreased body weight gain and fat mass increase in the obese Zucker rats and in the WD-fed mice. In addition, S-phenylpiracetam reduced the plasma glucose and leptin concentration and lowered hyperglycemia in a glucose tolerance test in both the mice and the rats. S-phenylpiracetam did not influence locomotor activity in the obese Zucker rats or in the WD-fed mice. The results demonstrate that S-phenylpiracetam reduces body weight gain and improves adaptation to hyperglycemia without stimulating locomotor activity. Our findings suggest that selective DAT inhibitors, such as S-phenylpiracetam, could be potentially useful for treating obesity in patients with metabolic syndrome with fewer adverse health consequences compared to other anorectic agents.

The combination of MDPV and ethanol results in decreased cathinone and increased alcohol levels. Study of such pharmacological interaction

Methylenedioxypyrovalerone (MDPV) is a new psychostimulant cathinone acting as a selective dopamine transporter blocker. Due to the concomitant consumption of ethanol (EtOH) and new psychoactive substances, it is of interest to explore a possible pharmacological interaction between MDPV and EtOH. In locomotor activity assays, EtOH (1g/kg i.p.) elicited a reduction in the stimulant effect induced by low doses of MDPV (0.1–0.3mg/kg, s.c.) in rats, jointly with a decrease in blood and brain MDPV concentrations. Experiments in rat liver microsomes showed different effects depending on the [MDPV]/[EtOH] relationship, evidencing, at certain concentrations, the enhancing effect of EtOH on MDPV metabolism. These suggest that EtOH interacts with MDPV at microsomal level, increasing its metabolic rate. The interaction between both substances was also supported by results in plasma EtOH concentration, which were significantly increased by MDPV, in such a manner that EtOH elimination rate was significantly reduced. The possible toxicological impact of this phenomenon deserves further investigation. In contrast, the rewarding properties of MDPV were unaltered by EtOH. Microdialysis experiments verified that, in the NAcc, both substances could also act synergistically, in such a manner that extracellular dopamine concentrations are maintained. Finally, if the psychostimulant effect induced by MDPV decreased with EtOH, it could favor the boosting and re-dosing in search of the desired effects. However, as the rewarding effect of each dose of the substance would not decrease, the addictive liability could increase considerably. Moreover, we must warn about the increase in EtOH concentrations when consumed concomitantly with MDPV.

Modafinil decreases anxiety-like behaviour in zebrafish.

Modafinil (2-((diphenylmethyl)sulfinyl)acetamide), a selective dopamine and norepinephrine transporter inhibitor, is most commonly prescribed for narcolepsy but has gained recent interest for treating a variety of disorders. Zebrafish (Danio rerio) are becoming a model of choice for pharmacological and behavioural research. To investigate the behavioural effects of modafinil on anxiety, we administered doses of 0, 2, 20, and 200 mg/L for 30 minutes then tested zebrafish in the novel approach test. In this test, the fish was placed into a circular arena with a novel object in the center and motion-tracking software was used to quantify the time the fish spent in the outer area of the arena (thigmotaxis zone), middle third of the arena (transition zone) and center of the arena, as well as total distance traveled, immobility and meandering. Modafinil caused a decrease in time spent in the thigmotaxis zone and increased time spent in the transition zone across all doses. Modafinil did not significantly alter the time spent in the center zone (near the novel object), the distance moved, meandering, or the duration of time spent immobile. We also validated this test as a measure of anxiety with the administration of ethanol (1%) which decreased time spent in the thigmotaxis zone and increased time spent in the transition zone. These results suggest that modafinil decreases anxiety-like behaviour in zebrafish.