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Sample records for dopamine transporters molecular

  1. Clinical and molecular characterisation of hereditary dopamine transporter deficiency syndrome: an observational cohort and experimental study

    PubMed Central

    Kurian, Manju A; Li, Yan; Zhen, Juan; Meyer, Esther; Hai, Nebula; Christen, Hans-Jürgen; Hoffmann, Georg F; Jardine, Philip; von Moers, Arpad; Mordekar, Santosh R; O'Callaghan, Finbar; Wassmer, Evangeline; Wraige, Elizabeth; Dietrich, Christa; Lewis, Timothy; Hyland, Keith; Heales, Simon JR; Sanger, Terence; Gissen, Paul; Assmann, Birgit E; Reith, Maarten EA; Maher, Eamonn R

    2010-01-01

    Summary Background Dopamine transporter deficiency syndrome is the first identified parkinsonian disorder caused by genetic alterations of the dopamine transporter. We describe a cohort of children with mutations in the gene encoding the dopamine transporter (SLC6A3) with the aim to improve clinical and molecular characterisation, reduce diagnostic delay and misdiagnosis, and provide insights into the pathophysiological mechanisms. Methods 11 children with a biochemical profile suggestive of dopamine transporter deficiency syndrome were enrolled from seven paediatric neurology centres in the UK, Germany, and the USA from February, 2009, and studied until June, 2010. The syndrome was characterised by detailed clinical phenotyping, biochemical and neuroradiological studies, and SLC6A3 mutation analysis. Mutant constructs of human dopamine transporter were used for in-vitro functional analysis of dopamine uptake and cocaine-analogue binding. Findings Children presented in infancy (median age 2·5 months, range 0·5–7) with either hyperkinesia (n=5), parkinsonism (n=4), or a mixed hyperkinetic and hypokinetic movement disorder (n=2). Seven children had been initially misdiagnosed with cerebral palsy. During childhood, patients developed severe parkinsonism-dystonia associated with an eye movement disorder and pyramidal tract features. All children had raised ratios of homovanillic acid to 5-hydroxyindoleacetic acid in cerebrospinal fluid, of range 5·0–13·2 (normal range 1·3–4·0). Homozygous or compound heterozygous SLC6A3 mutations were detected in all cases. Loss of function in all missense variants was recorded from in-vitro functional studies, and was supported by the findings of single photon emission CT DaTSCAN imaging in one patient, which showed complete loss of dopamine transporter activity in the basal nuclei. Interpretation Dopamine transporter deficiency syndrome is a newly recognised, autosomal recessive disorder related to impaired dopamine

  2. Vesicular monoamine transporter 2 and dopamine transporter are molecular targets of Pitx3 in the ventral midbrain dopamine neurons

    PubMed Central

    Hwang, Dong-Youn; Hong, Sunghoi; Jeong, Joo-Won; Choi, Sangdun; Kim, Hansoo; Kim, Jangwoo; Kim, Kwang-Soo

    2016-01-01

    Midbrain dopamine (mDA) neurons play critical roles in the regulation of voluntary movement and their dysfunction is associated with Parkinson’s disease. Pitx3 has been implicated in the proper development of mDA neurons in the substantia nigra pars compacta, which are selectively lost in Parkinson’s disease. However, the basic mechanisms underlying its role in mDA neuron development and/or survival are poorly understood. Toward this goal, we sought to identify downstream target genes of Pitx3 by comparing gene expression profiles in mDA neurons of wild-type and Pitx3-deficient aphakia mice. This global gene expression analysis revealed many potential target genes of Pitx3; in particular, the expression of vesicular monoamine transporter 2 and dopamine transporter, responsible for dopamine storage and reuptake, respectively, is greatly reduced in mDA neurons by Pitx3 ablation. In addition, gain-of-function analyses and chromatin immunoprecipitation strongly indicate that Pitx3 may directly activate transcription of vesicular monoamine transporter 2 and dopamine transporter genes, critically contributing to neurotransmission and/or survival of mDA neurons. As the two genes have been known to be regulated by Nurr1, another key dopaminergic transcription factor, we propose that Pitx3 and Nurr1 may coordinately regulate mDA specification and survival, at least in part, through a merging and overlapping downstream pathway. PMID:19780901

  3. Role of Histidine 547 of Human Dopamine Transporter in Molecular Interaction with HIV-1 Tat and Dopamine Uptake

    PubMed Central

    Yuan, Yaxia; Quizon, Pamela M.; Sun, Wei-Lun; Yao, Jianzhuang; Zhu, Jun; Zhan, Chang-Guo

    2016-01-01

    HIV-1 Tat plays an important role in HIV-associated neurocognitive disorders (HAND) by disrupting neurotransmission including dopamine uptake by human dopamine transporter (hDAT). Previous studies have demonstrated that HIV-1 Tat directly binds to hDAT and some amino-acid mutations that attenuate the hDAT-Tat binding also significantly decreased dopamine uptake activity of hDAT. This combined computational-experimental study demonstrates that histidine-547 (H547) of hDAT plays a crucial role in the hDAT-Tat binding and dopamine uptake by hDAT, and that the H547A mutation can not only considerably attenuate Tat-induced inhibition of dopamine uptake, but also significantly increase the Vmax of hDAT for dopamine uptake. The finding of such an unusual hDAT mutant capable of both increasing the Vmax of hDAT for dopamine uptake and disrupting the hDAT-Tat binding may provide an exciting knowledge basis for development of novel concepts for therapeutic treatment of the HAND. PMID:27250920

  4. Role of Histidine 547 of Human Dopamine Transporter in Molecular Interaction with HIV-1 Tat and Dopamine Uptake.

    PubMed

    Yuan, Yaxia; Quizon, Pamela M; Sun, Wei-Lun; Yao, Jianzhuang; Zhu, Jun; Zhan, Chang-Guo

    2016-01-01

    HIV-1 Tat plays an important role in HIV-associated neurocognitive disorders (HAND) by disrupting neurotransmission including dopamine uptake by human dopamine transporter (hDAT). Previous studies have demonstrated that HIV-1 Tat directly binds to hDAT and some amino-acid mutations that attenuate the hDAT-Tat binding also significantly decreased dopamine uptake activity of hDAT. This combined computational-experimental study demonstrates that histidine-547 (H547) of hDAT plays a crucial role in the hDAT-Tat binding and dopamine uptake by hDAT, and that the H547A mutation can not only considerably attenuate Tat-induced inhibition of dopamine uptake, but also significantly increase the Vmax of hDAT for dopamine uptake. The finding of such an unusual hDAT mutant capable of both increasing the Vmax of hDAT for dopamine uptake and disrupting the hDAT-Tat binding may provide an exciting knowledge basis for development of novel concepts for therapeutic treatment of the HAND. PMID:27250920

  5. Insights from molecular dynamics: the binding site of cocaine in the dopamine transporter and permeation pathways of substrates in the leucine and dopamine transporters

    PubMed Central

    Merchant, Bonnie A.; Madura, Jeffry D.

    2012-01-01

    The dopamine transporter (DAT) facilitates the regulation of synaptic neurotransmitter levels. As a target for therapeutic and illicit psycho-stimulant drugs like antidepressants and cocaine, DAT has been studied intensively. Despite a wealth of mutational and physiological data regarding DAT, the structure remains unsolved and details of the transport mechanism, binding sites and conformational changes remain debated. A bacterial homologue of DAT, the leucine transporter (LeuTAa) has been used as a template and framework for modeling and understanding DAT. Free energy profiles obtained from Multi-Configuration Thermodynamic Integration allowed us to correctly identify the primary and secondary binding pockets of LeuTAa. A comparison of free energy profiles for dopamine and cocaine in DAT suggests that the binding site of cocaine is located in a secondary pocket, not the primary substrate site. Two recurring primary pathways for intracellular substrate release from the primary pocket are identified in both transporters using the Random Acceleration Molecular Dynamics method. One pathway appears to follow transmembranes (TMs) 1a and 6b while the other pathway follows along TMs 6b and 8. Interestingly, we observe that a single sodium ion is co-transported with leucine during both simulations types. PMID:23079638

  6. The binding sites for cocaine and dopamine in the dopamine transporter overlap

    PubMed Central

    Beuming, Thijs; Kniazeff, Julie; Bergmann, Marianne L; Shi, Lei; Gracia, Luis; Raniszewska, Klaudia; Newman, Amy Hauck; Javitch, Jonathan A; Weinstein, Harel; Gether, Ulrik; Loland, Claus J

    2009-01-01

    Cocaine is a widely abused substance with psychostimulant effects that are attributed to inhibition of the dopamine transporter (DAT). We present molecular models for DAT binding of cocaine and cocaine analogs constructed from the high-resolution structure of the bacterial transporter homolog LeuT. Our models suggest that the binding site for cocaine and cocaine analogs is deeply buried between transmembrane segments 1, 3, 6 and 8, and overlaps with the binding sites for the substrates dopamine and amphetamine, as well as for benztropine-like DAT inhibitors. We validated our models by detailed mutagenesis and by trapping the radiolabeled cocaine analog [3H]CFT in the transporter, either by cross-linking engineered cysteines or with an engineered Zn2+-binding site that was situated extracellularly to the predicted common binding pocket. Our data demonstrate the molecular basis for the competitive inhibition of dopamine transport by cocaine. PMID:18568020

  7. Introducing Thermal Wave Transport Analysis (TWTA): A Thermal Technique for Dopamine Detection by Screen-Printed Electrodes Functionalized with Molecularly Imprinted Polymer (MIP) Particles.

    PubMed

    Peeters, Marloes M; van Grinsven, Bart; Foster, Christopher W; Cleij, Thomas J; Banks, Craig E

    2016-01-01

    A novel procedure is developed for producing bulk modified Molecularly Imprinted Polymer (MIP) screen-printed electrodes (SPEs), which involves the direct mixing of the polymer particles within the screen-printed ink. This allowed reduction of the sample preparation time from 45 min to 1 min, and resulted in higher reproducibility of the electrodes. The samples are measured with a novel detection method, namely, thermal wave transport analysis (TWTA), relying on the analysis of thermal waves through a functional interface. As a first proof-of-principle, MIPs for dopamine are developed and successfully incorporated within a bulk modified MIP SPE. The detection limits of dopamine within buffer solutions for the MIP SPEs are determined via three independent techniques. With cyclic voltammetry this was determined to be 4.7 × 10(-6) M, whereas by using the heat-transfer method (HTM) 0.35 × 10(-6) M was obtained, and with the novel TWTA concept 0.26 × 10(-6) M is possible. This TWTA technique is measured simultaneously with HTM and has the benefits of reducing measurement time to less than 5 min and increasing effect size by nearly a factor of two. The two thermal methods are able to enhance dopamine detection by one order of magnitude compared to the electrochemical method. In previous research, it was not possible to measure neurotransmitters in complex samples with HTM, but with the improved signal-to-noise of TWTA for the first time, spiked dopamine concentrations were determined in a relevant food sample. In summary, novel concepts are presented for both the sensor functionalization side by employing screen-printing technology, and on the sensing side, the novel TWTA thermal technique is reported. The developed bio-sensing platform is cost-effective and suitable for mass-production due to the nature of screen-printing technology, which makes it very interesting for neurotransmitter detection in clinical diagnostic applications. PMID:27128891

  8. Metaphit inhibits dopamine transport and binding of ( sup 3 H)methylphenidate, a proposed marker for the dopamine transport complex

    SciTech Connect

    Schweri, M.M. ); Jacobson, A.E.; Rice, K.C. ); Lessor, R.A.

    1989-01-01

    Metaphit, an acylating derivative of phencyclidine, was shown to interact with components of the dopamine nerve terminal in rat striatal tissue. This compound, previously demonstrated to be an irreversible inhibitor at the phencyclidine receptor, was shown in these experiments to irreversibly inhibit synaptosomal ({sup 3}H)dopamine uptake. It also inhibited binding of ({sup 3}H)methylphenidate to its recognition site, which is thought to be a subunit of the dopamine transporter. Although the inhibition was due primarily to a reduction in the binding and transport capacity of the systems studied, increases in the apparent K{sub D} of ({sup 3}H)methylphenidate and the K{sub m} of ({sup 3}H)dopamine were also observed. Differences in the behavior of Metaphit and phencylidine in these dopaminergic systems compared to their effects on the NMDA receptor-linked phencyclidine receptor suggest that Metaphit may be interacting with two distinct molecular sites in the rat striatum.

  9. Thermal Stability of Dopamine Transporters.

    PubMed

    Kukk, Siim; Stepanov, Vladimir; Järv, Jaak

    2015-08-01

    The thermal stabilities of the rat and mouse dopamine transporter (DAT) proteins were studied within the temperature range of 0-37°C. The inactivation of the protein was followed by monitoring changes in radioligand-specific binding. We found that the process followed a rate equation with first-order kinetics and was characterized by having a single rate constant k inact. The activation energies (E a) that were calculated from the Arrhenius plots (ln k inact vs. 1/T) were 43 ± 5 and 45 ± 6 kJ/mol for the rat (rDAT) and mouse (mDAT) transporters, respectively, and 44 ± 7 kJ/mol for rDAT from PC-6.3 cell line. These E a values were similar to the E a values of thermal inactivation of the muscarinic receptor from rat brain cortex and to the thermal inactivation of other transmembrane proteins. However, all of these activation energy values were significantly lower than the E a values for soluble single-subunit proteins of similar size. These results therefore suggest that the thermal stability of transmembrane proteins may be governed to a significant extent by cell membrane properties and by interactions between the membrane components and the protein. In contrast, the stability of soluble proteins seems to be mostly governed by protein structure and size, which determine the sum of the stabilizing intramolecular interactions within the protein molecule. It is therefore not surprising that cell membrane properties and composition may have significant effects on the functional properties of transmembrane proteins. PMID:25812533

  10. INTERACTION OF COCAINE-, BENZTROPINE-, AND GBR12909-LIKE COMPOUNDS WITH WILDTYPE AND MUTANT HUMAN DOPAMINE TRANSPORTERS: MOLECULAR FEATURES THAT DIFFERENTIALLY DETERMINE ANTAGONIST BINDING PROPERTIES

    PubMed Central

    Schmitt, Kyle C.; Zhen, Juan; Kharkar, Prashant; Mishra, Manoj; Chen, Nianhang; Dutta, Aloke K.; Reith, Maarten E.A.

    2009-01-01

    The widely abused psychostimulant cocaine is thought to elicit its reinforcing effects primarily via inhibition of the neuronal dopamine transporter (DAT). However, not all DAT inhibitors share cocaine’s behavioral profile, despite similar or greater affinity for the DAT. This may be due to differential molecular interactions with the DAT. Our previous work using transporter mutants with altered conformational equilibrium (W84L and D313N) indicated that benztropine and GBR12909 interact with the DAT in a different manner than cocaine. Here, we expand upon these previous findings, studying a number of structurally different DAT inhibitors for their ability to inhibit [3H]CFT binding to wildtype, W84L and D313N transporters. We systematically tested structural intermediates between cocaine and benztropine, structural hybrids of benztropine and GBR12909 and a number of other structurally heterologous inhibitors. Derivatives of the stimulant desoxypipradrol (2-benzhydrylpiperidine) exhibited a cocaine-like binding profile with respect to mutation, whereas compounds possessing the diphenylmethoxy moiety of benztropine and GBR12909 were dissimilar to cocaine-like compounds. In tests with specific isomers of cocaine and tropane analogues, compounds with 3α stereochemistry tended to exhibit benztropine-like binding, whereas those with 3β stereochemistry were more cocaine-like. Our results point to the importance of specific molecular features—most notably the presence of a diphenylmethoxy moiety—in determining a compound’s binding profile. This study furthers the concept of using DAT mutants to differentiate cocaine-like inhibitors from atypical inhibitors in vitro. Further studies of the molecular features that define inhibitor-transporter interaction could lead to the development of DAT inhibitors with differential clinical utility. PMID:18786172

  11. Interaction of cocaine-, benztropine-, and GBR12909-like compounds with wild-type and mutant human dopamine transporters: molecular features that differentially determine antagonist-binding properties.

    PubMed

    Schmitt, Kyle C; Zhen, Juan; Kharkar, Prashant; Mishra, Manoj; Chen, Nianhang; Dutta, Aloke K; Reith, Maarten E A

    2008-11-01

    The widely abused psychostimulant cocaine is thought to elicit its reinforcing effects primarily via inhibition of the neuronal dopamine transporter (DAT). However, not all DAT inhibitors share cocaine's behavioral profile, despite similar or greater affinity for the DAT. This may be due to differential molecular interactions with the DAT. Our previous work using transporter mutants with altered conformational equilibrium (W84L and D313N) indicated that benztropine and GBR12909 interact with the DAT in a different manner than cocaine. Here, we expand upon these previous findings, studying a number of structurally different DAT inhibitors for their ability to inhibit [(3)H]CFT binding to wild-type, W84L and D313N transporters. We systematically tested structural intermediates between cocaine and benztropine, structural hybrids of benztropine and GBR12909 and a number of other structurally heterologous inhibitors. Derivatives of the stimulant desoxypipradrol (2-benzhydrylpiperidine) exhibited a cocaine-like binding profile with respect to mutation, whereas compounds possessing the diphenylmethoxy moiety of benztropine and GBR12909 were dissimilar to cocaine-like compounds. In tests with specific isomers of cocaine and tropane analogues, compounds with 3alpha stereochemistry tended to exhibit benztropine-like binding, whereas those with 3beta stereochemistry were more cocaine-like. Our results point to the importance of specific molecular features--most notably the presence of a diphenylmethoxy moiety--in determining a compound's binding profile. This study furthers the concept of using DAT mutants to differentiate cocaine-like inhibitors from atypical inhibitors in vitro. Further studies of the molecular features that define inhibitor-transporter interaction could lead to the development of DAT inhibitors with differential clinical utility. PMID:18786172

  12. Psychostimulants affect dopamine transmission through both dopamine transporter-dependent and independent mechanisms.

    PubMed

    dela Peña, Ike; Gevorkiana, Ruzanna; Shi, Wei-Xing

    2015-10-01

    The precise mechanisms by which cocaine and amphetamine-like psychostimulants exert their reinforcing effects are not yet fully defined. It is widely believed, however, that these drugs produce their effects by enhancing dopamine neurotransmission in the brain, especially in limbic areas such as the nucleus accumbens, by inducing dopamine transporter-mediated reverse transport and/or blocking dopamine reuptake though the dopamine transporter. Here, we present the evidence that aside from dopamine transporter, non-dopamine transporter-mediated mechanisms also participate in psychostimulant-induced dopamine release and contribute to the behavioral effects of these drugs, such as locomotor activation and reward. Accordingly, psychostimulants could increase norepinephrine release in the prefrontal cortex, the latter then alters the firing pattern of dopamine neurons resulting in changes in action potential-dependent dopamine release. These alterations would further affect the temporal pattern of dopamine release in the nucleus accumbens, thereby modifying information processing in that area. Hence, a synaptic input to a nucleus accumbens neuron may be enhanced or inhibited by dopamine depending on its temporal relationship to dopamine release. Specific temporal patterns of dopamine release may also be required for certain forms of synaptic plasticity in the nucleus accumbens. Together, these effects induced by psychostimulants, mediated through a non-dopamine transporter-mediated mechanism involving norepinephrine and the prefrontal cortex, may also contribute importantly to the reinforcing properties of these drugs. PMID:26209364

  13. X-ray structure of dopamine transporter elucidates antidepressant mechanism.

    PubMed

    Penmatsa, Aravind; Wang, Kevin H; Gouaux, Eric

    2013-11-01

    Antidepressants targeting Na(+)/Cl(-)-coupled neurotransmitter uptake define a key therapeutic strategy to treat clinical depression and neuropathic pain. However, identifying the molecular interactions that underlie the pharmacological activity of these transport inhibitors, and thus the mechanism by which the inhibitors lead to increased synaptic neurotransmitter levels, has proven elusive. Here we present the crystal structure of the Drosophila melanogaster dopamine transporter at 3.0 Å resolution bound to the tricyclic antidepressant nortriptyline. The transporter is locked in an outward-open conformation with nortriptyline wedged between transmembrane helices 1, 3, 6 and 8, blocking the transporter from binding substrate and from isomerizing to an inward-facing conformation. Although the overall structure of the dopamine transporter is similar to that of its prokaryotic relative LeuT, there are multiple distinctions, including a kink in transmembrane helix 12 halfway across the membrane bilayer, a latch-like carboxy-terminal helix that caps the cytoplasmic gate, and a cholesterol molecule wedged within a groove formed by transmembrane helices 1a, 5 and 7. Taken together, the dopamine transporter structure reveals the molecular basis for antidepressant action on sodium-coupled neurotransmitter symporters and elucidates critical elements of eukaryotic transporter structure and modulation by lipids, thus expanding our understanding of the mechanism and regulation of neurotransmitter uptake at chemical synapses. PMID:24037379

  14. Classic Studies on the Interaction of Cocaine and the Dopamine Transporter.

    PubMed

    Verma, Vivek

    2015-12-31

    The dopamine transporter is responsible for recycling dopamine after release. Inhibitors of the dopamine transporter, such as cocaine, will stop the reuptake of dopamine and allow it to stay extracellularly, causing prominent changes at the molecular, cellular, and behavioral levels. There is much left to be known about the mechanism and site(s) of binding, as well as the effect that cocaine administration does to dopamine transporter-cocaine binding sites and gene expression which also plays a strong role in cocaine abusers and their behavioral characteristics. Thus, if more light is shed on the dopamine transporter-cocaine interaction, treatments for addiction and even other diseases of the dopaminergic system may not be too far ahead. As today's ongoing research expands on the shoulders of classic research done in the 1990s and 2000s, the foundation of core research done in that time period will be reviewed, which forms the basis of today's work and tomorrow's therapies. PMID:26598579

  15. Neurotransmitter and psychostimulant recognition by the dopamine transporter

    PubMed Central

    Wang, Kevin H.; Penmatsa, Aravind; Gouaux, Eric

    2015-01-01

    Na+/Cl−-coupled biogenic amine transporters are the primary targets of therapeutic and abused drugs, ranging from antidepressants to the psychostimulants cocaine and amphetamines, and to their cognate substrates. Here we determine x-ray crystal structures of the Drosophila melanogaster dopamine transporter (dDAT) bound to its substrate dopamine (DA), a substrate analogue 3,4-dichlorophenethylamine, the psychostimulants D-amphetamine, methamphetamine, or to cocaine and cocaine analogues. All ligands bind to the central binding site, located approximately halfway across the membrane bilayer, in close proximity to bound sodium and chloride ions. The central binding site recognizes three chemically distinct classes of ligands via conformational changes that accommodate varying sizes and shapes, thus illustrating molecular principles that distinguish substrates from inhibitors in biogenic amine transporters. PMID:25970245

  16. Cloning of the cocaine-sensitive bovine dopamine transporter

    SciTech Connect

    Usdin, T.B.; Chen, C.; Brownstein, M.J.; Hoffman, B.J. ); Mezey, E. )

    1991-12-15

    A cDNA encoding the dopamine transporter from bovine brain substantia nigra was identified on the basis of its structural homology to other, recently cloned, neurotransmitter transporters. The sequence of the 693-amino acid protein is quite similar to those of the rat {gamma}-aminobutyric acid, human norepinephrine, and rat serotonin transporters. Dopamine transporter mRNA was detected by in situ hybridization in the substantia nigra but not in the locus coeruleus, raphe, caudate, or other brain areas. ({sup 3}H)Dopamine accumulation in tissue culture cells transfected with the cDNA was inhibited by amphetamine, cocaine, and specific inhibitors of dopamine transports, including GBR12909.

  17. The dopamine transporter: role in neurotoxicity and human disease

    SciTech Connect

    Bannon, Michael J. . E-mail: mbannon@med.wayne.edu

    2005-05-01

    The dopamine transporter (DAT) is a plasma membrane transport protein expressed exclusively within a small subset of CNS neurons. It plays a crucial role in controlling dopamine-mediated neurotransmission and a number of associated behaviors. This review focuses on recent data elucidating the role of the dopamine transporter in neurotoxicity and a number of CNS disorders, including Parkinson disease, drug abuse, and attention deficit hyperactivity disorder (ADHD)

  18. Substrates dissociate dopamine transporter oligomers

    PubMed Central

    Chen, Nianhang; Reith, Maarten E. A.

    2016-01-01

    Substrate-induced endocytic trafficking of DAT has been observed, but little is known about the regulation of DAT oligomerization by substrate. The present study investigates the effect on substrates on DAT oligomerization and explores a potential link with the presence of DAT at the cell surface in HEK-293 cells transiently or stably expressing N-terminal tagged DAT constructs. DA (100 μM) or amphetamine (2–10 μM) reduced Myc-DAT coimmunoprecipitated along with Flag-DAT (oligomeric DAT) in tandem with a reduction in surface DAT determined by biotinylation. DA (10–1000 μM) and amphetamine (0.2–200 μM) reduced DAT oligomerization as assessed by cross-linking with copper sulfate phenanthroline (CuP) or Cu2+. Inhibition of endocytosis by 10 μM phenylarsine oxide (PAO) or 450 mM sucrose counteracted the effect of 10 μM DA or 2 μM amphetamine in reducing DAT cross-linking. In addition to overall similarities between the results with the two cross-linking agents and between the results with the two different endocytosis inhibitors, some differences were noted as well, likely related to the efficiency of the cross-linking process and the sulfhydryl-reactive properties of PAO, respectively. The present results are the first to indicate regulation of oligomerization of an SLC6 transporter, the DAT, by substrates that act at DAT. In addition, the present study opens up the possibility of an important linkage between between oligomerization of DAT and endocytic or other modulatory mechanisms impacting surface DAT. PMID:18088380

  19. Ethics of Preclinical Dopamine Transporter Imaging.

    PubMed

    Cochrane, Thomas I

    2016-08-01

    While dopamine transporter single-photon emission computed tomography (DAT-SPECT) imaging is sensitive and specific when performed in patients with signs or symptoms of parkinsonism, its predictive value is uncertain in healthy subjects, even with patients who have first-degree relatives affected by Parkinson disease. In deciding whether to honor a patient's request for a DAT-SPECT, neurologists must balance a patient's autonomy rights with beneficence and nonmaleficence and also consider the distributive justice implications of ordering the test. Generally speaking, the benefits of a DAT-SPECT will be too small to justify its use in an asymptomatic patient concerned about developing Parkinson disease. PMID:27495208

  20. Ceramide-induced alterations in dopamine transporter function.

    PubMed

    Riddle, Evan L; Rau, Kristi S; Topham, Matthew K; Hanson, Glen R; Fleckenstein, Annette E

    2003-01-01

    The purpose of this study was to determine the effects of ceramide on dopamine and serotonin (5-HT, 5-hydroxytryptamine) transporters. Exposure of rat striatal synaptosomes to C2-ceramide caused a reversible, concentration-dependent decrease in plasmalemmal dopamine uptake. In contrast, ceramide exposure increased striatal 5-HT synaptosomal uptake. This increase did not appear to be due to an increased uptake by the 5-HT transporter. Rather, the increase appeared to result from an increase in 5-HT transport through the dopamine transporter, an assertion evidenced by findings that this increase: (1) does not occur in hippocampal synaptosomes (i.e., a preparation largely devoid of dopamine transporters), (2) occurs in striatal synaptosomes prepared from para-chloroamphetamine-treated rats (i.e., a preparation lacking 5-HT transporters), (3) is attenuated by pretreatment with methylphenidate (i.e., a relatively selective dopamine reuptake inhibitor) and (4) is inhibited by exposure to exogenous dopamine (i.e., which presumably competes for uptake with 5-HT). Taken together, these results reveal that ceramide is a novel modulator of monoamine transporter function, and may alter the affinity of dopamine transporters for its primary substrate. PMID:12498904

  1. Importance of cholesterol in dopamine transporter function

    PubMed Central

    Jones, Kymry T.; Zhen, Juan; Reith, Maarten E.A.

    2012-01-01

    The conformation and function of the dopamine transporter (DAT) can be affected by manipulating membrane cholesterol, yet there is no agreement as to the impact of cholesterol on the activity of lipid-raft localized DATs compared to non-raft DATs. Given the paucity of information regarding the impact of cholesterol on substrate efflux by the DAT, this study explores its influence on the kinetics of DAT-mediated DA efflux induced by dextroamphetamine, as measured by rotating disk electrode voltammetry (RDEV). Treatment with methyl-β-cyclodextrin (mβCD), which effectively depletes total membrane cholesterol- uniformly affecting cholesterol-DAT interactions in both raft and non-raft membrane domains- reduced both DA uptake and efflux rate. In contrast, disruption of raft localized DAT by cholesterol chelation with nystatin had no effect, arguing against a vital role for raft-localized DAT in substrate uptake or efflux. Supra-normal repletion of cholesterol depleted cells with the analogue desmosterol, a non-raft promoting sterol, was as effective as cholesterol itself in restoring transport rates. Further studies with Zn2+ and the conformationally-biased W84L DAT mutant supported the idea that cholesterol is important for maintaining the outward-facing DAT with normal rates of conformational interconversions. Collectively, these results point to a role for direct cholesterol-DAT interactions in regulating DAT function. PMID:22957537

  2. Dopamine Transporters, D2 Receptors, and Dopamine Release in Generalized Social Anxiety Disorder

    PubMed Central

    Schneier, Franklin R.; Abi-Dargham, Anissa; Martinez, Diana; Slifstein, Mark; Hwang, Dah-Ren; Liebowitz, Michael R.; Laruelle, Marc

    2009-01-01

    Background Dopamine D2 receptor and dopamine transporter availability in the striatum have each been reported abnormal in generalized social anxiety disorder (GSAD) in studies using single photon computerized tomography (SPECT). D2 receptors and dopamine transporters have not previously been studied within the same GSAD subjects, however, and prior GSAD studies have not assessed dopamine release or subdivided striatum into functional subregions. Methods Unmedicated adults with GSAD (N=17) and matched healthy comparison subjects (HC, N=13) participated in this study. Of these, 15 GSAD and 13 HC subjects completed baseline assessment of D2 receptor availability using positron emission tomography (PET) with the radiotracer [11C] raclopride. Twelve GSAD and 13 HC subjects completed a repeat scan after intravenous administration of D-amphetamine, to study dopamine release. Twelve of the GSAD subjects and 10 of the HC subjects also completed SPECT with the radiotracer [123I] methyl 3ß-(4-iodophenyl) tropane-2ß-carboxylate ([123I] ß-CIT) to assess dopamine transporter availability. Results GSAD and HC groups did not differ significantly in striatal dopamine transporter availability, overall striatal or striatal subregion D2 receptor availability at baseline, or change in D2 receptor availability after D-amphetamine. Receptor availability and change after D-amphetamine were not significantly associated with severity of social anxiety or trait detachment. Conclusions These findings do not replicate previous findings of altered striatal dopamine transporter and D2 receptor availability in GSAD subjects assessed with SPECT. The differences from results of prior studies may be due to differences in imaging methods or characteristics of samples. PMID:19180583

  3. Classic Studies on the Interaction of Cocaine and the Dopamine Transporter

    PubMed Central

    Verma, Vivek

    2015-01-01

    The dopamine transporter is responsible for recycling dopamine after release. Inhibitors of the dopamine transporter, such as cocaine, will stop the reuptake of dopamine and allow it to stay extracellularly, causing prominent changes at the molecular, cellular, and behavioral levels. There is much left to be known about the mechanism and site(s) of binding, as well as the effect that cocaine administration does to dopamine transporter-cocaine binding sites and gene expression which also plays a strong role in cocaine abusers and their behavioral characteristics. Thus, if more light is shed on the dopamine transporter-cocaine interaction, treatments for addiction and even other diseases of the dopaminergic system may not be too far ahead. As today’s ongoing research expands on the shoulders of classic research done in the 1990s and 2000s, the foundation of core research done in that time period will be reviewed, which forms the basis of today’s work and tomorrow’s therapies. PMID:26598579

  4. Altered Neurocircuitry in the Dopamine Transporter Knockout Mouse Brain

    PubMed Central

    Zhang, Xiaowei; Bearer, Elaine L.; Boulat, Benoit; Hall, F. Scott; Uhl, George R.; Jacobs, Russell E.

    2010-01-01

    The plasma membrane transporters for the monoamine neurotransmitters dopamine, serotonin, and norepinephrine modulate the dynamics of these monoamine neurotransmitters. Thus, activity of these transporters has significant consequences for monoamine activity throughout the brain and for a number of neurological and psychiatric disorders. Gene knockout (KO) mice that reduce or eliminate expression of each of these monoamine transporters have provided a wealth of new information about the function of these proteins at molecular, physiological and behavioral levels. In the present work we use the unique properties of magnetic resonance imaging (MRI) to probe the effects of altered dopaminergic dynamics on meso-scale neuronal circuitry and overall brain morphology, since changes at these levels of organization might help to account for some of the extensive pharmacological and behavioral differences observed in dopamine transporter (DAT) KO mice. Despite the smaller size of these animals, voxel-wise statistical comparison of high resolution structural MR images indicated little morphological change as a consequence of DAT KO. Likewise, proton magnetic resonance spectra recorded in the striatum indicated no significant changes in detectable metabolite concentrations between DAT KO and wild-type (WT) mice. In contrast, alterations in the circuitry from the prefrontal cortex to the mesocortical limbic system, an important brain component intimately tied to function of mesolimbic/mesocortical dopamine reward pathways, were revealed by manganese-enhanced MRI (MEMRI). Analysis of co-registered MEMRI images taken over the 26 hours after introduction of Mn2+ into the prefrontal cortex indicated that DAT KO mice have a truncated Mn2+ distribution within this circuitry with little accumulation beyond the thalamus or contralateral to the injection site. By contrast, WT littermates exhibit Mn2+ transport into more posterior midbrain nuclei and contralateral mesolimbic structures at

  5. Dopamine receptor genes: new tools for molecular psychiatry.

    PubMed Central

    Niznik, H B; Van Tol, H H

    1992-01-01

    For over a decade it has been generally assumed that all the pharmacological and biochemical actions of dopamine within the central nervous system and periphery were mediated by two distinct dopamine receptors. These receptors, termed D1 and D2, were defined as those coupled to the stimulation or inhibition of adenylate cyclase, respectively, and by their selectivity and avidity for various drugs and compounds. The concept that two dopamine receptors were sufficient to account for all the effects mediated by dopamine was an oversimplification. Recent molecular biological studies have identified five distinct genes which encode at least eight functional dopamine receptors. The members of the expanded dopamine receptor family, however, can still be codifed by way of the original D1 and D2 receptor dichotomy. These include two genes encoding dopamine D1-like receptors (D1 [D1A]/D5 [D1B]) and three genes encoding D2-like receptors (D2/D3/D4). We review here our recent work on the cloning and characterization of some of the members of the dopamine receptor gene family (D1, D2, D4, D5), their relationship to neuropsychiatric disorders and their potential role in antipsychotic drug action. Images Fig. 1 PMID:1450188

  6. Differential regional effects of methamphetamine on dopamine transport.

    PubMed

    Chu, Pei-Wen; Seferian, Kristi S; Birdsall, Elisabeth; Truong, Jannine G; Riordan, James A; Metcalf, Cameron S; Hanson, Glen R; Fleckenstein, Annette E

    2008-08-20

    Multiple high-dose methamphetamine administrations cause long-lasting (>1 week) deficits in striatal dopaminergic neuronal function. This stimulant likewise causes rapid (within 1 h) and persistent (at least 48 h) decreases in activities of striatal: 1) dopamine transporters, as assessed in synaptosomes; and 2) vesicular monoamine transporter -2 (VMAT-2), as assessed in a non-membrane-associated (referred to herein as cytoplasmic) vesicular subcellular fraction. Importantly, not all brain areas are vulnerable to methamphetamine-induced long-lasting deficits. Similarly, the present study indicates that methamphetamine exerts differential acute effects on monoaminergic transporters according to brain region. In particular, results revealed that in the nucleus accumbens, methamphetamine rapidly, but reversibly (within 24 h), decreased plasmalemmal dopamine transporter function, without effect on plasmalemmal dopamine transporter immunoreactivity. Methamphetamine also rapidly and reversibly (within 48 h) decreased cytoplasmic VMAT-2 function in this region, with relatively little effect on cytoplasmic VMAT-2 immunoreactivity. In contrast, methamphetamine did not alter either dopamine transporter or VMAT-2 activity in the hypothalamus. Noteworthy, the nucleus accumbens and hypothalamus did not display the persistent long-lasting striatal dopamine depletions caused by the stimulant. Taken together, these data suggest that deficits in plasmalemmal and vesicular monoamine transporter activity lasting greater than 24-48 h may be linked to the long-lasting dopaminergic deficits caused by methamphetamine and appear to be region specific. PMID:18599036

  7. Recovery of dopamine transporters with methamphetamine detoxification is not linked to changes in dopamine release.

    PubMed

    Volkow, Nora D; Wang, Gene-Jack; Smith, Lisa; Fowler, Joanna S; Telang, Frank; Logan, Jean; Tomasi, Dardo

    2015-11-01

    Methamphetamine's widepread abuse and concerns that it might increase Parkinson's disease led us to assess if the reported loss of dopamine transporters (DAT) in methamphetamine abusers (MA) reflected damage to dopamine neurons. Using PET with [(11)C]cocaine to measure DAT, and with [(11)C]raclopride to measure dopamine release (assessed as changes in specific binding of [(11)C]raclopride between placebo and methylphenidate), which was used as a marker of dopamine neuronal function, we show that MA (n=16), tested during early detoxification, had lower DAT (20-30%) but overall normal DA release in striatum (except for a small decrease in left putamen), when compared to controls (n=15). In controls, DAT were positively correlated with DA release (higher DAT associated with larger DA increases), consistent with DAT serving as markers of DA terminals. In contrast, MA showed a trend for a negative correlation (p=0.07) (higher DAT associated with lower DA increases), consistent with reduced DA re-uptake following DAT downregulation. MA who remained abstinent nine-months later (n=9) showed significant increases in DAT (20%) but methylphenidate-induced dopamine increases did not change. In contrast, in controls, DAT did not change when retested 9 months later but methylphenidate-induced dopamine increases in ventral striatum were reduced (p=0.05). Baseline D2/D3 receptors in caudate were lower in MA than in controls and did not change with detoxification, nor did they change in the controls upon retest. The loss of DAT in the MA, which was not associated with a concomitant reduction in dopamine release as would have been expected if DAT loss reflected DA terminal degneration; as well as the recovery of DAT after protracted detoxification, which was not associated with increased dopamine release as would have been expected if DAT increases reflected terminal regeneration, indicate that the loss of DAT in these MA does not reflect degeneration of dopamine terminals. PMID

  8. Dopamine transport sites selectively labeled by a novel photoaffinity probe: 125I-DEEP

    SciTech Connect

    Grigoriadis, D.E.; Wilson, A.A.; Lew, R.; Sharkey, J.S.; Kuhar, M.J. )

    1989-08-01

    The dopamine transporter was labeled using a photosensitive compound related to GBR-12909, {sup 125}I-1-(2-(diphenylmethoxy)ethyl)-4-(2- (4-azido-3-iodophenyl)ethyl)piperazine ({sup 125}I-DEEP). {sup 125}I-DEEP bound reversibly and with high affinity to the dopamine transport protein in the absence of light and could be covalently attached to the protein following exposure to UV light. In rat striatal homogenates, {sup 125}I-DEEP was found to incorporate covalently into a protein with apparent molecular weight of 58,000 Da. The properties of this binding protein were characteristic of the dopamine transporter since covalent attachment could be inhibited by dopamine-uptake blockers with the proper pharmacological rank order of potencies. Covalent binding was also inhibited in a stereospecific manner by (+) and (-) cocaine, as well as other cocaine analogs. The protein was not found in the cerebellum. The dopamine transporter appears to exist in a glycosylated form since photoaffinity-labeled transport sites could adsorb to wheat germ-agglutinin and could be specifically eluted from the column by beta-N-acetylglucosamine.

  9. Urinary Dopamine as a Potential Index of the Transport Activity of Multidrug and Toxin Extrusion in the Kidney

    PubMed Central

    Kajiwara, Moto; Ban, Tsuyoshi; Matsubara, Kazuo; Nakanishi, Yoichi; Masuda, Satohiro

    2016-01-01

    Dopamine is a cationic natriuretic catecholamine synthesized in proximal tubular cells (PTCs) of the kidney before secretion into the lumen, a key site of its action. However, the molecular mechanisms underlying dopamine secretion into the lumen remain unclear. Multidrug and toxin extrusion (MATE) is a H+/organic cation antiporter that is highly expressed in the brush border membrane of PTCs and mediates the efflux of organic cations, including metformin and cisplatin, from the epithelial cells into the urine. Therefore, we hypothesized that MATE mediates dopamine secretion, a cationic catecholamine, into the tubule lumen, thereby regulating natriuresis. Here, we show that [3H]dopamine uptake in human (h) MATE1-, hMATE-2K- and mouse (m) MATE-expressing cells exhibited saturable kinetics. Fluid retention and decreased urinary excretion of dopamine and Na+ were observed in Mate1-knockout mice compared to that in wild-type mice. Imatinib, a MATE inhibitor, inhibited [3H]dopamine uptake by hMATE1-, hMATE2-K- and mMATE1-expressing cells in a concentration-dependent manner. At clinically-relevant concentrations, imatinib inhibited [3H]dopamine uptake by hMATE1- and hMATE2-K-expressing cells. The urinary excretion of dopamine and Na+ decreased and fluid retention occurred in imatinib-treated mice. In conclusion, MATE transporters secrete renally-synthesized dopamine, and therefore, urinary dopamine has the potential to be an index of the MATE transporter activity. PMID:27483254

  10. Oxygen radicals diminish dopamine transporter function in rat striatum.

    PubMed

    Fleckenstein, A E; Metzger, R R; Beyeler, M L; Gibb, J W; Hanson, G R

    1997-09-01

    Incubation of striatal synaptosomes with the oxygen radical generating enzyme, xanthine oxidase, decreased [3H]dopamine uptake: an effect attributable to a decreased Vmax. Concurrent incubation with the superoxide radical scavenger, superoxide dismutase, abolished the xanthine oxidase-induced decrease. These results indicate that, like methamphetamine administration in vivo, reactive oxygen species diminish dopamine transporter function in vitro. The significance of these findings to mechanisms responsible for effects of methamphetamine is discussed. PMID:9346337

  11. Affinities of methylphenidate derivatives for dopamine, norepinephrine and serotonin transporters.

    PubMed

    Gatley, S J; Pan, D; Chen, R; Chaturvedi, G; Ding, Y S

    1996-01-01

    We have synthesized several derivative of dl-threo-methylphenidate (Ritalin) bearing substituents on the phenyl ring. IC50 values for binding these compounds to rat brain monoamine transporters were assessed using [3H]WIN 35,428 (striatal membranes, dopamine transporters, DAT), [3H]nisoxetine (frontal cortex membranes, norepinephrine transporters, NET) and [3H]paroxetine (brain stem membranes, 5HT transporters, 5HTT). Affinities (1/Ki) decreased in the order: DAT > NET > 5HTT. Substitution at the para position of dl-threo-methylphenidate generally led to retained or increased affinity for the dopamine transporter (bromo > iodo > methoxy > hydroxy). Substitution at the meta position also increased affinity for the DAT (m-bromo > methylphenidate; m-iodo-p-hydroxy > p-hydroxy). Substitution at the ortho position with bromine considerably decreased affinity. Similar IC50 values for binding of o-bromomethylphenidate to the dopamine transporter were measured at 0, 22 and 37 degrees. N-Methylation of the piperidine ring of methylphenidate also considerably reduced affinity. The dl-erythro isomer of o-bromomethylphenidate did not bind to the DAT (IC50 > 50,000 nM). Affinities at the dopamine and norepinephrine transporters for substituted methylphenidate derivatives were well correlated (r2=0.90). Abilities of several methylphenidate derivatives to inhibit [3H]dopamine uptake in striatal synaptosomes corresponded well with inhibition of [3H]WIN 35, 428 binding. None of the compounds examined exhibited significant affinity to dopamine D1 or D2 receptors (IC50 > 500 or 5,000 nM, respectively), as assessed by inhibition of binding of [3H]SCH 23390 or [123I]epidepride, respectively, to striatal membranes. PMID:8786705

  12. Atypical Dopamine Uptake Inhibitors that Provide Clues About Cocaine's Mechanism at the Dopamine Transporter

    NASA Astrophysics Data System (ADS)

    Hauck Newman, Amy; Katz, Jonathan L.

    The dopamine transporter (DAT) has been a primary target for cocaine abuse/addiction medication discovery. However predicted addiction liability and limited clinical evaluation has provided a formidable challenge for development of these agents for human use. The unique and atypical pharmacological profile of the benztropine (BZT) class of dopamine uptake inhibitors, in preclinical models of cocaine effects and abuse, has encouraged further development of these agents. Moreover, in vivo studies have challenged the original DAT hypothesis and demonstrated that DAT occupancy and subsequent increases in dopamine produced by BZT analogues are significantly delayed and long lasting, as compared to cocaine. These important and distinctive elements are critical to the lack of abuse liability among BZT analogues, and improve their potential for development as treatments for cocaine abuse and possibly other neuropsychiatric disorders.

  13. A Physical Interaction between the Dopamine Transporter and DJ-1 Facilitates Increased Dopamine Reuptake

    PubMed Central

    Luk, Beryl; Mohammed, Mohinuddin; Liu, Fang; Lee, Frank J. S.

    2015-01-01

    The regulation of the dopamine transporter (DAT) impacts extracellular dopamine levels after release from dopaminergic neurons. Furthermore, a variety of protein partners have been identified that can interact with and modulate DAT function. In this study we show that DJ-1 can potentially modulate DAT function. Co-expression of DAT and DJ-1 in HEK-293T cells leads to an increase in [3H] dopamine uptake that does not appear to be mediated by increased total DAT expression but rather through an increase in DAT cell surface localization. In addition, through a series of GST affinity purifications and co-immunoprecipitations, we provide evidence that the DAT can be found in a complex with DJ-1, which involve distinct regions within both DAT and DJ-1. Using in vitro binding experiments we also show that this complex can be formed in part by a direct interaction between DAT and DJ-1. Co-expression of a mini-gene that can disrupt the DAT/DJ-1 complex appears to block the increase in [3H] dopamine uptake by DJ-1. Mutations in DJ-1 have been linked to familial forms of Parkinson’s disease, yet the normal physiological function of DJ-1 remains unclear. Our study suggests that DJ-1 may also play a role in regulating dopamine levels by modifying DAT activity. PMID:26305376

  14. Recovery of dopamine transporters with methamphetamine detoxification is not linked to changes in dopamine release

    SciTech Connect

    Volkow, Nora D.; Wang, Gene-Jack; Smith, Lisa; Fowler, Joanna S.; Telang, Frank; Logan, Jean; Tomasi, Dardo

    2015-07-21

    Metamphetamine’s widepread abuse and concerns that it may increase Parkinson’s disease led us to assess if the reported loss of dopamine transporters (DAT) in methamphetamine abusers (MA) reflected damage to dopamine neurons. Using PET with [11C]cocaine to measure DAT, and with [11C]raclopride to measure dopamine release (assessed as changes in specific binding of [11C]raclopride between placebo and methylphenidate), which was used as marker of dopamine neuronal function, we show that MA (n=16), tested during early detoxification, had lower DAT (20-30%) but overall normal DA release in striatum (except for a small decrease in left putamen), when compared to controls (n=15). In controls, DAT were positively correlated with DA release (higher DAT associated with larger DA increases), consistent with DAT serving as markers of DA terminals. In contrast, MA showed a trend for a negative correlation (p=0.07) (higher DAT associated with lower DA increases), consistent with reduced DA re-uptake following DAT downregulation. MA who remained abstinent nine-months later (n=9) showed significant increases in DAT (20%) but methylphenidate-induced dopamine increases did not change. In contrast, in controls, DAT did not change when retested 9 months later but methylphenidate-induced dopamine increases in ventral striatum were reduced (p=0.05). Baseline D2/D3 receptors in caudate were lower in MA than in controls and did not change with detoxification, nor did they change in the controls upon retest. The loss of DAT in the MA, which was not associated with a concomitant reduction in dopamine release as would have been expected if DAT loss reflected DA terminal degneration; as well as the recovery of DAT after protracted detoxification, which was not associated with increased dopamine release as would have been expected if DAT increases reflected terminal regeneration, indicate that the loss of DAT in these MA does not reflect degeneration of

  15. Recovery of dopamine transporters with methamphetamine detoxification is not linked to changes in dopamine release

    DOE PAGESBeta

    Volkow, Nora D.; Wang, Gene-Jack; Smith, Lisa; Fowler, Joanna S.; Telang, Frank; Logan, Jean; Tomasi, Dardo

    2015-07-21

    Metamphetamine’s widepread abuse and concerns that it may increase Parkinson’s disease led us to assess if the reported loss of dopamine transporters (DAT) in methamphetamine abusers (MA) reflected damage to dopamine neurons. Using PET with [11C]cocaine to measure DAT, and with [11C]raclopride to measure dopamine release (assessed as changes in specific binding of [11C]raclopride between placebo and methylphenidate), which was used as marker of dopamine neuronal function, we show that MA (n=16), tested during early detoxification, had lower DAT (20-30%) but overall normal DA release in striatum (except for a small decrease in left putamen), when compared to controls (n=15).more » In controls, DAT were positively correlated with DA release (higher DAT associated with larger DA increases), consistent with DAT serving as markers of DA terminals. In contrast, MA showed a trend for a negative correlation (p=0.07) (higher DAT associated with lower DA increases), consistent with reduced DA re-uptake following DAT downregulation. MA who remained abstinent nine-months later (n=9) showed significant increases in DAT (20%) but methylphenidate-induced dopamine increases did not change. In contrast, in controls, DAT did not change when retested 9 months later but methylphenidate-induced dopamine increases in ventral striatum were reduced (p=0.05). Baseline D2/D3 receptors in caudate were lower in MA than in controls and did not change with detoxification, nor did they change in the controls upon retest. The loss of DAT in the MA, which was not associated with a concomitant reduction in dopamine release as would have been expected if DAT loss reflected DA terminal degneration; as well as the recovery of DAT after protracted detoxification, which was not associated with increased dopamine release as would have been expected if DAT increases reflected terminal regeneration, indicate that the loss of DAT in these MA does not reflect degeneration of dopamine terminals.« less

  16. Dopamine Transporter Genotype Predicts Attentional Asymmetry in Healthy Adults

    ERIC Educational Resources Information Center

    Newman, Daniel P.; O'Connell, Redmond G.; Nathan, Pradeep J.; Bellgrove, Mark A.

    2012-01-01

    A number of recent studies suggest that DNA variation in the dopamine transporter gene (DAT1) influences spatial attention asymmetry in clinical populations such as ADHD, but confirmation in non-clinical samples is required. Since non-spatial factors such as attentional load have been shown to influence spatial biases in clinical conditions, here…

  17. Presence and Function of Dopamine Transporter (DAT) in Stallion Sperm: Dopamine Modulates Sperm Motility and Acrosomal Integrity

    PubMed Central

    Covarrubias, Alejandra A.; Rodríguez-Gil, Joan Enric; Ramírez-Reveco, Alfredo; Concha, Ilona I.

    2014-01-01

    Dopamine is a catecholamine with multiple physiological functions, playing a key role in nervous system; however its participation in reproductive processes and sperm physiology is controversial. High dopamine concentrations have been reported in different portions of the feminine and masculine reproductive tract, although the role fulfilled by this catecholamine in reproductive physiology is as yet unknown. We have previously shown that dopamine type 2 receptor is functional in boar sperm, suggesting that dopamine acts as a physiological modulator of sperm viability, capacitation and motility. In the present study, using immunodetection methods, we revealed the presence of several proteins important for the dopamine uptake and signalling in mammalian sperm, specifically monoamine transporters as dopamine (DAT), serotonin (SERT) and norepinephrine (NET) transporters in equine sperm. We also demonstrated for the first time in equine sperm a functional dopamine transporter using 4-[4-(Dimethylamino)styryl]-N-methylpyridinium iodide (ASP+), as substrate. In addition, we also showed that dopamine (1 mM) treatment in vitro, does not affect sperm viability but decreases total and progressive sperm motility. This effect is reversed by blocking the dopamine transporter with the selective inhibitor vanoxerine (GBR12909) and non-selective inhibitors of dopamine reuptake such as nomifensine and bupropion. The effect of dopamine in sperm physiology was evaluated and we demonstrated that acrosome integrity and thyrosine phosphorylation in equine sperm is significantly reduced at high concentrations of this catecholamine. In summary, our results revealed the presence of monoamine transporter DAT, NET and SERT in equine sperm, and that the dopamine uptake by DAT can regulate sperm function, specifically acrosomal integrity and sperm motility. PMID:25402186

  18. Association of attention-deficit disorder and the dopamine transporter gene

    SciTech Connect

    Cook, E.H. Jr.; Stein, M.A.; Krasowski, M.D.; Cox, N.J.; Olkon, D.M.; Kieffer, J.E.; Leventhal, B.L.

    1995-04-01

    Attention-deficit hyperactivity disorder (ADHD) has been shown to be familial and heritable, in previous studies. As with most psychiatric disorders, examination of pedigrees has not revealed a consistent Mendelian mode of transmission. The response of ADHD patients to medications that inhibit the dopamine transporter, including methylphenidate, amphetamine, pemoline, and bupropion, led us to consider the dopamine transporter as a primary candidate gene for ADHD. To avoid effects of population stratification and to avoid the problem of classification of relatives with other psychiatric disorders as affected or unaffected, we used the haplotype-based haplotype relative risk (HHRR) method to test for association between a VNTR polymorphism at the dopamine transporter locus (DAT1) and DSM-III-R-diagnosed ADHD (N = 49) and undifferentiated attention-deficit disorder (UADD) (N = 8) in trios composed of father, mother, and affected offspring. HHRR analysis revealed significant association between ADHS/UADD and the 480-bp DAT1 allele (X{sup 2} 7.51, 1 df, P = .006). When cases of UADD were dropped from the analysis, similar results were found (X{sup 2} 7.29, 1 df, P = .007). If these findings are replicated, molecular analysis of the dopamine transporter gene may identify mutations that increase susceptibility to ADHD/UADD. Biochemical analysis of such mutations may lead to development of more effective therapeutic interventions. 36 refs., 4 tabs.

  19. Tuberoinfundibular transport of intrahypothalamic-administered dopamine in normo- and hypertensive rats

    SciTech Connect

    Sim, M.K.

    1988-01-01

    The dopamine transport system in the tuberoinfundibular tract of the spontaneously hypertensive (SHR), Wistar Kyoto (WKY) and Sprague-Dawley (SD) rats was investigated. The results show that the rate of dopamine transport in this tract is strain-specific. SD rats transported twice as much dopamine (in 30 minutes) as WKY and SHR. The dopamine transport system in the SHR, being at par with that of the WKY, remained intact. These findings suggest that hypertension and the alleged reduced central dopaminergic activity in the SHR is not related to the transport of dopamine in the tuberoinfundibular tract.

  20. Pyrethroid pesticide-induced alterations in dopamine transporter function

    PubMed Central

    Elwan, Mohamed A.; Richardson, Jason R.; Guillot, Thomas S.; Caudle, W. Michael; Miller, Gary W.

    2016-01-01

    Parkinson’s disease (PD) is a progressive neurodegenerative disease affecting the nigrostriatal dopaminergic pathway. Several epidemiological studies have demonstrated an association between pesticide exposure and the incidence of PD. Studies from our laboratory and others have demonstrated that certain pesticides increase levels of the dopamine transporter (DAT), an integral component of dopaminergic neurotransmission and a gateway for dopaminergic neurotoxins. Here, we report that repeated exposure (3 injections over 2 weeks) of mice to two commonly used pyrethroid pesticides, deltamethrin (3 mg/kg) and permethrin (0.8 mg/kg), increases DAT-mediated dopamine uptake by 31 and 28%, respectively. Using cells stably expressing DAT, we determined that exposure (10 min) to deltamethrin and permethrin (1 nM–100 μM) had no effect on DAT-mediated dopamine uptake. Extending exposures to both pesticides for 30 min (10 μM) or 24 h (1, 5, and 10 μM) resulted in significant decrease in dopamine uptake. This reduction was not the result of competitive inhibition, loss of DAT protein, or cytotoxicity. However, there was an increase in DNA fragmentation, an index of apoptosis, in cells exhibiting reduced uptake at 30 min and 24 h. These data suggest that up-regulation of DAT by in vivo pyrethroid exposure is an indirect effect and that longer-term exposure of cells results in apoptosis. Since DAT can greatly affect the vulnerability of dopamine neurons to neurotoxicants, up-regulation of DAT by deltamethrin and permethrin may increase the susceptibility of dopamine neurons to toxic insult, which may provide insight into the association between pesticide exposure and PD. PMID:16005927

  1. Pyrethroid pesticide-induced alterations in dopamine transporter function

    SciTech Connect

    Elwan, Mohamed A.; Richardson, Jason R.; Guillot, Thomas S.; Caudle, W. Michael; Miller, Gary W. . E-mail: gary.miller@emory.edu

    2006-03-15

    Parkinson's disease (PD) is a progressive neurodegenerative disease affecting the nigrostriatal dopaminergic pathway. Several epidemiological studies have demonstrated an association between pesticide exposure and the incidence of PD. Studies from our laboratory and others have demonstrated that certain pesticides increase levels of the dopamine transporter (DAT), an integral component of dopaminergic neurotransmission and a gateway for dopaminergic neurotoxins. Here, we report that repeated exposure (3 injections over 2 weeks) of mice to two commonly used pyrethroid pesticides, deltamethrin (3 mg/kg) and permethrin (0.8 mg/kg), increases DAT-mediated dopamine uptake by 31 and 28%, respectively. Using cells stably expressing DAT, we determined that exposure (10 min) to deltamethrin and permethrin (1 nM-100 {mu}M) had no effect on DAT-mediated dopamine uptake. Extending exposures to both pesticides for 30 min (10 {mu}M) or 24 h (1, 5, and 10 {mu}M) resulted in significant decrease in dopamine uptake. This reduction was not the result of competitive inhibition, loss of DAT protein, or cytotoxicity. However, there was an increase in DNA fragmentation, an index of apoptosis, in cells exhibiting reduced uptake at 30 min and 24 h. These data suggest that up-regulation of DAT by in vivo pyrethroid exposure is an indirect effect and that longer-term exposure of cells results in apoptosis. Since DAT can greatly affect the vulnerability of dopamine neurons to neurotoxicants, up-regulation of DAT by deltamethrin and permethrin may increase the susceptibility of dopamine neurons to toxic insult, which may provide insight into the association between pesticide exposure and PD.

  2. Increased expression of the dopamine transporter leads to loss of dopamine neurons, oxidative stress and l-DOPA reversible motor deficits.

    PubMed

    Masoud, S T; Vecchio, L M; Bergeron, Y; Hossain, M M; Nguyen, L T; Bermejo, M K; Kile, B; Sotnikova, T D; Siesser, W B; Gainetdinov, R R; Wightman, R M; Caron, M G; Richardson, J R; Miller, G W; Ramsey, A J; Cyr, M; Salahpour, A

    2015-02-01

    The dopamine transporter is a key protein responsible for regulating dopamine homeostasis. Its function is to transport dopamine from the extracellular space into the presynaptic neuron. Studies have suggested that accumulation of dopamine in the cytosol can trigger oxidative stress and neurotoxicity. Previously, ectopic expression of the dopamine transporter was shown to cause damage in non-dopaminergic neurons due to their inability to handle cytosolic dopamine. However, it is unknown whether increasing dopamine transporter activity will be detrimental to dopamine neurons that are inherently capable of storing and degrading dopamine. To address this issue, we characterized transgenic mice that over-express the dopamine transporter selectively in dopamine neurons. We report that dopamine transporter over-expressing (DAT-tg) mice display spontaneous loss of midbrain dopamine neurons that is accompanied by increases in oxidative stress markers, 5-S-cysteinyl-dopamine and 5-S-cysteinyl-DOPAC. In addition, metabolite-to-dopamine ratios are increased and VMAT2 protein expression is decreased in the striatum of these animals. Furthermore, DAT-tg mice also show fine motor deficits on challenging beam traversal that are reversed with l-DOPA treatment. Collectively, our findings demonstrate that even in neurons that routinely handle dopamine, increased uptake of this neurotransmitter through the dopamine transporter results in oxidative damage, neuronal loss and l-DOPA reversible motor deficits. In addition, DAT over-expressing animals are highly sensitive to MPTP-induced neurotoxicity. The effects of increased dopamine uptake in these transgenic mice could shed light on the unique vulnerability of dopamine neurons in Parkinson's disease. PMID:25447236

  3. Urinary Dopamine as a Potential Index of the Transport Activity of Multidrug and Toxin Extrusion in the Kidney.

    PubMed

    Kajiwara, Moto; Ban, Tsuyoshi; Matsubara, Kazuo; Nakanishi, Yoichi; Masuda, Satohiro

    2016-01-01

    Dopamine is a cationic natriuretic catecholamine synthesized in proximal tubular cells (PTCs) of the kidney before secretion into the lumen, a key site of its action. However, the molecular mechanisms underlying dopamine secretion into the lumen remain unclear. Multidrug and toxin extrusion (MATE) is a H⁺/organic cation antiporter that is highly expressed in the brush border membrane of PTCs and mediates the efflux of organic cations, including metformin and cisplatin, from the epithelial cells into the urine. Therefore, we hypothesized that MATE mediates dopamine secretion, a cationic catecholamine, into the tubule lumen, thereby regulating natriuresis. Here, we show that [³H]dopamine uptake in human (h) MATE1-, hMATE-2K- and mouse (m) MATE-expressing cells exhibited saturable kinetics. Fluid retention and decreased urinary excretion of dopamine and Na⁺ were observed in Mate1-knockout mice compared to that in wild-type mice. Imatinib, a MATE inhibitor, inhibited [³H]dopamine uptake by hMATE1-, hMATE2-K- and mMATE1-expressing cells in a concentration-dependent manner. At clinically-relevant concentrations, imatinib inhibited [³H]dopamine uptake by hMATE1- and hMATE2-K-expressing cells. The urinary excretion of dopamine and Na⁺ decreased and fluid retention occurred in imatinib-treated mice. In conclusion, MATE transporters secrete renally-synthesized dopamine, and therefore, urinary dopamine has the potential to be an index of the MATE transporter activity. PMID:27483254

  4. N-8-Substituted benztropinamine analogs as selective dopamine transporter ligands.

    PubMed

    Grundt, Peter; Kopajtic, Theresa A; Katz, Jonathan L; Newman, Amy Hauck

    2005-12-15

    A series of N-8-substituted benztropinamines was synthesized and evaluated for binding at the dopamine (DAT), serotonin (SERT), norepinephrine (NET) transporters, and muscarinic M1 receptors. In general, the isosteric replacement of the C-3 benzhydrol ether of benztropine by a benzhydryl amino group was well tolerated at the DAT. However, for certain N-8 substituted derivatives, selectivity over muscarinic M1 receptor affinity was reduced. PMID:16213721

  5. Cocaine induction of dopamine transporter trafficking to the plasma membrane.

    PubMed

    Little, Karley Y; Elmer, Lawrence W; Zhong, Huailing; Scheys, Joshua O; Zhang, Lian

    2002-02-01

    Several previous human postmortem experiments have detected an increase in striatal [(3)H]WIN 35428 binding to the dopamine transporter (DAT) in chronic cocaine users. However, animal experiments have found considerable variability in DAT radioligand binding levels in brain after cocaine administration, perhaps caused by length and dose of treatment and type of radioligand used. The present experiments tested the hypothesis that [(3)H]WIN 35428 binding and [(3)H]dopamine uptake would be increased by exposure to cocaine through alterations in DAT cellular trafficking, rather than increased protein synthesis. Experiments were conducted in stably hDAT-transfected N2A cells and assessed the dose response and time course of cocaine effects on [(3)H]WIN 35428 binding to the DAT, [(3)H]dopamine uptake, measures of DAT protein and mRNA, as well as DAT subcellular location. Cocaine doses of 10(-6) M caused statistically significant increases in [(3)H]WIN 35428 binding and [(3)H]dopamine uptake after 12 and 3 h, respectively. Despite these increases in DAT function, there was no change in DAT total protein or mRNA. Immunofluorescence and biotinylation experiments indicated that cocaine treatment induced increases in plasma membrane DAT immunoreactivity and intracellular decreases. The present model system may further our understanding of regulatory alterations in DAT radioligand binding and function caused by cocaine exposure. PMID:11809869

  6. Missense dopamine transporter mutations associate with adult parkinsonism and ADHD

    PubMed Central

    Hansen, Freja H.; Skjørringe, Tina; Yasmeen, Saiqa; Arends, Natascha V.; Sahai, Michelle A.; Erreger, Kevin; Andreassen, Thorvald F.; Holy, Marion; Hamilton, Peter J.; Neergheen, Viruna; Karlsborg, Merete; Newman, Amy H.; Pope, Simon; Heales, Simon J.R.; Friberg, Lars; Law, Ian; Pinborg, Lars H.; Sitte, Harald H.; Loland, Claus; Shi, Lei; Weinstein, Harel; Galli, Aurelio; Hjermind, Lena E.; Møller, Lisbeth B.; Gether, Ulrik

    2014-01-01

    Parkinsonism and attention deficit hyperactivity disorder (ADHD) are widespread brain disorders that involve disturbances of dopaminergic signaling. The sodium-coupled dopamine transporter (DAT) controls dopamine homeostasis, but its contribution to disease remains poorly understood. Here, we analyzed a cohort of patients with atypical movement disorder and identified 2 DAT coding variants, DAT-Ile312Phe and a presumed de novo mutant DAT-Asp421Asn, in an adult male with early-onset parkinsonism and ADHD. According to DAT single-photon emission computed tomography (DAT-SPECT) scans and a fluoro-deoxy-glucose-PET/MRI (FDG-PET/MRI) scan, the patient suffered from progressive dopaminergic neurodegeneration. In heterologous cells, both DAT variants exhibited markedly reduced dopamine uptake capacity but preserved membrane targeting, consistent with impaired catalytic activity. Computational simulations and uptake experiments suggested that the disrupted function of the DAT-Asp421Asn mutant is the result of compromised sodium binding, in agreement with Asp421 coordinating sodium at the second sodium site. For DAT-Asp421Asn, substrate efflux experiments revealed a constitutive, anomalous efflux of dopamine, and electrophysiological analyses identified a large cation leak that might further perturb dopaminergic neurotransmission. Our results link specific DAT missense mutations to neurodegenerative early-onset parkinsonism. Moreover, the neuropsychiatric comorbidity provides additional support for the idea that DAT missense mutations are an ADHD risk factor and suggests that complex DAT genotype and phenotype correlations contribute to different dopaminergic pathologies. PMID:24911152

  7. Dopamine Transporter Activity Is Modulated by α-Synuclein.

    PubMed

    Butler, Brittany; Saha, Kaustuv; Rana, Tanu; Becker, Jonas P; Sambo, Danielle; Davari, Paran; Goodwin, J Shawn; Khoshbouei, Habibeh

    2015-12-01

    The duration and strength of the dopaminergic signal are regulated by the dopamine transporter (DAT). Drug addiction and neurodegenerative and neuropsychiatric diseases have all been associated with altered DAT activity. The membrane localization and the activity of DAT are regulated by a number of intracellular proteins. α-Synuclein, a protein partner of DAT, is implicated in neurodegenerative disease and drug addiction. Little is known about the regulatory mechanisms of the interaction between DAT and α-synuclein, the cellular location of this interaction, and the functional consequences of this interaction on the basal, amphetamine-induced DAT-mediated dopamine efflux, and membrane microdomain distribution of the transporter. Here, we found that the majority of DAT·α-synuclein protein complexes are found at the plasma membrane of dopaminergic neurons or mammalian cells and that the amphetamine-mediated increase in DAT activity enhances the association of these proteins at the plasma membrane. Further examination of the interaction of DAT and α-synuclein revealed a transient interaction between these two proteins at the plasma membrane. Additionally, we found DAT-induced membrane depolarization enhances plasma membrane localization of α-synuclein, which in turn increases dopamine efflux and enhances DAT localization in cholesterol-rich membrane microdomains. PMID:26442590

  8. Functional characterization of dopamine transporter in vivo using Drosophila melanogaster behavioral assays.

    PubMed

    Ueno, Taro; Kume, Kazuhiko

    2014-01-01

    Dopamine mediates diverse functions such as motivation, reward, attention, learning/memory and sleep/arousal. Recent studies using model organisms including the fruit fly, have elucidated various physiological functions of dopamine, and identified specific neural circuits for these functions. Flies with mutations in the Drosophila dopamine transporter (dDAT) gene show enhanced dopamine signaling, and short sleep and memory impairment phenotypes. However, understanding the mechanism by which dopamine signaling causes these phenotypes requires an understanding of the dynamics of dopamine release. Here we report the effects of dDAT expression on behavioral traits. We show that dDAT expression in a subset of dopaminergic neurons is sufficient for normal sleep. dDAT expression in other cell types such as Kenyon cells and glial cells can also rescue the short sleep phenotype of dDAT mutants. dDAT mutants also show a down-regulation of the D1-like dopamine receptor dDA1, and this phenotype is rescued when dDAT is expressed in the same cell types in which it rescues sleep. On the other hand, dDAT overexpression in mushroom bodies, which are the target of memory forming dopamine neurons, abolishes olfactory aversive memory. Our data demonstrate that expression of extrasynaptic dopamine transporters can rescue some aspects of dopamine signaling in dopamine transporter mutants. These results provide novel insights into regulatory systems that modulate dopamine signaling. PMID:25232310

  9. Relationship between cocaine-induced subjective effects and dopamine transporter occupancy

    SciTech Connect

    Volkow, N.D.; Fischman, M.; Wang, G.J.

    1997-05-01

    The ability of cocaine to occupy the dopamine transporter has been linked to its reinforcing properties. However, such a relationship has not been demonstrated in humans. Methods: Positron Emission Tomography and [C-11]cocaine were used to estimate dopamine transporter occupancies after different doses of cocaine in 18 active cocaine abusers. The ratio of the distribution volume of [C-11]cocaine in striatum to that in cerebellum, which corresponds to Bmax/Kd +1 and is insensitive to changes in cerebral blood flow, was our measure of dopamine transporter availability. In parallel subjective effects were measured to assess the relationship between dopamine transporter occupancy and cocaines behavioral effects. Intravenous cocaine produced a significant dose,-dependent blockade of dopamine transporters: 73 % for 0.6 mg/kg; 601/6 for 0.3 mg/kg; 48 % for 0.1 mg/kg iv and 40 % for 0.05 mg/kg. In addition, dopamine transporter occupancies were significantly correlated with cocaine plasma concentration (r = 0.55 p < 0.001). Cocaine also produced dose-dependent increases in self-reported ratings of {open_quotes}high{close_quotes} which were significantly correlated with the levels of dopamine transporter blockade. Discussion: These results provide the first documentation in humans that dopamine transporter occupancy is associated with cocaine induced subjective effects. They also suggest that dopamine transporter occupancies equal to or greater than 60% are required to produce significant effects on ratings of {open_quotes}high{close_quotes}.

  10. Molecular dynamics investigation of the adhesion mechanism acting between dopamine and the surface of dopamine-processed aramid fibers.

    PubMed

    Chai, Dongliang; Xie, Zhimin; Wang, Youshan; Liu, Li; Yum, Young-Jin

    2014-10-22

    Dopamine, as a universal material for surface treatment, can effectively improve the surface performance of aramid fibers. However, directly processing the surface of aramid fibers using dopamine currently incurs a high cost. To seek dopamine substitutes, one must first explore the adhesion mechanism responsible for binding the dopamine to the surface of the fiber. In this study, we construct an all-atomic molecular dynamics model of an aramid fiber before and after surface modification using dopamine. A force field based on condensed-phase optimized molecular potentials for atomistic simulation studies (COMPASS) is used. Using it, we analyze the surface adhesion mechanism of polydopamines aggregated by 21 kinds of molecular structures typically found on the surface of aramid fibers. The results show that a clear and smooth interface is formed between the polydopamine nanofilm layer and the surface of the aramid fiber. The high atomic density of the polydopamine in the small interface region is found to be conducive to noncovalent bonds of polydopamines with the surface of the aramid fiber. In addition, we investigate the works of adhesion of the 21 molecular structures typically found on the surface of aramid fibers. The results suggest that the work of adhesion of 5,6-indolequinone is the highest, followed by annular eumelanin molecules with annular planar structure. Straight-chain shaped dimers proved to be the molecules with the highest adhesion ability of the dihydroxyindole chain oligomers. Therefore, there is reason to suppose that more molecular structures (as above) can be formed by processing the surface of aramid fibers using dopamine by controlling the processing conditions. These molecular structures help improve the adhesion ability of the dopamine on the surface of the aramid fiber. Additionally, if these polydopamine molecules with high adhesion ability can be synthesized on a large scale, then new surface-processing materials are possible. PMID

  11. Nicotine increases dopamine transporter function in rat striatum through a trafficking-independent mechanism

    PubMed Central

    Middleton, Lisa S.; Apparsundaram, Subbu; King-Pospisil, Kelley A.; Dwoskin, Linda P.

    2007-01-01

    In previous in vivo voltammetry studies, acute nicotine administration increased striatal dopamine clearance. The current study aimed to determine whether nicotine also increases [3H]dopamine uptake across the time course of the previous voltammetry studies and whether dopamine transporter trafficking to the cell surface mediates the nicotine-induced augmentation of dopamine clearance in striatum. Rats were administered nicotine (0.32 mg/kg, s.c.); striatal synaptosomes were obtained 5, 10, 40 or 60 min later. Nicotine increased (25%) the Vmax of [3H]dopamine uptake at 10 and 40 min. To determine whether the increase in Vmax was due to an increase in dopamine transporter density, [3H]GBR 12935 (1-(2-[bis(4-fluorophenyl)methoxy]ethyl)-4-(3-phenylpropyl)piperazine dihydrochloride) binding was performed using rat striatal membranes; no differences were found between nicotine and saline control groups at 5, 10 or 40 min post-injection, indicating that nicotine did not increase striatal dopamine transporter density; however, [3H]GBR 12935 binding assays determine both cell surface and intracellular dopamine transporter. Changes in cellular dopamine transporter localization in striatum were determined using biotinylation and subfractionation approaches; no differences between nicotine and saline control groups were observed at 10 and 40 min post-injection. These results suggest that the nicotine-induced increase in dopamine uptake and clearance in striatum may occur via a trafficking-independent mechanism. PMID:17141211

  12. SKF-83566, a D1-dopamine receptor antagonist, inhibits the dopamine transporter.

    PubMed

    Stouffer, Melissa A; Ali, Solav; Reith, Maarten E A; Patel, Jyoti C; Sarti, Federica; Carr, Kenneth D; Rice, Margaret E

    2011-09-01

    Dopamine (DA) is an important transmitter in both motor and limbic pathways. We sought to investigate the role of D(1)-receptor activation in axonal DA release regulation in dorsal striatum using a D(1)-receptor antagonist, SKF-83566. Evoked DA release was monitored in rat striatal slices using fast-scan cyclic voltammetry. SKF-83566 caused a concentration-dependent increase in peak single-pulse evoked extracellular DA concentration, with a maximum increase of ∼ 65% in 5 μM SKF-83566. This was accompanied by a concentration-dependent increase in extracellular DA concentration clearance time. Both effects were occluded by nomifensine (1 μM), a dopamine transporter (DAT) inhibitor, suggesting that SKF-83566 acted via the DAT. We tested this by examining [(3)H]DA uptake into LLc-PK cells expressing rat DAT, and confirmed that SKF-83566 is a competitive DAT inhibitor with an IC(50) of 5.7 μM. Binding studies with [(3)H]CFT, a cocaine analog, showed even more potent action of SKF-83566 at the DAT cocaine binding site (IC(50) = 0.51 μM). Thus, data obtained using SKF-83566 as a D(1) DA-receptor antagonist may be confounded by concurrent DAT inhibition. More positively, however, SKF-83566 might be a candidate to attenuate cocaine effects in vivo because of the greater potency of this drug at the cocaine versus DA binding site of the DAT. PMID:21689106

  13. Restoration of the Dopamine Transporter through Cell Therapy Improves Dyskinesia in a Rat Model of Parkinson’s Disease

    PubMed Central

    Tomas, D.; Stanic, D.; Chua, H. K.; White, K.; Boon, W. C.; Horne, M.

    2016-01-01

    The dyskinesia of Parkinson's Disease is most likely due to excess levels of dopamine in the striatum. The mechanism may be due to aberrant synthesis but also, a deficiency or absence of the Dopamine Transporter. In this study we have examined the proposition that reinstating Dopamine Transporter expression in the striatum would reduce dyskinesia. We transplanted c17.2 cells that stably expressed the Dopamine Transporter into dyskinetic rats. There was a reduction in dyskinesia in rats that received grafts expressing the Dopamine Transporter. Strategies designed to increase Dopamine Transporter in the striatum may be useful in treating the dyskinesia associated with human Parkinson's Disease. PMID:27077649

  14. Low and high affinity dopamine transporter inhibitors block dopamine uptake within 5 sec of intravenous injection

    PubMed Central

    Yorgason, Jordan T.; Jones, Sara R.; España, Rodrigo A.

    2011-01-01

    Extensive evidence suggests that the reinforcing effects of cocaine involve inhibition of dopamine transporters (DAT) and subsequent increases in dopamine (DA) levels in the striatum. We have previously reported that cocaine inhibits the DAT within 4–5 sec of intravenous injection, matching the temporal profile of the behavioral and subjective effects of cocaine. Intravenous injection of GBR-12909, a high affinity, long-acting DAT inhibitor, also inhibits DA uptake within 5 sec. Given that high affinity, long-acting drugs are considered to have relatively low abuse potential, we found it intriguing that GBR-12909 had an onset profile similar to that of cocaine. To further explore the onset kinetics of both low and high affinity DAT inhibitors, we examined the effects of intravenous cocaine (1.5 mg/kg), methylphenidate (1.5 mg/kg), nomifensine (1.5 mg/kg), GBR-12909 (1.5 mg/kg), PTT (0.5 mg/kg), and WF23 (0.5 mg/kg) on electrically-evoked DA release and uptake in the nucleus accumbens core. Results indicate that all of the DAT inhibitors significantly inhibited DA uptake within 5 sec of injection. However, the timing of peak uptake inhibition varied greatly between the low and high affinity uptake inhibitors. Uptake inhibition following cocaine, methylphenidate, and nomifensine peaked 30 sec following injection. In contrast, peak effects for GBR-12909, PTT, and WF23 occurred between 20 and 60 min following injection. These observations suggest that the initial onset for intravenous DAT inhibitors is extremely rapid and does not appear to be dictated by a drug’s affinity. PMID:21402130

  15. Molecular Umbrella Transport

    PubMed Central

    Mehiri, Mohamed; Chen, Wen-Hua; Janout, Vaclav; Regen, Steven L.

    2009-01-01

    The ability of a series of molecular umbrellas, derived from cholic acid, L-lysine, spermidine and Cascade Blue, to cross fluid liposomal membranes made from 1-palmitoyl-2-oleyol-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol (POPG) (95/5, mol/mol) has been determined. In sharp contrast to the clasic “size/lipophilicity” rule of membrane transport, those molecular umbrellas that were larger in size and less lipophilic crossed these liposomal membranes more readily. The likely origin for this unusual behavior is briefly discussed. PMID:19140686

  16. Brain Region-Specific Trafficking of the Dopamine Transporter

    PubMed Central

    Block, Ethan R.; Nuttle, Jacob; Balcita-Pedicino, Judith Joyce; Caltagarone, John; Watkins, Simon C.

    2015-01-01

    The dopamine (DA) transporter (DAT) controls dopaminergic neurotransmission by removing extracellular DA. Although DA reuptake is proposed to be regulated by DAT traffic to and from the cell surface, the membrane trafficking system involved in the endocytic cycling of DAT in the intact mammalian brain has not been characterized. Hence, we performed immunolabeling and quantitative analysis of the subcellular and regional distribution of DAT using the transgenic knock-in mouse expressing hemagglutinin (HA) epitope-tagged DAT (HA-DAT) and by using a combination of electron microscopy and a novel method for immunofluorescence labeling of HA-DAT in acute sagittal brain slices. Both approaches demonstrated that, in midbrain somatodendritic regions, HA-DAT was present in the plasma membrane, endoplasmic reticulum, and Golgi complex, with a small fraction in early and recycling endosomes and an even smaller fraction in late endosomes and lysosomes. In the striatum and in axonal tracts between the midbrain and striatum, HA-DAT was detected predominantly in the plasma membrane, and quantitative analysis revealed increased DAT density in striatal compared with midbrain plasma membranes. Endosomes were strikingly rare and lysosomes were absent in striatal axons, in which there was little intracellular HA-DAT. Acute administration of amphetamine in vivo (60 min) or to slices ex vivo (10–60 min) did not result in detectable changes in DAT distribution. Altogether, these data provide evidence for regional differences in DAT plasma membrane targeting and retention and suggest a surprisingly low level of endocytic trafficking of DAT in the striatum along with limited DAT endocytic activity in somatodendritic areas. SIGNIFICANCE STATEMENT The dopamine transporter (DAT) is the key regulator of the dopamine neurotransmission in the CNS. In the present study, we developed a new approach for studying DAT localization and dynamics in intact neurons in acute sagittal brain slices from

  17. Monoamine transporters: insights from molecular dynamics simulations

    PubMed Central

    Grouleff, Julie; Ladefoged, Lucy Kate; Koldsø, Heidi; Schiøtt, Birgit

    2015-01-01

    The human monoamine transporters (MATs) facilitate the reuptake of the neurotransmitters serotonin, dopamine, and norepinephrine from the synaptic cleft. Imbalance in monoaminergic neurotransmission is linked to various diseases including major depression, attention deficit hyperactivity disorder, schizophrenia, and Parkinson’s disease. Inhibition of the MATs is thus an important strategy for treatment of such diseases. The MATs are sodium-coupled transport proteins belonging to the neurotransmitter/Na+ symporter (NSS) family, and the publication of the first high-resolution structure of a NSS family member, the bacterial leucine transporter LeuT, in 2005, proved to be a major stepping stone for understanding this family of transporters. Structural data allows for the use of computational methods to study the MATs, which in turn has led to a number of important discoveries. The process of substrate translocation across the membrane is an intrinsically dynamic process. Molecular dynamics simulations, which can provide atomistic details of molecular motion on ns to ms timescales, are therefore well-suited for studying transport processes. In this review, we outline how molecular dynamics simulations have provided insight into the large scale motions associated with transport of the neurotransmitters, as well as the presence of external and internal gates, the coupling between ion and substrate transport, and differences in the conformational changes induced by substrates and inhibitors. PMID:26528185

  18. An Artificial Molecular Transporter

    PubMed Central

    Schäfer, Christian; Ragazzon, Giulio; Colasson, Benoit; La Rosa, Marcello; Silvi, Serena

    2015-01-01

    Abstract The transport of substrates is one of the main tasks of biomolecular machines in living organisms. We report a synthetic small‐molecule system designed to catch, displace, and release molecular cargo in solution under external control. The system consists of a bistable rotaxane that behaves as an acid–base controlled molecular shuttle, whose ring component bears a tether ending with a nitrile group. The latter can be coordinated to a ruthenium complex that acts as the load, and dissociated upon irradiation with visible light. The cargo loading/unloading and ring transfer/return processes are reversible and can be controlled independently. The robust coordination bond ensures that the cargo remains attached to the device while the transport takes place. PMID:27308223

  19. A heterocyclic compound CE-103 inhibits dopamine reuptake and modulates dopamine transporter and dopamine D1-D3 containing receptor complexes.

    PubMed

    Sase, Ajinkya; Aher, Yogesh D; Saroja, Sivaprakasam R; Ganesan, Minu Karthika; Sase, Sunetra; Holy, Marion; Höger, Harald; Bakulev, Vasiliy; Ecker, Gerhard F; Langer, Thierry; Sitte, Harald H; Leban, Johann; Lubec, Gert

    2016-03-01

    A series of compounds have been reported to enhance memory via the DA system and herein a heterocyclic compound was tested for working memory (WM) enhancement. 2-((benzhydrylsulfinyl)methyl)thiazole (CE-103) was synthesized in a six-step synthesis. Binding of CE-103 to the dopamine (DAT), serotonin (SERT) and norepinephrine (NET) transporters and dopamine reuptake inhibition was tested as well as blood brain permeation and a screen for GPCR targets. 60 male Sprague Dawley rats were divided into six groups: CE-103 treated 1-10 mg/kg body weight, trained (TDI) and yoked (YDI) and vehicle treated, trained (TVI) and yoked (YVI) rats. Daily single intraperitoneal injections for a period of 10 days were administered and rats were tested in a radial arm maze (RAM). Hippocampi were taken 6 h following the last day of training and complexes containing the unphosphorylated or phosphorylated dopamine transporter (DAT) and complexes containing the D1-3 dopamine receptor subunits were determined. CE-103 was binding to the DAT but insignificantly to SERT or NET and dopamine reuptake was blocked specifically (IC50 = 14.73 μM). From day eight the compound was decreasing WM errors in the RAM significantly at both doses tested as compared to the vehicle controls. In the trained CE-103-treated group levels of the complex containing the phosphorylated dopamine transporter (pDAT) as well as D1R were decreased while levels of complexes containing D2R and D3R were significantly increased. CE-103 was shown to enhance spatial WM and DA reuptake inhibition with subsequent modulation of D1-3 receptors is proposed as a possible mechanism of action. PMID:26407764

  20. Living without DAT: Loss and compensation of the dopamine transporter gene in sauropsids (birds and reptiles)

    PubMed Central

    Lovell, P. V.; Kasimi, B.; Carleton, J.; Velho, T. A.; Mello, C. V.

    2015-01-01

    The dopamine transporter (DAT) is a major regulator of synaptic dopamine (DA) availability. It plays key roles in motor control and motor learning, memory formation, and reward-seeking behavior, is a major target of cocaine and methamphetamines, and has been assumed to be conserved among vertebrates. We have found, however, that birds, crocodiles, and lizards lack the DAT gene. We also found that the unprecedented loss of this important gene is compensated for by the expression of the noradrenaline transporter (NAT) gene, and not the serotonin transporter genes, in dopaminergic cells, which explains the peculiar pharmacology of the DA reuptake activity previously noted in bird striatum. This unexpected pattern contrasts with that of ancestral vertebrates (e.g. fish) and mammals, where the NAT gene is selectively expressed in noradrenergic cells. DA circuits in birds/reptiles and mammals thus operate with an analogous reuptake mechanism exerted by different genes, bringing new insights into gene expression regulation in dopaminergic cells and the evolution of a key molecular player in reward and addiction pathways. PMID:26364979

  1. Dopamine transporter occupancy by RTI-55, inhibition of dopamine transport and stimulation of locomotor activity

    SciTech Connect

    Gatley, S.J.; Gifford, A.N.; Volkow, N.D.

    1997-05-01

    Cocaine analogs such as RTI-55 (or {beta}CIT) with a higher affinity for the DAT are potentially useful as therapeutic drugs in cocaine abuse as well as for radiopharmaceutical use. Previously we showed that in mice RTI-55 (2 mg/Kg, i/p) reduced H-3 cocaine striatum-to-cerebellum ratios (St/Cb, {lg_bullet}) from 1.6 to 1.2 at 3 h after administration, with recovery by 12 h. In the present study we demonstrate a very similar time-course for transport {triangle} measured in striatal homo within 2 min of sacrifice. The maximum inhibition of uptake at about 1 h corresponded to about 80% of the control uptake rate, similar to the percent reduction in St/Cb. The time-course of the effect of this dose of RTI-55 on locomotor activity ({sq_bullet}) was complex, with a drop in the activity measure at 7 h, after a further injection of RTI-55, but activity remained higher than in saline controls. In spite of this complexity, which may be associated with stereotypies and/or exhaustion, the duration of increased activity is consistent with the duration of transporter blockade. These experiments support the notion that PET/SPECT measures of transporter occupancy accurately reflect transporter inhibition.

  2. Prolonged treatment with pramipexole promotes physical interaction of striatal dopamine D3 autoreceptors with dopamine transporters to reduce dopamine uptake.

    PubMed

    Castro-Hernández, Javier; Afonso-Oramas, Domingo; Cruz-Muros, Ignacio; Salas-Hernández, Josmar; Barroso-Chinea, Pedro; Moratalla, Rosario; Millan, Mark J; González-Hernández, Tomás

    2015-02-01

    The dopamine (DA) transporter (DAT), a membrane glycoprotein expressed in dopaminergic neurons, clears DA from extracellular space and is regulated by diverse presynaptic proteins like protein kinases, α-synuclein, D2 and D3 autoreceptors. DAT dysfunction is implicated in Parkinson's disease and depression, which are therapeutically treated by dopaminergic D2/D3 receptor (D2/D3R) agonists. It is, then, important to improve our understanding of interactions between D3R and DAT. We show that prolonged administration of pramipexole (0.1mg/kg/day, 6 to 21 days), a preferential D3R agonist, leads to a decrease in DA uptake in mouse striatum that reflects a reduction in DAT affinity for DA in the absence of any change in DAT density or subcellular distribution. The effect of pramipexole was absent in mice with genetically-deleted D3R (D3R(-/-)), yet unaffected in mice genetically deprived of D2R (D2R(-/-)). Pramipexole treatment induced a physical interaction between D3R and DAT, as assessed by co-immunoprecipitation and in situ proximity ligation assay. Furthermore, it promoted the formation of DAT dimers and DAT association with both D2R and α-synuclein, effects that were abolished in D3R(-/-) mice, yet unaffected in D2R(-/-) mice, indicating dependence upon D3R. Collectively, these data suggest that prolonged treatment with dopaminergic D3 agonists provokes a reduction in DA reuptake by dopaminergic neurons related to a hitherto-unsuspected modification of the DAT interactome. These observations provide novel insights into the long-term antiparkinson, antidepressant and additional clinical actions of pramipexole and other D3R agonists. PMID:25511804

  3. Neurotensin effect on dopamine release and calcium transport in rat striatum: interactions with diphenylalkylamine calcium antagonists.

    PubMed

    Battaini, F; Govoni, S; Di Giovine, S; Trabucchi, M

    1986-03-01

    The release of dopamine was investigated in rat striatal slices exposed in vitro to neurotensin. This peptide increased basal and K+-evoked dopamine release. Moreover neurotensin antagonized the flunarizine-induced inhibition of K+-stimulated dopamine release. The K+-evoked 45Ca2+ accumulation was also inhibited by flunarizine. This effect was antagonized by neurotensin. The results suggest that dopamine release in rat striatum is regulated by different molecular events also of peptidergic nature having as possible mechanism of action an influence on calcium ion movements. PMID:3713871

  4. ADHD-associated dopamine transporter, latrophilin and neurofibromin share a dopamine-related locomotor signature in Drosophila

    PubMed Central

    van der Voet, M; Harich, B; Franke, B; Schenck, A

    2016-01-01

    Attention-deficit/hyperactivity disorder (ADHD) is a common, highly heritable neuropsychiatric disorder with hyperactivity as one of the hallmarks. Aberrant dopamine signaling is thought to be a major theme in ADHD, but how this relates to the vast majority of ADHD candidate genes is illusive. Here we report a Drosophila dopamine-related locomotor endophenotype that is shared by pan-neuronal knockdown of orthologs of the ADHD-associated genes Dopamine transporter (DAT1) and Latrophilin (LPHN3), and of a gene causing a monogenic disorder with frequent ADHD comorbidity: Neurofibromin (NF1). The locomotor signature was not found in control models and could be ameliorated by methylphenidate, validating its relevance to symptoms of the disorder. The Drosophila ADHD endophenotype can be further exploited in high throughput to characterize the growing number of candidate genes. It represents an equally useful outcome measure for testing chemical compounds to define novel treatment options. PMID:25962619

  5. ADHD-associated dopamine transporter, latrophilin and neurofibromin share a dopamine-related locomotor signature in Drosophila.

    PubMed

    van der Voet, M; Harich, B; Franke, B; Schenck, A

    2016-04-01

    Attention-deficit/hyperactivity disorder (ADHD) is a common, highly heritable neuropsychiatric disorder with hyperactivity as one of the hallmarks. Aberrant dopamine signaling is thought to be a major theme in ADHD, but how this relates to the vast majority of ADHD candidate genes is illusive. Here we report a Drosophila dopamine-related locomotor endophenotype that is shared by pan-neuronal knockdown of orthologs of the ADHD-associated genes Dopamine transporter (DAT1) and Latrophilin (LPHN3), and of a gene causing a monogenic disorder with frequent ADHD comorbidity: Neurofibromin (NF1). The locomotor signature was not found in control models and could be ameliorated by methylphenidate, validating its relevance to symptoms of the disorder. The Drosophila ADHD endophenotype can be further exploited in high throughput to characterize the growing number of candidate genes. It represents an equally useful outcome measure for testing chemical compounds to define novel treatment options. PMID:25962619

  6. Dopamine Transporter Blockade Increases LTP in the CA1 Region of the Rat Hippocampus via Activation of the D3 Dopamine Receptor

    ERIC Educational Resources Information Center

    Swant, Jarod; Wagner, John J.

    2006-01-01

    Dopamine has been demonstrated to be involved in the modulation of long-term potentiation (LTP) in the CA1 region of the hippocampus. As monoamine transporter blockade will increase the actions of endogenous monoamine neurotransmitters, the effect of a dopamine transporter (DAT) antagonist on LTP was assessed using field excitatory postsynaptic…

  7. Effects of Ketamine and Ketamine Metabolites on Evoked Striatal Dopamine Release, Dopamine Receptors, and Monoamine Transporters.

    PubMed

    Can, Adem; Zanos, Panos; Moaddel, Ruin; Kang, Hye Jin; Dossou, Katinia S S; Wainer, Irving W; Cheer, Joseph F; Frost, Douglas O; Huang, Xi-Ping; Gould, Todd D

    2016-10-01

    Following administration at subanesthetic doses, (R,S)-ketamine (ketamine) induces rapid and robust relief from symptoms of depression in treatment-refractory depressed patients. Previous studies suggest that ketamine's antidepressant properties involve enhancement of dopamine (DA) neurotransmission. Ketamine is rapidly metabolized to (2S,6S)- and (2R,6R)-hydroxynorketamine (HNK), which have antidepressant actions independent of N-methyl-d-aspartate glutamate receptor inhibition. These antidepressant actions of (2S,6S;2R,6R)-HNK, or other metabolites, as well as ketamine's side effects, including abuse potential, may be related to direct effects on components of the dopaminergic (DAergic) system. Here, brain and blood distribution/clearance and pharmacodynamic analyses at DA receptors (D1-D5) and the DA, norepinephrine, and serotonin transporters were assessed for ketamine and its major metabolites (norketamine, dehydronorketamine, and HNKs). Additionally, we measured electrically evoked mesolimbic DA release and decay using fast-scan cyclic voltammetry following acute administration of subanesthetic doses of ketamine (2, 10, and 50 mg/kg, i.p.). Following ketamine injection, ketamine, norketamine, and multiple hydroxynorketamines were detected in the plasma and brain of mice. Dehydronorketamine was detectable in plasma, but concentrations were below detectable limits in the brain. Ketamine did not alter the magnitude or kinetics of evoked DA release in the nucleus accumbens in anesthetized mice. Neither ketamine's enantiomers nor its metabolites had affinity for DA receptors or the DA, noradrenaline, and serotonin transporters (up to 10 μM). These results suggest that neither the side effects nor antidepressant actions of ketamine or ketamine metabolites are associated with direct effects on mesolimbic DAergic neurotransmission. Previously observed in vivo changes in DAergic neurotransmission following ketamine administration are likely indirect. PMID:27469513

  8. Amphetamine activates calcium channels through dopamine transporter-mediated depolarization.

    PubMed

    Cameron, Krasnodara N; Solis, Ernesto; Ruchala, Iwona; De Felice, Louis J; Eltit, Jose M

    2015-11-01

    Amphetamine (AMPH) and its more potent enantiomer S(+)AMPH are psychostimulants used therapeutically to treat attention deficit hyperactivity disorder and have significant abuse liability. AMPH is a dopamine transporter (DAT) substrate that inhibits dopamine (DA) uptake and is implicated in DA release. Furthermore, AMPH activates ionic currents through DAT that modify cell excitability presumably by modulating voltage-gated channel activity. Indeed, several studies suggest that monoamine transporter-induced depolarization opens voltage-gated Ca(2+) channels (CaV), which would constitute an additional AMPH mechanism of action. In this study we co-express human DAT (hDAT) with Ca(2+) channels that have decreasing sensitivity to membrane depolarization (CaV1.3, CaV1.2 or CaV2.2). Although S(+)AMPH is more potent than DA in transport-competition assays and inward-current generation, at saturating concentrations both substrates indirectly activate voltage-gated L-type Ca(2+) channels (CaV1.3 and CaV1.2) but not the N-type Ca(2+) channel (CaV2.2). Furthermore, the potency to achieve hDAT-CaV electrical coupling is dominated by the substrate affinity on hDAT, with negligible influence of L-type channel voltage sensitivity. In contrast, the maximal coupling-strength (defined as Ca(2+) signal change per unit hDAT current) is influenced by CaV voltage sensitivity, which is greater in CaV1.3- than in CaV1.2-expressing cells. Moreover, relative to DA, S(+)AMPH showed greater coupling-strength at concentrations that induced relatively small hDAT-mediated currents. Therefore S(+)AMPH is not only more potent than DA at inducing hDAT-mediated L-type Ca(2+) channel currents but is a better depolarizing agent since it produces tighter electrical coupling between hDAT-mediated depolarization and L-type Ca(2+) channel activation. PMID:26162812

  9. 5-Hydroxytryptamine and dopamine transport by rat and human blood platelets

    PubMed Central

    Gordon, J.L.; Olverman, H.J.

    1978-01-01

    1 Uptake of 5-hydroxytryptamine (5-HT) by rat platelets in plasma was very rapid and diffusion did not contribute significantly at substrate concentrations that did not saturate the active transport. 2 Under conditions which allowed measurement of initial rates of uptake, kinetic analysis revealed a high affinity uptake mechanism for 5-HT (Km = 0.7 μM). 3 Uptake of dopamine was relatively slow and involved a lower affinity (Km = 70 μM) active transport process. Diffusion contributed significantly at concentrations that did not saturate the active transport. 4 5-HT competitively inhibited uptake of dopamine, and vice versa; Ki values for both amines were similar to their respective Km values for uptake. 5 Chlorimipramine, desmethylimipramine and benztropine were tested as uptake inhibitors. Each was equipotent against 5-HT and dopamine, although the absolute potency of the drugs varied greatly. Chlorimipramine was the most potent (Ki## 100 nM), and kinetic analysis revealed that the inhibition was competitive against both 5-HT and dopamine. 6 Similar results were obtained in studies with human platelets: Km values for 5-HT and dopamine were about 1 μM and 100 μM respectively. Activity profiles of inhibitors were also similar: each compound tested was equipotent against 5-HT and dopamine, and the two amines each competitively inhibited uptake of the other. 7 We conclude that dopamine is actively transported by platelets via the 5-HT uptake mechanism, but with a much lower affinity. There is no high-affinity dopamine-specific mechanism corresponding to that in the corpus striatum. Consequently although platelets may be valid models of transport in 5-hydroxytryptaminergic neurones, they should not be regarded as models for the dopamine transport mechanism found in dopaminergic neurones. PMID:623937

  10. Dopamine Transporter Genotype Conveys Familial Risk of Attention-Deficit/Hyperactivity Disorder through Striatal Activation

    ERIC Educational Resources Information Center

    Durston, Sarah; Fossella, John A.; Mulder, Martijn J.; Casey B. J.; Ziermans, Tim B.; Vessaz, M. Nathalie; Van Engeland, Herman

    2008-01-01

    The study examines the effect of the dopamine transporter (DAT1) genotype in attention-deficit/hyperactivity disorder (ADHD). The results confirm that DAT1 translates the genetic risk of ADHD through striatal activation.

  11. Estrogen mediated inhibition of dopamine transport in the striatum: regulation by G alpha i/o.

    PubMed

    Thompson, Tina L; Certain, Matthew E

    2005-03-28

    In the current study, the interaction between estrogen priming and dopamine D2 receptor activation on dopamine uptake in the striatum of ovariectomized female rats was investigated. Basal ADP-[(32)P(i)]ribosylation of G(i/o) was examined in synaptosomal membranes prepared from ovariectomized, estrogen primed or N-p-(isothiocyanatophenethyl) spiperone (NIPS) treated rats. [(32)P(i)]-incorporation was significantly increased (141%) in tissue from NIPS treated animals but attenuated (57%) in tissue from estrogen primed animals. Dopamine uptake kinetics were measured in vivo following manipulation of the heterotrimeric G-protein by pertussis toxin (0.5 microg, 48 h). Pertussis toxin significantly inhibited dopamine uptake at all concentrations of dopamine examined. Co-treatment with estrogen and pertussis toxin resulted in a further attenuation of dopamine transport at high but not low dopamine concentrations. These data are consistent with an estrogen mediated alteration of G-protein activity and support the hypothesis that estrogen may alter transporter activity through a modulation of dopamine D2 autoreceptor/G alpha(i/o) protein coupling. PMID:15792779

  12. Behavioral, biological, and chemical perspectives on atypical agents targeting the dopamine transporterɸ

    PubMed Central

    Reith, Maarten E.A.; Blough, Bruce E.; Hong, Weimin C.; Jones, Kymry T.; Schmitt, Kyle C.; Baumann, Michael H.; Partilla, John S.; Rothman, Richard B.; Katz, Jonathan L.

    2014-01-01

    Background Treatment of Stimulant-Use Disorders remains a formidable challenge, and the dopamine transporter (DAT) remains a potential target for antagonist or agonist-like substitution therapies. Methods This review focuses on DAT ligands, such as benztropine, GBR 12909, modafinil, and DAT substrates derived from phenethylamine or cathinone that have atypical DAT-inhibitor effects, either in vitro or in vivo. The compounds are described from a molecular mechanistic, behavioral, and medicinal-chemical perspective. Results Possible mechanisms for atypicality at the molecular level can be deduced from the conformational cycle for substrate translocation. For each conformation, a crystal structure of a bacterial homolog is available, with a possible role of cholesterol, which is also present in the crystal of drosophila DAT. Although there is a direct relationship between behavioral potencies of most DAT inhibitors and their DAT affinities, a number of compounds bind to the DAT and inhibit dopamine uptake but do not share cocaine-like effects. Such atypical behavior, depending on the compound, may be related to slow DAT association, combined sigma-receptor actions, or bias for cytosol-facing DAT. Some structures are sterically small enough to serve as DAT substrates but large enough to also inhibit transport. Such compounds may display partial DA releasing effects, and may be combined with release or uptake inhibition at other monoamine transporters. Conclusions Mechanisms of atypical DAT inhibitors may serve as targets for the development of treatments for stimulant abuse. These mechanisms are novel and their further exploration may produce compounds with unique therapeutic potential as treatments for stimulant abuse. PMID:25548026

  13. Chronic methylphenidate alters locomotor activity and dopamine transporters differently from cocaine.

    PubMed

    Izenwasser, S; Coy, A E; Ladenheim, B; Loeloff, R J; Cadet, J L; French, D

    1999-06-01

    Continuous infusion of cocaine produces partial behavioral tolerance to its locomotor activating effects, while daily injections produce sensitization. Methylphenidate binds with a similar affinity to cocaine at the dopamine transporter, but has a much lower affinity for the serotonin transporter than does cocaine. This study was done to compare the effects of chronic methylphenidate with chronic cocaine. The pattern of locomotor activity over a 7 day treatment period was significantly different from cocaine. Methylphenidate elevated activity on each day, compared to saline, yet neither tolerance to a continuous infusion of the drug, nor sensitization to repeated daily injections was produced. We have previously shown that neither of these treatments with cocaine produces significant alterations in dopamine transporter density 1 day after the end of treatment. In contrast, methylphenidate injections significantly decreased dopamine transporters in rostral caudate putamen, with no change in nucleus accumbens. Continuous infusion of methylphenidate had no effect on dopamine transporters in either brain region. These findings provide further evidence that different classes of dopamine uptake inhibitors may interact with the dopamine transporter in qualitatively different manners. Furthermore, it is possible that the inhibition of serotonin uptake by cocaine may contribute to the adaptations in behavioral activity that are seen during chronic treatment. PMID:10414438

  14. Dopamine transporter ligands: recent developments and therapeutic potential.

    PubMed

    Runyon, Scott P; Carroll, F Ivy

    2006-01-01

    The dopamine transporter (DAT) is a target for the development of pharmacotherapies for a number of central disorders including Parkinson's disease, Alzheimer's disease, schizophrenia, Tourette's syndrome, Lesch-Nyhan disease, attention deficit hyperactivity disorder (ADHD), obesity, depression, and stimulant abuse as well as normal aging. Considerable effort continues to be devoted to the development of new ligands for the DAT. In this review, we present some of the more interesting ligands developed during the last few years from the 3-phenytropane, 1,4-dialkylpiperazine, phenylpiperidine, and benztropine classes of DAT uptake inhibitors as well as a few less studied miscellaneous DAT uptake inhibitors. Studies related to the therapeutic potential of some of the more studied compounds are presented. A few of the compounds have been studied as pharmacotherapies for Parkinson's disease, ADHD, and obesity. However, most of the drug discovery studies have been directed toward pharmacotherapies for stimulant abuse (mainly cocaine). A number of the compounds showed decreased cocaine maintained responding in rhesus monkeys trained to self-administer cocaine. One compound, GBR 12,909, was evaluated in a Phase 1 clinical trial. PMID:17017960

  15. Association between amygdala reactivity and a dopamine transporter gene polymorphism.

    PubMed

    Bergman, O; Åhs, F; Furmark, T; Appel, L; Linnman, C; Faria, V; Bani, M; Pich, E M; Bettica, P; Henningsson, S; Manuck, S B; Ferrell, R E; Nikolova, Y S; Hariri, A R; Fredrikson, M; Westberg, L; Eriksson, E

    2014-01-01

    Essential for detection of relevant external stimuli and for fear processing, the amygdala is under modulatory influence of dopamine (DA). The DA transporter (DAT) is of fundamental importance for the regulation of DA transmission by mediating reuptake inactivation of extracellular DA. This study examined if a common functional variable number tandem repeat polymorphism in the 3' untranslated region of the DAT gene (SLC6A3) influences amygdala function during the processing of aversive emotional stimuli. Amygdala reactivity was examined by comparing regional cerebral blood flow, measured with positron emission tomography and [(15)O]water, during exposure to angry and neutral faces, respectively, in a Swedish sample comprising 32 patients with social anxiety disorder and 17 healthy volunteers. In a separate US sample, comprising 85 healthy volunteers studied with blood oxygen level-dependent functional magnetic resonance imaging, amygdala reactivity was assessed by comparing the activity during exposure to threatening faces and neutral geometric shapes, respectively. In both the Swedish and the US sample, 9-repeat carriers displayed higher amygdala reactivity than 10-repeat homozygotes. The results suggest that this polymorphism contributes to individual variability in amygdala reactivity. PMID:25093598

  16. Association between amygdala reactivity and a dopamine transporter gene polymorphism

    PubMed Central

    Bergman, O; Åhs, F; Furmark, T; Appel, L; Linnman, C; Faria, V; Bani, M; Pich, E M; Bettica, P; Henningsson, S; Manuck, S B; Ferrell, R E; Nikolova, Y S; Hariri, A R; Fredrikson, M; Westberg, L; Eriksson, E

    2014-01-01

    Essential for detection of relevant external stimuli and for fear processing, the amygdala is under modulatory influence of dopamine (DA). The DA transporter (DAT) is of fundamental importance for the regulation of DA transmission by mediating reuptake inactivation of extracellular DA. This study examined if a common functional variable number tandem repeat polymorphism in the 3′ untranslated region of the DAT gene (SLC6A3) influences amygdala function during the processing of aversive emotional stimuli. Amygdala reactivity was examined by comparing regional cerebral blood flow, measured with positron emission tomography and [15O]water, during exposure to angry and neutral faces, respectively, in a Swedish sample comprising 32 patients with social anxiety disorder and 17 healthy volunteers. In a separate US sample, comprising 85 healthy volunteers studied with blood oxygen level-dependent functional magnetic resonance imaging, amygdala reactivity was assessed by comparing the activity during exposure to threatening faces and neutral geometric shapes, respectively. In both the Swedish and the US sample, 9-repeat carriers displayed higher amygdala reactivity than 10-repeat homozygotes. The results suggest that this polymorphism contributes to individual variability in amygdala reactivity. PMID:25093598

  17. Photoinduced diffusion molecular transport

    NASA Astrophysics Data System (ADS)

    Rozenbaum, Viktor M.; Dekhtyar, Marina L.; Lin, Sheng Hsien; Trakhtenberg, Leonid I.

    2016-08-01

    We consider a Brownian photomotor, namely, the directed motion of a nanoparticle in an asymmetric periodic potential under the action of periodic rectangular resonant laser pulses which cause charge redistribution in the particle. Based on the kinetics for the photoinduced electron redistribution between two or three energy levels of the particle, the time dependence of its potential energy is derived and the average directed velocity is calculated in the high-temperature approximation (when the spatial amplitude of potential energy fluctuations is small relative to the thermal energy). The thus developed theory of photoinduced molecular transport appears applicable not only to conventional dichotomous Brownian motors (with only two possible potential profiles) but also to a much wider variety of molecular nanomachines. The distinction between the realistic time dependence of the potential energy and that for a dichotomous process (a step function) is represented in terms of relaxation times (they can differ on the time intervals of the dichotomous process). As shown, a Brownian photomotor has the maximum average directed velocity at (i) large laser pulse intensities (resulting in short relaxation times on laser-on intervals) and (ii) excited state lifetimes long enough to permit efficient photoexcitation but still much shorter than laser-off intervals. A Brownian photomotor with optimized parameters is exemplified by a cylindrically shaped semiconductor nanocluster which moves directly along a polar substrate due to periodically photoinduced dipole moment (caused by the repetitive excited electron transitions to a non-resonant level of the nanocylinder surface impurity).

  18. Increased vesicular monoamine transporter enhances dopamine release and opposes Parkinson disease-related neurodegeneration in vivo.

    PubMed

    Lohr, Kelly M; Bernstein, Alison I; Stout, Kristen A; Dunn, Amy R; Lazo, Carlos R; Alter, Shawn P; Wang, Minzheng; Li, Yingjie; Fan, Xueliang; Hess, Ellen J; Yi, Hong; Vecchio, Laura M; Goldstein, David S; Guillot, Thomas S; Salahpour, Ali; Miller, Gary W

    2014-07-01

    Disruption of neurotransmitter vesicle dynamics (transport, capacity, release) has been implicated in a variety of neurodegenerative and neuropsychiatric conditions. Here, we report a novel mouse model of enhanced vesicular function via bacterial artificial chromosome (BAC)-mediated overexpression of the vesicular monoamine transporter 2 (VMAT2; Slc18a2). A twofold increase in vesicular transport enhances the vesicular capacity for dopamine (56%), dopamine vesicle volume (33%), and basal tissue dopamine levels (21%) in the mouse striatum. The elevated vesicular capacity leads to an increase in stimulated dopamine release (84%) and extracellular dopamine levels (44%). VMAT2-overexpressing mice show improved outcomes on anxiety and depressive-like behaviors and increased basal locomotor activity (41%). Finally, these mice exhibit significant protection from neurotoxic insult by the dopaminergic toxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), as measured by reduced dopamine terminal damage and substantia nigra pars compacta cell loss. The increased release of dopamine and neuroprotection from MPTP toxicity in the VMAT2-overexpressing mice suggest that interventions aimed at enhancing vesicular capacity may be of therapeutic benefit in Parkinson disease. PMID:24979780

  19. Increased vesicular monoamine transporter enhances dopamine release and opposes Parkinson disease-related neurodegeneration in vivo

    PubMed Central

    Lohr, Kelly M.; Bernstein, Alison I.; Stout, Kristen A.; Dunn, Amy R.; Lazo, Carlos R.; Alter, Shawn P.; Wang, Minzheng; Li, Yingjie; Fan, Xueliang; Hess, Ellen J.; Yi, Hong; Vecchio, Laura M.; Goldstein, David S.; Guillot, Thomas S.; Salahpour, Ali; Miller, Gary W.

    2014-01-01

    Disruption of neurotransmitter vesicle dynamics (transport, capacity, release) has been implicated in a variety of neurodegenerative and neuropsychiatric conditions. Here, we report a novel mouse model of enhanced vesicular function via bacterial artificial chromosome (BAC)-mediated overexpression of the vesicular monoamine transporter 2 (VMAT2; Slc18a2). A twofold increase in vesicular transport enhances the vesicular capacity for dopamine (56%), dopamine vesicle volume (33%), and basal tissue dopamine levels (21%) in the mouse striatum. The elevated vesicular capacity leads to an increase in stimulated dopamine release (84%) and extracellular dopamine levels (44%). VMAT2-overexpressing mice show improved outcomes on anxiety and depressive-like behaviors and increased basal locomotor activity (41%). Finally, these mice exhibit significant protection from neurotoxic insult by the dopaminergic toxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), as measured by reduced dopamine terminal damage and substantia nigra pars compacta cell loss. The increased release of dopamine and neuroprotection from MPTP toxicity in the VMAT2-overexpressing mice suggest that interventions aimed at enhancing vesicular capacity may be of therapeutic benefit in Parkinson disease. PMID:24979780

  20. Dual Action of Zn2+ on the Transport Cycle of the Dopamine Transporter*

    PubMed Central

    Li, Yang; Hasenhuetl, Peter S.; Schicker, Klaus; Sitte, Harald H.; Freissmuth, Michael; Sandtner, Walter

    2015-01-01

    The dopamine transporter shapes dopaminergic neurotransmission by clearing extracellular dopamine and by replenishing vesicular stores. The dopamine transporter carries an endogenous binding site for Zn2+, but the nature of the Zn2+-dependent modulation has remained elusive: both, inhibition and stimulation of DAT have been reported. Here, we exploited the high time resolution of patch-clamp recordings to examine the effects of Zn2+ on the transport cycle of DAT: we recorded peak currents associated with substrate translocation and steady-state currents reflecting the forward transport mode of DAT. Zn2+ depressed the peak current but enhanced the steady-state current through DAT. The parsimonious explanation is preferential binding of Zn2+ to the outward facing conformation of DAT, which allows for an allosteric activation of DAT, in both, the forward transport mode and substrate exchange mode. We directly confirmed that Zn2+ dissociated more rapidly from the inward- than from the outward-facing state of DAT. Finally, we formulated a kinetic model for the action of Zn2+ on DAT that emulated all current experimental observations and accounted for all previous (in part contradictory) findings. Importantly, the model predicts that the intracellular Na+ concentration determines whether substrate uptake by DAT is stimulated or inhibited by Zn2+. This prediction was directly verified. The mechanistic framework provided by the current model is of relevance for the rational design of allosteric activators of DAT. These are of interest for treating de novo loss-of-function mutations of DAT associated with neuropsychiatric disorders such as attention deficit hyperactivity disorder (ADHD). PMID:26504078

  1. Dopamine transporter availability in clinically normal aging is associated with individual differences in white matter integrity

    PubMed Central

    Rieckmann, Anna; Hedden, Trey; Younger, Alayna P.; Sperling, Reisa A.; Johnson, Keith A.; Buckner, Randy L.

    2016-01-01

    Aging-related differences in white matter integrity, the presence of amyloid plaques, and density of biomarkers indicative of dopamine functions can be detected and quantified with in vivo human imaging. The primary aim of the present study was to investigate whether these imaging-based measures constitute independent imaging biomarkers in older adults, which would speak to the hypothesis that the aging brain is characterized by multiple independent neurobiological cascades. We assessed MRI-based markers of white matter integrity and PET-based marker of dopamine transporter density and amyloid deposition in the same set of 53 clinically normal individuals (age 65–87). A multiple regression analysis demonstrated that dopamine transporter availability is predicted by white matter integrity, which was detectable even after controlling for chronological age. Further post-hoc exploration revealed that dopamine transporter availability was further associated with systolic blood pressure, mirroring the established association between cardiovascular health and white matter integrity. Dopamine transporter availability was not associated with the presence of amyloid burden. Neurobiological correlates of dopamine transporter measures in aging are therefore likely unrelated to Alzheimer’s disease but are aligned with white matter integrity and cardiovascular risk. More generally, these results suggest that two common imaging markers of the aging brain that are typically investigated separately do not reflect independent neurobiological processes. PMID:26542307

  2. Sensitized Nucleus Accumbens Dopamine Terminal Responses to Methylphenidate and Dopamine Transporter Releasers after Intermittent-Access Self-Administration

    PubMed Central

    Calipari, Erin S.; Jones, Sara R.

    2014-01-01

    Long-access methylphenidate (MPH) self-administration has been shown to produce enhanced amphetamine potency at the dopamine transporter and concomitant changes in reinforcing efficacy, suggesting that MPH abuse may change the dopamine system in a way that promotes future drug abuse. While long-access self-administration paradigms have translational validity for cocaine, it may not be as relevant a model of MPH abuse, as it has been suggested that people often take MPH intermittently. Although previous work outlined the neurochemical and behavioral consequences of long-access MPH self-administration, it was not clear whether intermittent access (6 h session; 5min access/30min) would result in similar changes. For cocaine, long-access self-administration resulted in tolerance to cocaine’s effects on dopamine and behavior while intermittent-access resulted in sensitization. Here we assessed the neurochemical consequences of intermittent-access MPH self-administration on dopamine terminal function. We found increased maximal rates of uptake, increased stimulated release, and subsensitive D2-like autoreceptors. Consistent with previous work using extended-access MPH paradigms, the potencies of amphetamine and MPH, but not cocaine, were increased, demonstrating that unlike cocaine, MPH effects were not altered by the pattern of intake. Although the potency results suggest that MPH may share properties with releasers, dopamine release was increased following acute application of MPH, similar to cocaine, and in contrast to the release decreasing effects of amphetamine. Taken together, these data demonstrate that MPH exhibits properties of both blockers and releasers, and that the compensatory changes produced by MPH self-administration may increase the abuse liability of amphetamines, independent of the pattern of administration. PMID:24632529

  3. Sensitized nucleus accumbens dopamine terminal responses to methylphenidate and dopamine transporter releasers after intermittent-access self-administration.

    PubMed

    Calipari, Erin S; Jones, Sara R

    2014-07-01

    Long-access methylphenidate (MPH) self-administration has been shown to produce enhanced amphetamine potency at the dopamine transporter and concomitant changes in reinforcing efficacy, suggesting that MPH abuse may change the dopamine system in a way that promotes future drug abuse. While long-access self-administration paradigms have translational validity for cocaine, it may not be as relevant a model of MPH abuse, as it has been suggested that people often take MPH intermittently. Although previous work outlined the neurochemical and behavioral consequences of long-access MPH self-administration, it was not clear whether intermittent access (6 h session; 5 min access/30 min) would result in similar changes. For cocaine, long-access self-administration resulted in tolerance to cocaine's effects on dopamine and behavior while intermittent-access resulted in sensitization. Here we assessed the neurochemical consequences of intermittent-access MPH self-administration on dopamine terminal function. We found increased maximal rates of uptake, increased stimulated release, and subsensitive D2-like autoreceptors. Consistent with previous work using extended-access MPH paradigms, the potencies of amphetamine and MPH, but not cocaine, were increased, demonstrating that unlike cocaine, MPH effects were not altered by the pattern of intake. Although the potency results suggest that MPH may share properties with releasers, dopamine release was increased following acute application of MPH, similar to cocaine, and in contrast to the release decreasing effects of amphetamine. Taken together, these data demonstrate that MPH exhibits properties of both blockers and releasers, and that the compensatory changes produced by MPH self-administration may increase the abuse liability of amphetamines, independent of the pattern of administration. PMID:24632529

  4. Probes for the dopamine transporter: new leads toward a cocaine-abuse therapeutic--A focus on analogues of benztropine and rimcazole.

    PubMed

    Newman, Amy Hauck; Kulkarni, Santosh

    2002-09-01

    In an attempt to discover a cocaine-abuse pharmacotherapeutic, extensive investigation has been directed toward elucidating the molecular mechanisms underlying the reinforcing effects of this psychostimulant drug. The results of these studies have been consistent with the inhibition of dopamine uptake, at the dopamine transporter (DAT), which results in a rapid and excessive accumulation of extracellular dopamine in the synapse as being the mechanism primarily responsible for the locomotor stimulant actions of cocaine. Nevertheless, investigation of the serotonin (SERT) and norepinephrine (NET) transporters, as well as other receptor systems, with which cocaine either directly or indirectly interacts, has suggested that the DAT is not solely responsible for the reinforcing effects of cocaine. In an attempt to further elucidate the roles of these systems in the reinforcing effects of cocaine, selective molecular probes, in the form of drug molecules, have been designed, synthesized, and characterized. Many of these compounds bind potently and selectively to the DAT, block dopamine reuptake, and are behaviorally cocaine-like in animal models of psychostimulant abuse. However, there have been exceptions noted in several classes of dopamine uptake inhibitors that demonstrate behavioral profiles that are distinctive from cocaine. Structure-activity relationships between chemically diverse dopamine uptake inhibitors have suggested that different binding interactions, at the molecular level on the DAT, as well as divergent actions at the other monoamine transporters may be related to the differing pharmacological actions of these compounds, in vivo. These studies suggest that novel dopamine uptake inhibitors, which are structurally and pharmacologically distinct from cocaine, may be developed as potential cocaine-abuse therapeutics. PMID:12210554

  5. Correlation of striatal dopamine transporter imaging with post mortem substantia nigra cell counts.

    PubMed

    Kraemmer, Julia; Kovacs, Gabor G; Perju-Dumbrava, Laura; Pirker, Susanne; Traub-Weidinger, Tatiana; Pirker, Walter

    2014-12-01

    Dopamine transporter imaging is widely used for the differential diagnosis of parkinsonism. Only limited data are available on the relationship between striatal dopamine transporter binding and dopaminergic cell loss in the substantia nigra (SN). We analyzed postmortem SN cell counts in patients who had previously undergone dopamine transporter single-photon emission computed tomography (SPECT). Pathological diagnoses included Parkinson's disease (n = 1), dementia with Lewy bodies (n = 2), multiple system atrophy (n = 1), corticobasal degeneration (n = 2), atypical parkinsonism with multiple pathological conditions (n = 1), Alzheimer's disease (n = 1), and Creutzfeldt-Jakob disease (n = 1). [(12) (3) I]β-CIT SPECT had been performed in all subjects using a standardized protocol on the same triple-head gamma camera. The density of neuromelanin-containing and tyrosine hydroxylase-positive substantia nigra neurons/mm(2) was evaluated in paraffin-embedded tissue sections by morphometric methods. Mean disease duration at the time of dopamine transporter imaging was 2.3 years, and the mean interval from imaging to death was 29.3 months (range, 4-68 months). Visual analysis of dopamine transporter images showed reduced striatal uptake in all seven patients with neurodegenerative parkinsonism, but not in Alzheimer's and Creutzfeldt-Jakob disease cases. Averaged [(right+left)/2] striatal uptake was highly correlated with averaged SN cell counts (rs  = 0.98, P < 0.0005 for neuromelanin- and rs  = 0.96, P < 0.0005 for tyrosine hydroxylase-positive cells). Similar strong correlations were found in separate analyses for the right and left sides. Striatal dopamine transporter binding highly correlated with postmortem SN cell counts, confirming the validity of dopamine transporter imaging as an excellent in vivo marker of nigrostriatal dopaminergic degeneration. PMID:25048738

  6. Vesicular Glutamate Transport Promotes Dopamine Storage and Glutamate Corelease In Vivo

    PubMed Central

    Hnasko, Thomas S.; Chuhma, Nao; Zhang, Hui; Goh, Germaine Y.; Sulzer, David; Palmiter, Richard D.; Rayport, Stephen; Edwards, Robert H.

    2010-01-01

    SUMMARY Dopamine neurons in the ventral tegmental area (VTA) play an important role in the motivational systems underlying drug addiction, and recent work has suggested that they also release the excitatory neurotransmitter glutamate. To assess a physiological role for glutamate corelease, we disrupted the expression of vesicular glutamate transporter 2 selectively in dopamine neurons. The conditional knockout abolishes glutamate release from midbrain dopamine neurons in culture and severely reduces their excitatory synaptic output in mesoaccumbens slices. Baseline motor behavior is not affected, but stimulation of locomotor activity by cocaine is impaired, apparently through a selective reduction of dopamine stores in the projection of VTA neurons to ventral striatum. Glutamate co-entry promotes monoamine storage by increasing the pH gradient that drives vesicular monoamine transport. Remarkably, low concentrations of glutamate acidify synaptic vesicles more slowly but to a greater extent than equimolar Cl−, indicating a distinct, presynaptic mechanism to regulate quantal size. PMID:20223200

  7. Dopamine Transporters in Striatum Correlated with Deactivation in the Default Mode Network during Visuospatial Attention

    SciTech Connect

    Tomasi, D.; Fowler, J.; Tomasi, D.; Volkow, N.D.; Wang, R.L.; Telang, F.; Wang, Chang, L.; Ernst, T.; /Fowler, J.S.

    2009-06-01

    Dopamine and dopamine transporters (DAT, which regulate extracellular dopamine in the brain) are implicated in the modulation of attention but their specific roles are not well understood. Here we hypothesized that dopamine modulates attention by facilitation of brain deactivation in the default mode network (DMN). Thus, higher striatal DAT levels, which would result in an enhanced clearance of dopamine and hence weaker dopamine signals, would be associated to lower deactivation in the DMN during an attention task. For this purpose we assessed the relationship between DAT in striatum (measured with positron emission tomography and [{sup 11}C]cocaine used as DAT radiotracer) and brain activation and deactivation during a parametric visual attention task (measured with blood oxygenation level dependent functional magnetic resonance imaging) in healthy controls. We show that DAT availability in caudate and putamen had a negative correlation with deactivation in ventral parietal regions of the DMN (precuneus, BA 7) and a positive correlation with deactivation in a small region in the ventral anterior cingulate gyrus (BA 24/32). With increasing attentional load, DAT in caudate showed a negative correlation with load-related deactivation increases in precuneus. These findings provide evidence that dopamine transporters modulate neural activity in the DMN and anterior cingulate gyrus during visuospatial attention. Our findings suggest that dopamine modulates attention in part by regulating neuronal activity in posterior parietal cortex including precuneus (region involved in alertness) and cingulate gyrus (region deactivated in proportion to emotional interference). These findings suggest that the beneficial effects of stimulant medications (increase dopamine by blocking DAT) in inattention reflect in part their ability to facilitate the deactivation of the DMN.

  8. Dopamine D1, D2, D3 Receptors, Vesicular Monoamine Transporter Type-2 (VMAT2) and Dopamine Transporter (DAT) Densities in Aged Human Brain

    PubMed Central

    Sun, Jianjun; Xu, Jinbin; Cairns, Nigel J.; Perlmutter, Joel S.; Mach, Robert H.

    2012-01-01

    The dopamine D1, D2, D3 receptors, vesicular monoamine transporter type-2 (VMAT2), and dopamine transporter (DAT) densities were measured in 11 aged human brains (aged 77–107.8, mean: 91 years) by quantitative autoradiography. The density of D1 receptors, VMAT2, and DAT was measured using [3H]SCH23390, [3H]dihydrotetrabenazine, and [3H]WIN35428, respectively. The density of D2 and D3 receptors was calculated using the D3-preferring radioligand, [3H]WC-10 and the D2-preferring radioligand [3H]raclopride using a mathematical model developed previously by our group. Dopamine D1, D2, and D3 receptors are extensively distributed throughout striatum; the highest density of D3 receptors occurred in the nucleus accumbens (NAc). The density of the DAT is 10–20-fold lower than that of VMAT2 in striatal regions. Dopamine D3 receptor density exceeded D2 receptor densities in extrastriatal regions, and thalamus contained a high level of D3 receptors with negligible D2 receptors. The density of dopamine D1 linearly correlated with D3 receptor density in the thalamus. The density of the DAT was negligible in the extrastriatal regions whereas the VMAT2 was expressed in moderate density. D3 receptor and VMAT2 densities were in similar level between the aged human and aged rhesus brain samples, whereas aged human brain samples had lower range of densities of D1 and D2 receptors and DAT compared with the aged rhesus monkey brain. The differential density of D3 and D2 receptors in human brain will be useful in the interpretation of PET imaging studies in human subjects with existing radiotracers, and assist in the validation of newer PET radiotracers having a higher selectivity for dopamine D2 or D3 receptors. PMID:23185343

  9. Inhibition by dizocilpine (MK-801) of striatal dopamine release induced by MPTP and MPP+: possible action at the dopamine transporter.

    PubMed

    Clarke, P B; Reuben, M

    1995-01-01

    release evoked by amphetamine 0.3 microM and MPP+ 10 flM, attenuated responses to MPTP 50 AM and did not affect responses to 12 mM K+. MK-801 100 microM evinced a similar profile but was less effective.7. MK-801 inhibited [3H]-dopamine uptake in striatal synaptosomes with an IC5o of 115 M.8. It is concluded that high concentrations of MK-801 inhibit the acute dopamine release evoked by MPTP and MPP+ in synaptosomes. This antagonism may occur, at least in part, through inhibition of the cell membrane dopamine transporter. MPTP and MPP+ also appear to interact with brain nicotinic cholinoceptors but the functional consequences of this interaction are not yet clear. PMID:7881731

  10. Electron transport through molecular junctions

    NASA Astrophysics Data System (ADS)

    Zimbovskaya, Natalya A.; Pederson, Mark R.

    2011-12-01

    At present, metal-molecular tunnel junctions are recognized as important active elements in molecular electronics. This gives a strong motivation to explore physical mechanisms controlling electron transport through molecules. In the last two decades, an unceasing progress in both experimental and theoretical studies of molecular conductance has been demonstrated. In the present work we give an overview of theoretical methods used to analyze the transport properties of metal-molecular junctions as well as some relevant experiments and applications. After a brief general description of the electron transport through molecules we introduce a Hamiltonian which can be used to analyze electron-electron, electron-phonon and spin-orbit interactions. Then we turn to description of the commonly used transport theory formalisms including the nonequilibrium Green’s functions based approach and the approach based on the “master” equations. We discuss the most important effects which could be manifested through molecules in electron transport phenomena such as Coulomb, spin and Frank-Condon blockades, Kondo peak in the molecular conductance, negative differential resistance and some others. Bearing in mind that first principles electronic structure calculations are recognized as the indispensable basis of the theory of electron transport through molecules, we briefly discuss the main equations and some relevant applications of the density functional theory which presently is often used to analyze important characteristics of molecules and molecular clusters. Finally, we discuss some kinds of nanoelectronic devices built using molecules and similar systems such as carbon nanotubes, various nanowires and quantum dots.

  11. Atrial Natriuretic Peptide Stimulates Dopamine Tubular Transport by Organic Cation Transporters: A Novel Mechanism to Enhance Renal Sodium Excretion

    PubMed Central

    Kouyoumdzian, Nicolás M.; Rukavina Mikusic, Natalia L.; Kravetz, María C.; Lee, Brenda M.; Carranza, Andrea; Del Mauro, Julieta S.; Pandolfo, Marcela; Gironacci, Mariela M.; Gorzalczany, Susana; Toblli, Jorge E.; Fernández, Belisario E.

    2016-01-01

    The aim of this study was to demonstrate the effects of atrial natriuretic peptide (ANP) on organic cation transporters (OCTs) expression and activity, and its consequences on dopamine urinary levels, Na+, K+-ATPase activity and renal function. Male Sprague Dawley rats were infused with isotonic saline solution during 120 minutes and randomized in nine different groups: control, pargyline plus tolcapone (P+T), ANP, dopamine (DA), D-22, DA+D-22, ANP+D-22, ANP+DA and ANP+DA+D-22. Renal functional parameters were determined and urinary dopamine concentration was quantified by HPLC. Expression of OCTs and D1-receptor in membrane preparations from renal cortex tissues were determined by western blot and Na+, K+-ATPase activity was determined using in vitro enzyme assay. 3H-DA renal uptake was determined in vitro. Compared to P+T group, ANP and dopamine infusion increased diuresis, urinary sodium and dopamine excretion significantly. These effects were more pronounced in ANP+DA group and reversed by OCTs blockade by D-22, demonstrating that OCTs are implied in ANP stimulated-DA uptake and transport in renal tissues. The activity of Na+, K+-ATPase exhibited a similar fashion when it was measured in the same experimental groups. Although OCTs and D1-receptor protein expression were not modified by ANP, OCTs-dependent-dopamine tubular uptake was increased by ANP through activation of NPR-A receptor and protein kinase G as signaling pathway. This effect was reflected by an increase in urinary dopamine excretion, natriuresis, diuresis and decreased Na+, K+-ATPase activity. OCTs represent a novel target that links the activity of ANP and dopamine together in a common mechanism to enhance their natriuretic and diuretic effects. PMID:27392042

  12. The Roles of Dopamine Transport Inhibition and Dopamine Release Facilitation in Wake Enhancement and Rebound Hypersomnolence Induced by Dopaminergic Agents

    PubMed Central

    Gruner, John A.; Marcy, Val R.; Lin, Yin-Guo; Bozyczko-Coyne, Donna; Marino, Michael J.; Gasior, Maciej

    2009-01-01

    Study Objective: Rebound hypersomnolence (RHS: increased sleep following increased wake) is a limiting side-effect of many wake-promoting agents. In particular, RHS in the first few hours following wake appears to be associated with dopamine (DA)-releasing agents, e.g., amphetamine, but whether it can also be produced by DA transporter (DAT) inhibition alone is unknown. In these studies, DA-releasing and DAT-inhibiting agents and their interaction were systematically examined for their ability to increase wake and induce RHS. Design: Chronically implanted rats were evaluated in a blinded, pseudo-randomized design. Participants: 237 rats were used in these studies with 1 week between repeat tests. Interventions: Animals were habituated overnight and dosed the next day, 5 h after lights on, with test agents. Measurements and Results: Sleep/wake activity and RHS were evaluated using EEG/EMG recording up to 22 h post dosing. In vitro dopamine release was evaluated in rat synaptosomes. At doses that produced equal increases in wake, DA-releasing (amphetamine, methamphetamine, phentermine) and several DAT-inhibiting agents (cocaine, bupropion, and methylphenidate) produced RHS during the first few hours after the onset of sleep recovery. However, other DAT-inhibiting agents (mazindol, nomifensine, GBR-12909, and GBR-12935) did not produce RHS. Combination treatment with amphetamine and nomifensine produced waking activity greater than the sum of their individual activities alone while ameliorating the amphetamine-like RHS. In rat synaptosomes, nomifensine reduced the potency of amphetamine to induce DA release ∼270-fold, potentially explaining its action in ameliorating amphetamine-induced RHS. Conclusions: All DA releasing agents tested, and some DAT-inhibiting agents, produced RHS at equal wake-promoting doses. Thus amphetamine-like DA release appears sufficient for inducing RHS, but additional properties (pharmacologic and/or pharmacokinetic) evidently underlie RHS

  13. Axonal transport of muscarinic receptors in vesicles containing noradrenaline and dopamine-beta-hydroxylase.

    PubMed

    Laduron, P M

    1984-01-01

    Presynaptic muscarinic receptors labeled with [3H]dexetimide and noradrenaline in dog splenic nerves accumulated proximally to a ligature at the same rate of axonal transport. After fractionation by differential centrifugation, specific [3H]quinuclidinyl benzilate or [3H]dexetimide binding revealed a distribution profile similar to that of dopamine-beta-hydroxylase and noradrenaline. Subfractionation by density gradient centrifugation showed two peaks of muscarinic receptors; the peak of density 1.17 contained noradrenaline and dopamine-beta-hydroxylase whereas that of density 1.14 was devoid of noradrenaline. Therefore the foregoing experiments provide evidence that presynaptic muscarinic receptors are transported in sympathetic nerves in synaptic vesicles which are similar to those containing noradrenaline and dopamine-beta-hydroxylase. This suggests a possible coexistence of receptor and neurotransmitter in the same vesicle. PMID:6198205

  14. A novel heterocyclic compound targeting the dopamine transporter improves performance in the radial arm maze and modulates dopamine receptors D1-D3.

    PubMed

    Saroja, Sivaprakasam R; Aher, Yogesh D; Kalaba, Predrag; Aher, Nilima Y; Zehl, Martin; Korz, Volker; Subramaniyan, Saraswathi; Miklosi, Andras G; Zanon, Lisa; Neuhaus, Winfried; Höger, Harald; Langer, Thierry; Urban, Ernst; Leban, Johann; Lubec, Gert

    2016-10-01

    A series of compounds targeting the dopamine transporter (DAT) haS been shown to improve memory performance most probably by re-uptake inhibition. Although specific DAT inhibitors are available, there is limited information about specificity, mechanism and in particular the effect on dopamine receptors. It was therefore the aim of the study to test the DAT inhibitor 4-(diphenyl-methanesulfinylmethyl)-2-methyl-thiazole (code: CE-111), synthetized in our laboratory for the specificity to target DAT, for the effects upon spatial memory and for induced dopamine receptor modulation. Re-uptake inhibition was tested for DAT (IC50=3.2μM), serotonin transporter, SERT (IC50=272291μM) and noradrenaline transporter, NET (IC50=174μM). Spatial memory was studied in the radial arm maze (RAM) in male Sprague-Dawley rats that were intraperitoneally injected with CE-111 (1 or 10mg/kg body weight). Performance in the RAM was improved using 1 and 10mg/kg body weight of CE-111. Training and treatment effects on presynaptic, postsynaptic and extrasynaptic D1 and D2- receptors and dopamine receptor containing complexes as well as on activated DAT were observed. CE-111 was crossing the blood-brain barrier comparable to modafinil and was identified as effective to improve memory performance in the RAM. Dopamine re-uptake inhibition along with modulations in dopamine receptors are proposed as potential underlying mechanisms. PMID:27288589

  15. A Conserved Salt Bridge between Transmembrane Segments 1 and 10 Constitutes an Extracellular Gate in the Dopamine Transporter*

    PubMed Central

    Pedersen, Anders V.; Andreassen, Thorvald F.; Loland, Claus J.

    2014-01-01

    Neurotransmitter transporters play an important role in termination of synaptic transmission by mediating reuptake of neurotransmitter, but the molecular processes behind translocation are still unclear. The crystal structures of the bacterial homologue, LeuT, provided valuable insight into the structural and dynamic requirements for substrate transport. These structures support the existence of gating domains controlling access to a central binding site. On the extracellular side, access is controlled by the “thin gate” formed by an interaction between Arg-30 and Asp-404. In the human dopamine transporter (DAT), the corresponding residues are Arg-85 and Asp-476. Here, we present results supporting the existence of a similar interaction in DAT. The DAT R85D mutant has a complete loss of function, but the additional insertion of an arginine in opposite position (R85D/D476R), causing a charge reversal, results in a rescue of binding sites for the cocaine analogue [3H]CFT. Also, the coordination of Zn2+ between introduced histidines (R85H/D476H) caused a ∼2.5-fold increase in [3H]CFT binding (Bmax). Importantly, Zn2+ also inhibited [3H]dopamine transport in R85H/D476H, suggesting that a dynamic interaction is required for the transport process. Furthermore, cysteine-reactive chemistry shows that mutation of the gating residues causes a higher proportion of transporters to reside in the outward facing conformation. Finally, we show that charge reversal of the corresponding residues (R104E/E493R) in the serotonin transporter also rescues [3H](S)-citalopram binding, suggesting a conserved feature. Taken together, these data suggest that the extracellular thin gate is present in monoamine transporters and that a dynamic interaction is required for substrate transport. PMID:25339174

  16. A flow cytometry-based dopamine transporter binding assay using antagonist-conjugated quantum dots

    SciTech Connect

    Kovtun, Oleg; Ross, Emily; Tomlinson, Ian; Rosenthal, Sandra

    2012-01-01

    Here we present the development and validation of a flow cytometry-based dopamine transporter (DAT) binding assay that uses antagonist-conjugated quantum dots (QDs).We anticipate that our QD-based assay is of immediate value to the high throughput screening of novel DAT modulators.

  17. Phase I Report: Technetium Radiotracers for the Dopamine Transporter. [September 1998 - March 1999

    SciTech Connect

    Baldwin, R.N.

    1999-03-17

    This project (a) demonstrated specific dopamine transporter (DAT) uptake in vivo and metabolic stability of a radiolabelled cycloplentadieny rhenium compound in rats and baboons, (b) showed that cyclopentadieny tricarbonyl rhenium and technetium compounds conjugated tropanel could be made by metal transfer with ferrocenes; and (c) explored new methods of synthesizing these compounds under mild conditions.

  18. Interaction of Dopamine Transporter (DAT1) Genotype and Maltreatment for ADHD: A Latent Class Analysis

    ERIC Educational Resources Information Center

    Li, James J.; Lee, Steve S.

    2012-01-01

    Background: Although the association of the dopamine transporter (DAT1) gene and attention-deficit/hyperactivity disorder (ADHD) has been widely studied, far less is known about its potential interaction with environmental risk factors. Given that maltreatment is a replicated risk factor for ADHD, we explored the interaction between DAT1 and…

  19. Molecular size of the canine and human brain D2 dopamine receptor as determined by radiation inactivation

    SciTech Connect

    Lilly, L.; Fraser, C.M.; Jung, C.Y.; Seeman, P.; Venter, J.C.

    1983-07-01

    Target-size analysis (radiation inactivation) has been utilized for determination of the molecular size of the striatal D2 dopamine receptor of both canine and human membranes. The dog and human receptors were found to have a molecular size of 123,000 daltons. The identity of molecular size values is consistent with available pharmacological and biochemical evidence supporting D2 dopamine receptor identity in canine and human tissues. These data suggest that the canine receptor may be a valid model for molecular and structural investigation of the human D2 dopamine receptor.

  20. Functionally distinct dopamine and octopamine transporters in the CNS of the cabbage looper moth.

    PubMed

    Gallant, Pamela; Malutan, Tabita; McLean, Heather; Verellen, LouAnn; Caveney, Stanley; Donly, Cam

    2003-02-01

    A cDNA was cloned from the cabbage looper Trichoplusia ni based on similarity to other cloned dopamine transporters (DATs). The total nucleotide sequence is 3.8 kb in length and contains an open reading frame for a protein of 612 amino acids. The predicted moth DAT protein (TrnDAT) has greatest amino acid sequence identity with Drosophila melanogasterDAT (73%) and Caenorhabditis elegansDAT (51%). TrnDAT shares only 45% amino acid sequence identity with an octopamine transporter (TrnOAT) cloned recently from this moth. The functional properties of TrnDAT and TrnOAT were compared through transient heterologous expression in Sf9 cells. Both transporters have similar transport affinities for DA (Km 2.43 and 2.16 micro m, respectively). However, the competitive substrates octopamine and tyramine are more potent blockers of [3H]dopamine (DA) uptake by TrnOAT than by TrnDAT. D-Amphetamine is a strong inhibitor and l-norepinephrine a weak inhibitor of both transporters. TrnDAT-mediated DA uptake is approximately 100-fold more sensitive to selective blockers of vertebrate transporters of dopamine and norepinephrine, such as nisoxetine, nomifensine and dibenzazepine antidepressants, than TrnOAT-mediated DA uptake. TrnOAT is 10-fold less sensitive to cocaine than TrnDAT. None of the 15 monoamine uptake blockers tested was TrnOAT-selective. In situ hybridization shows that TrnDAT and TrnOAT transcripts are expressed by different sets of neurons in caterpillar brain and ventral nerve cord. These results show that the caterpillar CNS contains both a phenolamine transporter and a catecholamine transporter whereas in the three invertebrates whose genomes have been completely sequenced only a dopamine-selective transporter is found. PMID:12581206

  1. Protein kinase C-mediated phosphorylation and functional regulation of dopamine transporters in striatal synaptosomes.

    PubMed

    Vaughan, R A; Huff, R A; Uhl, G R; Kuhar, M J

    1997-06-13

    Dopamine transporters (DATs) are members of a family of Na+- and Cl--dependent neurotransmitter transporters responsible for the rapid clearance of dopamine from synaptic clefts. The predicted primary sequence of DAT contains numerous consensus phosphorylation sites. In this report we demonstrate that DATs undergo endogenous phosphorylation in striatal synaptosomes that is regulated by activators of protein kinase C. Rat striatal synaptosomes were metabolically labeled with [32P]orthophosphate, and solubilized homogenates were subjected to immunoprecipitation with an antiserum specific for DAT. Basal phosphorylation occurred in the absence of exogenous treatments, and the phosphorylation level was rapidly increased when synaptosomes were treated with the phosphatase inhibitors okadaic acid or calyculin. Treatment of synaptosomes with the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) also increased the level of phosphate incorporation. This occurred within 10 min and was dosedependent between 0.1 and 1 microM PMA. DAT phosphorylation was also significantly increased by two other protein kinase C activators, (-)-indolactam V and 1-oleoyl-2-acetyl-sn-glycerol. The inactive phorbol ester 4alpha-phorbol 12,13-didecanoate at 10 microM was without effect, and PMA-induced phosphorylation was blocked by treatment of synaptosomes with the protein kinase C inhibitors staurosporine and bisindoylmaleimide. These results indicate that DATs undergo rapid in vivo phosphorylation in response to protein kinase C activation and that a robust mechanism exists in synaptosomes for DAT dephosphorylation. Dopamine transport activity in synaptosomes was reduced by all treatments that promoted DAT phosphorylation, with comparable dose, time, and inhibitor characteristics. The change in transport activity was produced by a reduction in Vmax with no significant effect on the Km for dopamine. These results suggest that synaptosomal dopamine transport activity is regulated by

  2. Synthesis, Pharmacological Evaluation and Molecular Modeling Studies of Triazole Containing Dopamine D3 Receptor Ligands

    PubMed Central

    Peng, Xin; Wang, Qi; Mishra, Yogesh; Xu, Jinbin; Reichert, David E.; Malik, Maninder; Taylor, Michelle; Luedtke, Robert R.; Mach, Robert H.

    2015-01-01

    A series of 2-methoxyphenyl piperazine analogues containing a triazole ring were synthesized and their in vitro binding affinities at human dopamine D2 and D3 receptors were evaluated. Compounds 5b, 5c, 5d, and 4g, demonstrate high affinity for dopamine D3 receptors and moderate selectivity for the dopamine D3 versus D2 receptor subtypes. To further examine their potential as therapeutic agents, their intrinsic efficacy at both D2 and D3 receptors was determined using a forskolin-dependent adenylyl cyclase inhibition assay. Affinity at dopamine D4 and serotonin 5-HT1A receptors was also determined. In addition, information from previous molecular modeling studies of the binding of a panel of 163 structurally-related benzamide analogues at dopamine D2 and D3 receptors was applied to this series of compounds. The results of the modeling studies were consistent with our previous experimental data. More importantly, the modeling study results explained why the replacement of the amide linkage with the hetero-aromatic ring leads to a reduction in the affinity of these compounds at D3 receptors. PMID:25556097

  3. Intranasal Dopamine Reduces In Vivo [123I]FP-CIT Binding to Striatal Dopamine Transporter: Correlation with Behavioral Changes and Evidence for Pavlovian Conditioned Dopamine Response

    PubMed Central

    de Souza Silva, Maria A.; Mattern, Claudia; Decheva, Cvetana; Huston, Joseph P.; Sadile, Adolfo G.; Beu, Markus; Müller, H.-W.; Nikolaus, Susanne

    2016-01-01

    Purpose: Dopamine (DA), which does not cross the blood-brain barrier, has central and behavioral effects when administered via the nasal route. Neither the mechanisms of central action of intranasal dopamine (IN-DA), nor its mechanisms of diffusion and transport into the brain are well understood. We here examined whether IN-DA application influences dopamine transporter (DAT) binding in the dorsal striatum and assessed the extent of binding in relation to motor and exploratory behaviors. We hypothesized that, based on the finding of increased extracellular DA in the striatum induced by application of IN-DA, binding of [123I]FP-CIT to the DAT should be decreased due to competition at the receptor. Methods: Rats were administered 3 mg/kg IN-DA and vehicle (VEH), with IN-DA injection either preceding or following VEH. Then motor and exploratory behaviors (traveled distance, velocity, center time, sitting, rearing, head-shoulder motility, grooming) were assessed for 30 min in an open field prior to administration of [123I]FP-CIT. DAT binding after IN-DA and VEH was measured with small animal SPECT 2 h following administration of the radioligand. Results: (1) After IN-DA application, striatal DAT binding was significantly lower as compared to VEH, indicating that the nasally delivered DA had central action and increased DA levels comparable to that found previously with L-DOPA administration; and (2) DAT binding in response to intranasal VEH was lower when IN-DA application preceded VEH treatment. This finding is suggestive of Pavlovian conditioning of DA at the level of the DAT, since the DA treatment modified (decreased) the binding in response to the subsequent VEH treatment. VEH treatment also reduced motor and exploratory behaviors more when applied before, as compared to when it followed IN-DA application, also indicative of behavioral Pavlovian conditioning akin to that found upon application of various psychostimulant drugs. Conclusions: The results: (a

  4. Dissociable roles of dopamine and serotonin transporter function in a rat model of negative urgency.

    PubMed

    Yates, Justin R; Darna, Mahesh; Gipson, Cassandra D; Dwoskin, Linda P; Bardo, Michael T

    2015-09-15

    Negative urgency is a facet of impulsivity that reflects mood-based rash action and is associated with various maladaptive behaviors in humans. However, the underlying neural mechanisms of negative urgency are not fully understood. Several brain regions within the mesocorticolimbic pathway, as well as the neurotransmitters dopamine (DA) and serotonin (5-HT), have been implicated in impulsivity. Extracellular DA and 5-HT concentrations are regulated by DA transporters (DAT) and 5-HT transporters (SERT); thus, these transporters may be important molecular mechanisms underlying individual differences in negative urgency. The current study employed a reward omission task to model negative urgency in rats. During reward trials, a cue light signaled the non-contingent delivery of one sucrose pellet; immediately following the non-contingent reward, rats responded on a lever to earn sucrose pellets (operant phase). Omission trials were similar to reward trials, except that non-contingent sucrose was omitted following the cue light prior to the operant phase. As expected, contingent responding was higher following omission of expected reward than following delivery of expected reward, thus reflecting negative urgency. Upon completion of behavioral training, Vmax and Km were obtained from kinetic analysis of [(3)H]DA and [(3)H]5-HT uptake using synaptosomes prepared from nucleus accumbens (NAc), dorsal striatum (Str), medial prefrontal cortex (mPFC), and orbitofrontal cortex (OFC) isolated from individual rats. Vmax for DAT in NAc and for SERT in OFC were positively correlated with negative urgency scores. The current findings suggest that mood-based impulsivity (negative urgency) is associated with enhanced DAT function in NAc and SERT function in OFC. PMID:26005123

  5. Assessment of the in vitro binding of JHW 007, a dopamine transport inhibitor that blocks the effects of cocaine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Benztropine (BZT) and its analogues, like cocaine, bind to the dopamine transporter and block dopamine uptake. However, while BZT analogues bind the DAT with high affinity, they generally do not have cocaine-like behavioral effects. JHW 007 is a BZT analogue that displaces [3H]WIN 35,428 from the D...

  6. Altered dopamine transporter function and phosphorylation following chronic cocaine self-administration and extinction in rats.

    PubMed

    Ramamoorthy, Sammanda; Samuvel, Devadoss J; Balasubramaniam, Annamalai; See, Ronald E; Jayanthi, Lankupalle D

    2010-01-15

    Cocaine binds with the dopamine transporter (DAT), an effect that has been extensively implicated in its reinforcing effects. However, persisting adaptations in DAT regulation after cocaine self-administration have not been extensively investigated. Here, we determined the changes in molecular mechanisms of DAT regulation in the caudate-putamen (CPu) and nucleus accumbens (NAcc) of rats with a history of cocaine self-administration, followed by 3weeks of withdrawal under extinction conditions (i.e., no cocaine available). DA uptake was significantly higher in the CPu of cocaine-experienced animals as compared to saline-yoked controls. DAT V(max) was elevated in the CPu without changes in apparent affinity for DA. In spite of elevated CPu DAT activity, total and surface DAT density and DAT-PP2Ac (protein phosphatase 2A catalytic subunit) interaction remained unaltered, although p-Ser- DAT phosphorylation was elevated. In contrast to the CPu, there were no differences between cocaine and saline rats in the levels of DA uptake, DAT V(max) and K(m) values, total and surface DAT, p-Ser-DAT phosphorylation, or DAT-PP2Ac interactions in the NAcc. These results show that chronic cocaine self-administration leads to lasting, regionally specific alterations in striatal DA uptake and DAT-Ser phosphorylation. Such changes may be related to habitual patterns of cocaine-seeking observed during relapse. PMID:20035724

  7. Impact of disruption of secondary binding site S2 on dopamine transporter function.

    PubMed

    Zhen, Juan; Reith, Maarten E A

    2016-09-01

    The structures of the leucine transporter, drosophila dopamine transporter, and human serotonin transporter show a secondary binding site (designated S2 ) for drugs and substrate in the extracellular vestibule toward the membrane exterior in relation to the primary substrate recognition site (S1 ). The present experiments are aimed at disrupting S2 by mutating Asp476 and Ile159 to Ala. Both mutants displayed a profound decrease in [(3) H]DA uptake compared with wild-type associated with a reduced turnover rate kcat . This was not caused by a conformational bias as the mutants responded to Zn(2+) (10 μM) similarly as WT. The dopamine transporters with either the D476A or I159A mutation both displayed a higher Ki for dopamine for the inhibition of [3H](-)-2-β-carbomethoxy-3-β-(4-fluorophenyl)tropane binding than did the WT transporter, in accordance with an allosteric interaction between the S1 and S2 sites. The results provide evidence in favor of a general applicability of the two-site allosteric model of the Javitch/Weinstein group from LeuT to dopamine transporter and possibly other monoamine transporters. X-ray structures of transporters closely related to the dopamine (DA) transporter show a secondary binding site S2 in the extracellular vestibule proximal to the primary binding site S1 which is closely linked to one of the Na(+) binding sites. This work examines the relationship between S2 and S1 sites. We found that S2 site impairment severely reduced DA transport and allosterically reduced S1 site affinity for the cocaine analog [(3) H]CFT. Our results are the first to lend direct support for the application of the two-site allosteric model, advanced for bacterial LeuT, to the human DA transporter. The model states that, after binding of the first DA molecule (DA1 ) to the primary S1 site (along with Na(+) ), binding of a second DA (DA2 ) to the S2 site triggers, through an allosteric interaction, the release of DA1 and Na(+) into the cytoplasm. PMID

  8. Homology Modeling of Dopamine D2 and D3 Receptors: Molecular Dynamics Refinement and Docking Evaluation

    PubMed Central

    Platania, Chiara Bianca Maria; Salomone, Salvatore; Leggio, Gian Marco; Drago, Filippo; Bucolo, Claudio

    2012-01-01

    Dopamine (DA) receptors, a class of G-protein coupled receptors (GPCRs), have been targeted for drug development for the treatment of neurological, psychiatric and ocular disorders. The lack of structural information about GPCRs and their ligand complexes has prompted the development of homology models of these proteins aimed at structure-based drug design. Crystal structure of human dopamine D3 (hD3) receptor has been recently solved. Based on the hD3 receptor crystal structure we generated dopamine D2 and D3 receptor models and refined them with molecular dynamics (MD) protocol. Refined structures, obtained from the MD simulations in membrane environment, were subsequently used in molecular docking studies in order to investigate potential sites of interaction. The structure of hD3 and hD2L receptors was differentiated by means of MD simulations and D3 selective ligands were discriminated, in terms of binding energy, by docking calculation. Robust correlation of computed and experimental Ki was obtained for hD3 and hD2L receptor ligands. In conclusion, the present computational approach seems suitable to build and refine structure models of homologous dopamine receptors that may be of value for structure-based drug discovery of selective dopaminergic ligands. PMID:22970199

  9. Homology modeling of dopamine D2 and D3 receptors: molecular dynamics refinement and docking evaluation.

    PubMed

    Platania, Chiara Bianca Maria; Salomone, Salvatore; Leggio, Gian Marco; Drago, Filippo; Bucolo, Claudio

    2012-01-01

    Dopamine (DA) receptors, a class of G-protein coupled receptors (GPCRs), have been targeted for drug development for the treatment of neurological, psychiatric and ocular disorders. The lack of structural information about GPCRs and their ligand complexes has prompted the development of homology models of these proteins aimed at structure-based drug design. Crystal structure of human dopamine D(3) (hD(3)) receptor has been recently solved. Based on the hD(3) receptor crystal structure we generated dopamine D(2) and D(3) receptor models and refined them with molecular dynamics (MD) protocol. Refined structures, obtained from the MD simulations in membrane environment, were subsequently used in molecular docking studies in order to investigate potential sites of interaction. The structure of hD(3) and hD(2L) receptors was differentiated by means of MD simulations and D(3) selective ligands were discriminated, in terms of binding energy, by docking calculation. Robust correlation of computed and experimental K(i) was obtained for hD(3) and hD(2L) receptor ligands. In conclusion, the present computational approach seems suitable to build and refine structure models of homologous dopamine receptors that may be of value for structure-based drug discovery of selective dopaminergic ligands. PMID:22970199

  10. A Role for Accumbal Glycine Receptors in Modulation of Dopamine Release by the Glycine Transporter-1 Inhibitor Org25935

    PubMed Central

    Lidö, Helga Höifödt; Ericson, Mia; Marston, Hugh; Söderpalm, Bo

    2010-01-01

    Accumbal glycine modulates basal and ethanol-induced dopamine levels in the nucleus accumbens (nAc) as well as voluntary ethanol consumption. Also, systemic administration of the glycine transporter-1 inhibitor Org25935 elevates dopamine levels in nAc, prevents a further ethanol-induced dopamine elevation and robustly and dose-dependently decreases ethanol consumption in rats. Here we investigated whether Org25935 applied locally in nAc modulates dopamine release, and whether accumbal glycine receptors or NMDA receptors are involved in this tentative effect. We also addressed whether Org25935 and ethanol applied locally in nAc interact with dopamine levels, as seen after systemic administration. We used in vivo microdialysis coupled to HPLC-ED in freely moving male Wistar rats to monitor dopamine output in nAc after local perfusion of Org25935 alone, with ethanol, or Org25935-perfusion after pre-treatment with the glycine receptor antagonist strychnine or the NMDA receptor glycine site antagonist L-701.324. Local Org25935 increased extracellular dopamine levels in a subpopulation of rats. Local strychnine, but not systemic L-701.324, antagonized the dopamine-activating effect of Org25935. Ethanol failed to induce a dopamine overflow in the subpopulation responding to Org25935 with a dopamine elevation. The study supports a role for accumbal glycine receptors rather than NMDA receptor signaling in the dopamine-activating effect of Org25935. The results further indicate that the previously reported systemic Org25935–ethanol interaction with regard to accumbal dopamine is localized to the nAc. This adds to the growing evidence for the glycine receptor as an important player in the dopamine reward circuitry and in ethanol's effects within this system. PMID:21556278

  11. Development of the dopamine transporter selective RTI-336 as a pharmacotherapy for cocaine abuse.

    PubMed

    Carroll, F Ivy; Howard, James L; Howell, Leonard L; Fox, Barbara S; Kuhar, Michael J

    2006-01-01

    The discovery and preclinical development of selective dopamine reuptake inhibitors as potential pharmacotherapies for treating cocaine addiction are presented. The studies are based on the hypothesis that a dopamine reuptake inhibitor is expected to partially substitute for cocaine, thus decreasing cocaine self-administration and minimizing the craving for cocaine. This type of indirect agonist therapy has been highly effective for treating smoking addiction (nicotine replacement therapy) and heroin addiction (methadone). To be an effective pharmacotherapy for cocaine addiction, the potential drug must be safe, long-acting, and have minimal abuse potential. We have developed several 3-phenyltropane analogs that are potent dopamine uptake inhibitors, and some are selective for the dopamine transporter relative to the serotonin and norepinephrine transporters. In animal studies, these compounds substitute for cocaine, reduce the intake of cocaine in rats and rhesus monkeys trained to self-administer cocaine, and have demonstrated a slow onset and long duration of action and lack of sensitization. The 3-phenyltropane analogs were also tested in a rhesus monkey self-administration model to define their abuse potential relative to cocaine. Based on these studies, 3beta-(4-chlorophenyl)-2beta-[3-(4'-methylphenyl)isoxazol-5-yl]tropane (RTI-336) has been selected for preclinical development. PMID:16584128

  12. Nonclassical Pharmacology of the Dopamine Transporter: Atypical Inhibitors, Allosteric Modulators, and Partial Substrates

    PubMed Central

    Rothman, Richard B.; Reith, Maarten E. A.

    2013-01-01

    The dopamine transporter (DAT) is a sodium-coupled symporter protein responsible for modulating the concentration of extraneuronal dopamine in the brain. The DAT is a principle target of various psychostimulant, nootropic, and antidepressant drugs, as well as certain drugs used recreationally, including the notoriously addictive stimulant cocaine. DAT ligands have traditionally been divided into two categories: cocaine-like inhibitors and amphetamine-like substrates. Whereas inhibitors block monoamine uptake by the DAT but are not translocated across the membrane, substrates are actively translocated and trigger DAT-mediated release of dopamine by reversal of the translocation cycle. Because both inhibitors and substrates increase extraneuronal dopamine levels, it is often assumed that all DAT ligands possess an addictive liability equivalent to that of cocaine. However, certain recently developed ligands, such as atypical benztropine-like DAT inhibitors with reduced or even a complete lack of cocaine-like rewarding effects, suggest that addictiveness is not a constant property of DAT-affecting compounds. These atypical ligands do not conform to the classic preconception that all DAT inhibitors (or substrates) are functionally and mechanistically alike. Instead, they suggest the possibility that the DAT exhibits some of the ligand-specific pleiotropic functional qualities inherent to G-protein–coupled receptors. That is, ligands with different chemical structures induce specific conformational changes in the transporter protein that can be differentially transduced by the cell, ultimately eliciting unique behavioral and psychological effects. The present overview discusses compounds with conformation-specific activity, useful not only as tools for studying the mechanics of dopamine transport, but also as leads for medication development in addictive disorders. PMID:23568856

  13. Spontaneous Inward Opening of the Dopamine Transporter Is Triggered by PIP2-Regulated Dynamics of the N-Terminus

    PubMed Central

    2015-01-01

    We present the dynamic mechanism of concerted motions in a full-length molecular model of the human dopamine transporter (hDAT), a member of the neurotransmitter/sodium symporter (NSS) family, involved in state-to-state transitions underlying function. The findings result from an analysis of unbiased atomistic molecular dynamics simulation trajectories (totaling >14 μs) of the hDAT molecule immersed in lipid membrane environments with or without phosphatidylinositol 4,5-biphosphate (PIP2) lipids. The N-terminal region of hDAT (N-term) is shown to have an essential mechanistic role in correlated rearrangements of specific structural motifs relevant to state-to-state transitions in the hDAT. The mechanism involves PIP2-mediated electrostatic interactions between the N-term and the intracellular loops of the transporter molecule. Quantitative analyses of collective motions in the trajectories reveal that these interactions correlate with the inward-opening dynamics of hDAT and are allosterically coupled to the known functional sites of the transporter. The observed large-scale motions are enabled by specific reconfiguration of the network of ionic interactions at the intracellular end of the protein. The isomerization to the inward-facing state in hDAT is accompanied by concomitant movements in the extracellular vestibule and results in the release of an Na+ ion from the Na2 site and destabilization of the substrate dopamine in the primary substrate binding S1 site. The dynamic mechanism emerging from the findings highlights the involvement of the PIP2-regulated interactions between the N-term and the intracellular loop 4 in the functionally relevant conformational transitions that are also similar to those found to underlie state-to-state transitions in the leucine transporter (LeuT), a prototypical bacterial homologue of the NSS. PMID:26255829

  14. Amphetamine activates Rho GTPase signaling to mediate dopamine transporter internalization and acute behavioral effects of amphetamine

    PubMed Central

    Wheeler, David S.; Underhill, Suzanne M.; Stolz, Donna B.; Murdoch, Geoffrey H.; Thiels, Edda; Romero, Guillermo; Amara, Susan G.

    2015-01-01

    Acute amphetamine (AMPH) exposure elevates extracellular dopamine through a variety of mechanisms that include inhibition of dopamine reuptake, depletion of vesicular stores, and facilitation of dopamine efflux across the plasma membrane. Recent work has shown that the DAT substrate AMPH, unlike cocaine and other nontransported blockers, can also stimulate endocytosis of the plasma membrane dopamine transporter (DAT). Here, we show that when AMPH enters the cytoplasm it rapidly stimulates DAT internalization through a dynamin-dependent, clathrin-independent process. This effect, which can be observed in transfected cells, cultured dopamine neurons, and midbrain slices, is mediated by activation of the small GTPase RhoA. Inhibition of RhoA activity with C3 exotoxin or a dominant-negative RhoA blocks AMPH-induced DAT internalization. These actions depend on AMPH entry into the cell and are blocked by the DAT inhibitor cocaine. AMPH also stimulates cAMP accumulation and PKA-dependent inactivation of RhoA, thus providing a mechanism whereby PKA- and RhoA-dependent signaling pathways can interact to regulate the timing and robustness of AMPH’s effects on DAT internalization. Consistent with this model, the activation of D1/D5 receptors that couple to PKA in dopamine neurons antagonizes RhoA activation, DAT internalization, and hyperlocomotion observed in mice after AMPH treatment. These observations support the existence of an unanticipated intracellular target that mediates the effects of AMPH on RhoA and cAMP signaling and suggest new pathways to target to disrupt AMPH action. PMID:26553986

  15. Amphetamine activates Rho GTPase signaling to mediate dopamine transporter internalization and acute behavioral effects of amphetamine.

    PubMed

    Wheeler, David S; Underhill, Suzanne M; Stolz, Donna B; Murdoch, Geoffrey H; Thiels, Edda; Romero, Guillermo; Amara, Susan G

    2015-12-22

    Acute amphetamine (AMPH) exposure elevates extracellular dopamine through a variety of mechanisms that include inhibition of dopamine reuptake, depletion of vesicular stores, and facilitation of dopamine efflux across the plasma membrane. Recent work has shown that the DAT substrate AMPH, unlike cocaine and other nontransported blockers, can also stimulate endocytosis of the plasma membrane dopamine transporter (DAT). Here, we show that when AMPH enters the cytoplasm it rapidly stimulates DAT internalization through a dynamin-dependent, clathrin-independent process. This effect, which can be observed in transfected cells, cultured dopamine neurons, and midbrain slices, is mediated by activation of the small GTPase RhoA. Inhibition of RhoA activity with C3 exotoxin or a dominant-negative RhoA blocks AMPH-induced DAT internalization. These actions depend on AMPH entry into the cell and are blocked by the DAT inhibitor cocaine. AMPH also stimulates cAMP accumulation and PKA-dependent inactivation of RhoA, thus providing a mechanism whereby PKA- and RhoA-dependent signaling pathways can interact to regulate the timing and robustness of AMPH's effects on DAT internalization. Consistent with this model, the activation of D1/D5 receptors that couple to PKA in dopamine neurons antagonizes RhoA activation, DAT internalization, and hyperlocomotion observed in mice after AMPH treatment. These observations support the existence of an unanticipated intracellular target that mediates the effects of AMPH on RhoA and cAMP signaling and suggest new pathways to target to disrupt AMPH action. PMID:26553986

  16. Association study of a dopamine transporter polymorphism and attention deficit hyperactivity disorder in UK and Turkish samples.

    PubMed

    Curran, S; Mill, J; Tahir, E; Kent, L; Richards, S; Gould, A; Huckett, L; Sharp, J; Batten, C; Fernando, S; Ozbay, F; Yazgan, Y; Simonoff, E; Thompson, M; Taylor, E; Asherson, P

    2001-07-01

    Molecular genetic studies in attention deficit hyperactivity disorder (ADHD) have focussed on candidate genes within the dopamine system, which is thought to be the main site of action of stimulant drugs, the primary pharmacological treatment of the disorder. Of particular interest are findings with the dopamine transporter gene (DAT1), since stimulant drugs interact directly with the transporter protein. To date, there have been eight published association studies of ADHD with a 480 base-pair allele of a variable number tandem repeat (VNTR) polymorphism in the 3'-untranslated region of the gene, five that support an association and three against. We have analysed the same VNTR marker in a dataset of UK Caucasian children and an independent dataset of Turkish Caucasian children with DSM-IV ADHD, using the transmission disequilibrium test (TDT). Results from the UK (chi(2) = 8.97, P = 0.001, OR = 1.95), but not the Turkish sample (chi(2) = 0.93, P = 0.34) support association and linkage between genetic variation at the DAT1 locus and ADHD. When considered alongside evidence from other published reports, there is only modest evidence for the association, consistent with a very small main effect for the 480-bp allele (chi(2) = 3.45, P = 0.06, OR = 1.15), however we find significant evidence of heterogeneity between the combined dataset (chi(2) = 22.64, df = 8, P = 0.004). PMID:11443527

  17. Fluorine-18 Radiolabeled PET Tracers for Imaging Monoamine Transporters: Dopamine, Serotonin, and Norepinephrine

    PubMed Central

    Stehouwer, Jeffrey S.; Goodman, Mark M.

    2009-01-01

    Synopsis This review focuses on the development of fluorine-18 radiolabeled PET tracers for imaging the dopamine transporter (DAT), serotonin transporter (SERT), and norepinephrine transporter (NET). All successful DAT PET tracers reported to date are members of the 3β-phenyl tropane class and are synthesized from cocaine. Currently available carbon-11 SERT PET tracers come from both the diphenylsulfide and 3β-phenyl nortropane class, but so far only the nortropanes have found success with fluorine-18 derivatives. NET imaging has so far employed carbon-11 and fluorine-18 derivatives of reboxetine but due to defluorination of the fluorine-18 derivatives further research is still necessary. PMID:20216936

  18. Population genetic study of the human dopamine transporter gene (DAT1).

    PubMed

    Doucette-Stamm, L A; Blakely, D J; Tian, J; Mockus, S; Mao, J I

    1995-01-01

    The human dopamine transporter gene, DAT1, acts to transport released dopamine into presynaptic terminals of the brain. The possibility that the DAT1 gene plays a role in genetic diseases of the brain has led to studies of DAT1 in several psychiatric and neurological disorders. Previous sequence analysis of DAT1 revealed a 40-bp repeat in the 3' end of the gene. In order to identify all potential alleles for this VNTR marker a population database was established. One thousand seventy-four unrelated individuals were screened by PCR for the region containing the 40 bp repeat. Allele frequency differences were found between black Americans and Caucasians or Hispanics but no differences were observed between Caucasians and Hispanics. A previously unreported allele was detected in all three populations. Thus, we have shown that screening a large population identifies new alleles and generates more accurate allele frequencies. PMID:7557351

  19. Reduced striatal dopamine transporter density associated with working memory deficits in opioid-dependent male subjects: a SPECT study.

    PubMed

    Liang, Chih-Sung; Ho, Pei-Shen; Yen, Che-Hung; Yeh, Yi-Wei; Kuo, Shin-Chang; Huang, Chang-Chih; Chen, Chun-Yen; Shih, Mei-Chen; Ma, Kuo-Hsing; Huang, San-Yuan

    2016-01-01

    Research on the effects of repeated opioid use on striatal dopamine transporters has yielded inconsistent results, possibly confounded by a history of methamphetamine or methadone exposure in opioid-dependent individuals. Previous studies have shown that striatal dopamine transporter density is positively correlated with the cognitive performance of healthy volunteers. This study aimed to investigate changes in striatal dopamine transporter density and their functional significance in opioid-dependent individuals. Single-photon emission computed tomography with [(99m) Tc]TRODAT-1 as a ligand was used to measure striatal dopamine transporter levels in 20 opioid-dependent individuals and 20 age- and sex-matched healthy controls. Opioid-dependent individuals had no history of methamphetamine or methadone use. The Wisconsin Card Sorting Test (WCST) was performed to assess neurocognitive function. We found that compared with healthy controls, opioid-dependent individuals showed a significant reduction in striatal dopamine transporter density. They also showed poorer performance on the WCST in terms of the trials administered, total errors, perseverative responses, perseverative errors, and non-perseverative errors. Striatal dopamine transporter levels negatively correlated with non-perseverative errors not only in opioid-dependent individuals but also in healthy controls. These findings suggest that in human, repeated opioid exposure reduces striatal dopamine transporter density, which can be associated with non-perseverative errors. Non-perseverative errors may be one of the more sensitive parameters in WCST to identify working memory deficits associated with striatal dopamine transporter reduction. Moreover, we suggest that whether opioid-associated neurotoxicity is reversible depends on the brain region. PMID:25439653

  20. Serotonin and Dopamine Transporter Binding in Children with Autism Determined by SPECT

    ERIC Educational Resources Information Center

    Makkonen, Ismo; Riikonen, Raili; Kokki, Hannu; Airaksinen, Mauno M.; Kuikka, Jyrki T.

    2008-01-01

    Disturbances in the serotonergic system have been recognized in autism. To investigate the association between serotonin and dopamine transporters and autism, we studied 15 children (14 males, one female; mean age 8y 8mo [SD 3y 10mo]) with autism and 10 non-autistic comparison children (five males, five females; mean age 9y 10mo [SD 2y 8mo]) using…

  1. Preserved serotonin transporter binding in de novo Parkinson's disease: negative correlation with the dopamine transporter.

    PubMed

    Strecker, Karl; Wegner, Florian; Hesse, Swen; Becker, Georg-Alexander; Patt, Marianne; Meyer, Philipp M; Lobsien, Donald; Schwarz, Johannes; Sabri, Osama

    2011-01-01

    Recent imaging and neuropathological studies indicate reduced serotonin transporter (SERT) in advanced Parkinson's disease (PD). However, data on SERT in early PD patients are sparse. Following the hypothesis that the serotonergic system is damaged early in PD, the aim of our study was to investigate SERT availability by means of PET imaging. Since the loss of dopaminergic neurons is the pathologic hallmark of PD and SERT might be associated with psychiatric co-morbidity, we further sought to correlate SERT availability with the availability of dopamine transporter (DAT) and depressive or motor symptoms in early PD. We prospectively recruited nine early PD patients (4 female, 5 male; 42-76 years) and nine age matched healthy volunteers (5 female, 4 male; 42-72 years). Diagnosis of PD was confirmed by the UK brain bank criteria and DAT imaging. SERT availability was measured by means of [11C]DASB PET. For neuropsychiatric assessment done on the day of PET we applied UPDRS parts I, II and III, Beck's Depression Inventory, Hamilton Rating Scale for Depression, Mini-Mental State Examination and Demtect. SERT was not reduced in any of 14 investigated regions of interest in the nine PD patients compared to healthy controls (p>0.13). SERT was negatively associated with DAT in the striatum (r=-0.69; p=0.04) but not within the midbrain. There was no correlation of SERT availability with depressive symptoms. No alteration of SERT binding in our patients suggests that the serotonergic system is remarkably preserved in early PD. Correlation with DAT might point to a compensatory regulation of the serotonergic system in early stages of PD. PMID:20644949

  2. Nicotine-dopamine-transporter interactions during reward-based decision making.

    PubMed

    Kambeitz, Joseph; la Fougère, Christian; Werner, Natalie; Pogarell, Oliver; Riedel, Michael; Falkai, Peter; Ettinger, Ulrich

    2016-06-01

    Our everyday-life comprises a multitude of decisions that we take whilst trying to maximize advantageous outcomes, limit risks and update current needs. The cognitive processes that guide decision making as well as the brain circuits they are based on are only poorly understood. Numerous studies point to a potential role of dopamine and nicotine in decision making but less is known about their interactions. Here, 26 healthy male subjects performed the Iowa Gambling Task (IGT) in two sessions following the administration of either nicotine or placebo. Striatal dopamine transporter (DAT) binding was measured by single-photon emission computed tomography (SPECT). Results indicate that lower DAT levels were associated with better performance in the IGT (p=0.0004). Cognitive modelling analysis using the prospect valence learning (PVL) model indicated that low DAT subjects' performance deteriorated following nicotine administration as indicated by an increased learning rate and a decreased response consistency. Our results shed light on the neurochemistry underlying reward-based decision making in humans by demonstrating a significant interaction between nicotine and the DAT. The observed interaction is consistent with the hypothesized associations between DAT expression and extracellular dopamine levels, suggestive of an inverted U-shape relationship between baseline dopamine and magnitude in response to a pro-dopaminergic compound. Our findings are of particular interest in the context of psychiatric disorders where aberrant decision making represents a part of the core symptomatology, such as addiction, schizophrenia or depression. PMID:27112968

  3. Brain dopamine-serotonin vesicular transport disease presenting as a severe infantile hypotonic parkinsonian disorder.

    PubMed

    Jacobsen, Jessie C; Wilson, Callum; Cunningham, Vicki; Glamuzina, Emma; Prosser, Debra O; Love, Donald R; Burgess, Trent; Taylor, Juliet; Swan, Brendan; Hill, Rosamund; Robertson, Stephen P; Snell, Russell G; Lehnert, Klaus

    2016-03-01

    Two male siblings from a consanguineous union presented in early infancy with marked truncal hypotonia, a general paucity of movement, extrapyramidal signs and cognitive delay. By mid-childhood they had made little developmental progress and remained severely hypotonic and bradykinetic. They developed epilepsy and had problems with autonomic dysfunction and oculogyric crises. They had a number of orthopaedic problems secondary to their hypotonia. Cerebrospinal fluid (CSF) neurotransmitters were initially normal, apart from mildly elevated 5-hydroxyindolacetic acid, and the children did not respond favourably to a trial of levodopa-carbidopa. The youngest died from respiratory complications at 10 years of age. Repeat CSF neurotransmitters in the older sibling at eight years of age showed slightly low homovanillic acid and 5-hydroxyindoleacetic acid levels. Whole-exome sequencing revealed a novel mutation homozygous in both children in the monoamine transporter gene SLC18A2 (p.Pro237His), resulting in brain dopamine-serotonin vesicular transport disease. This is the second family to be described with a mutation in this gene. Treatment with the dopamine agonist pramipexole in the surviving child resulted in mild improvements in alertness, communication, and eye movements. This case supports the identification of the causal mutation in the original case, expands the clinical phenotype of brain dopamine-serotonin vesicular transport disease and confirms that pramipexole treatment may lead to symptomatic improvement in affected individuals. PMID:26497564

  4. Enhanced Dopamine Release by Dopamine Transport Inhibitors Described by a Restricted Diffusion Model and Fast-Scan Cyclic Voltammetry.

    PubMed

    Hoffman, Alexander F; Spivak, Charles E; Lupica, Carl R

    2016-06-15

    Fast-scan cyclic voltammetry (FSCV) using carbon fiber electrodes is widely used to rapidly monitor changes in dopamine (DA) levels in vitro and in vivo. Current analytical approaches utilize parameters such as peak oxidation current amplitude and decay times to estimate release and uptake processes, respectively. However, peak amplitude changes are often observed with uptake inhibitors, thereby confounding the interpretation of these parameters. To overcome this limitation, we demonstrate that a simple five-parameter, two-compartment model mathematically describes DA signals as a balance of release (r/ke) and uptake (ku), summed with adsorption (kads and kdes) of DA to the carbon electrode surface. Using nonlinear regression, we demonstrate that our model precisely describes measured DA signals obtained in brain slice recordings. The parameters extracted from these curves were then validated using pharmacological manipulations that selectively alter vesicular release or DA transporter (DAT)-mediated uptake. Manipulation of DA release through altering the Ca(2+)/Mg(2+) ratio or adding tetrodotoxin reduced the release parameter with no effect on the uptake parameter. DAT inhibitors methylenedioxypyrovalerone, cocaine, and nomifensine significantly reduced uptake and increased vesicular DA release. In contrast, a low concentration of amphetamine reduced uptake but had no effect on DA release. Finally, the kappa opioid receptor agonist U50,488 significantly reduced vesicular DA release but had no effect on uptake. Together, these data demonstrate a novel analytical approach to distinguish the effects of manipulations on DA release or uptake that can be used to interpret FSCV data. PMID:27018734

  5. Dissociable effects of the prodrug phendimetrazine and its metabolite phenmetrazine at dopamine transporters

    PubMed Central

    Solis, Ernesto; Suyama, Julie A.; Lazenka, Matthew F.; DeFelice, Louis J.; Negus, S. Stevens; Blough, Bruce E.; Banks, Matthew L.

    2016-01-01

    Phendimetrazine (PDM) is a clinically available anorectic and a candidate pharmacotherapy for cocaine addiction. PDM has been hypothesized to function as a prodrug that requires metabolism to the amphetamine-like monoamine transporter substrate phenmetrazine (PM) to produce its pharmacological effects; however, whether PDM functions as an inactive prodrug or has pharmacological activity on its own remains unclear. The study aim was to determine PDM pharmacological mechanisms using electrophysiological, neurochemical, and behavioral procedures. PDM blocked the endogenous basal hDAT (human dopamine transporter) current in voltage-clamped (−60 mV) oocytes consistent with a DAT inhibitor profile, whereas its metabolite PM induced an inward hDAT current consistent with a DAT substrate profile. PDM also attenuated the PM-induced inward current during co-application, providing further evidence that PDM functions as a DAT inhibitor. PDM increased nucleus accumbens dopamine levels and facilitated electrical brain stimulation reinforcement within 10 min in rats, providing in vivo evidence supporting PDM pharmacological activity. These results demonstrate that PDM functions as a DAT inhibitor that may also interact with the pharmacological effects of its metabolite PM. Overall, these results suggest a novel mechanism for PDM therapeutic effects via initial PDM DAT inhibition followed by PM DAT substrate-induced dopamine release. PMID:27514281

  6. Dissociable effects of the prodrug phendimetrazine and its metabolite phenmetrazine at dopamine transporters.

    PubMed

    Solis, Ernesto; Suyama, Julie A; Lazenka, Matthew F; DeFelice, Louis J; Negus, S Stevens; Blough, Bruce E; Banks, Matthew L

    2016-01-01

    Phendimetrazine (PDM) is a clinically available anorectic and a candidate pharmacotherapy for cocaine addiction. PDM has been hypothesized to function as a prodrug that requires metabolism to the amphetamine-like monoamine transporter substrate phenmetrazine (PM) to produce its pharmacological effects; however, whether PDM functions as an inactive prodrug or has pharmacological activity on its own remains unclear. The study aim was to determine PDM pharmacological mechanisms using electrophysiological, neurochemical, and behavioral procedures. PDM blocked the endogenous basal hDAT (human dopamine transporter) current in voltage-clamped (-60 mV) oocytes consistent with a DAT inhibitor profile, whereas its metabolite PM induced an inward hDAT current consistent with a DAT substrate profile. PDM also attenuated the PM-induced inward current during co-application, providing further evidence that PDM functions as a DAT inhibitor. PDM increased nucleus accumbens dopamine levels and facilitated electrical brain stimulation reinforcement within 10 min in rats, providing in vivo evidence supporting PDM pharmacological activity. These results demonstrate that PDM functions as a DAT inhibitor that may also interact with the pharmacological effects of its metabolite PM. Overall, these results suggest a novel mechanism for PDM therapeutic effects via initial PDM DAT inhibition followed by PM DAT substrate-induced dopamine release. PMID:27514281

  7. Mice expressing markedly reduced striatal dopamine transporters exhibit increased locomotor activity, dopamine uptake turnover rate, and cocaine responsiveness.

    PubMed

    Rao, Anjali; Sorkin, Alexander; Zahniser, Nancy R

    2013-10-01

    Variations in the expression levels of the dopamine transporter (DAT) can influence responsiveness to psychostimulant drugs like cocaine. To better understand this relationship, we studied a new DAT-low expresser (DAT-LE) mouse model and performed behavioral and biochemical studies with it. Immunoblotting and [(3) H]WIN 35,428 binding analyses revealed that these mice express ∼35% of wildtype (WT) mouse striatal DAT levels. Compared to WT mice, DAT-LE mice were hyperactive in a novel open-field environment. Despite their higher basal locomotor activity, cocaine (10 or 20 mg/kg, i.p.) induced greater locomotor activation in DAT-LE mice than in WT mice. The maximal velocity (Vmax ) of DAT-mediated [(3) H]DA uptake into striatal synaptosomes was reduced by 46% in DAT-LE mice, as compared to WT. Overall, considering the reduced number of DAT binding sites (Bmax ) along with the reduced Vmax in DAT-LE mice, a 2-fold increase in DA uptake turnover rate (Vmax /Bmax ) was found, relative to WT mice. This suggests that neuroadaptive changes have occurred in the DAT-LE mice that would help to compensate for their low DAT numbers. Interestingly, these changes do not include a reduction in tyrosine hydroxylase levels, as was previously reported in DAT knockout homozygous and heterozygous animals. Further, these changes are not sufficient to prevent elevated novelty- and cocaine-induced locomotor activity. Hence, these mice represent a unique model for studying changes of in vivo DAT function and regulation that result from markedly reduced levels of DAT expression. PMID:23564231

  8. A behavioral defect of temporal association memory in mice that partly lack dopamine reuptake transporter

    PubMed Central

    Deng, Shining; Zhang, Lingli; Zhu, Tailin; Liu, Yan-Mei; Zhang, Hailong; Shen, Yiping; Li, Wei-Guang; Li, Fei

    2015-01-01

    Temporal association memory, like working memory, is a type of episodic memory in which temporally discontinuous elements are associated. However, the mechanisms that govern this association remain incompletely understood. Here, we identify a crucial role of dopaminergic action in temporal association memory. We used hemizygote hyperdopaminergic mutant mice with reduced dopamine transporter (DAT) expression, referred to as DAT+/− mice. We found that mice with this modest dopamine imbalance exhibited significantly impaired trace fear conditioning, which necessitates the association of temporally discontinuous elements, and intact delay auditory fear conditioning, which does not. Moreover, the DAT+/− mice displayed substantial impairments in non-matching-to-place spatial working-memory tasks. Interestingly, these temporal association and working memory deficits could be mimicked by a low dose of the dopamine D2 receptor antagonist haloperidol. The shared phenotypes resulting from either the genetic reduction of DAT or the pharmacological inhibition of the D2 receptor collectively indicate that temporal association memory necessitates precise regulation of dopaminergic signaling. The particular defect in temporal association memory due to partial lack of DAT provides mechanistic insights on the understanding of cognitive impairments in multiple neurodevelopmental disorders. PMID:26658842

  9. Essential Oils from the Medicinal Herbs Upregulate Dopamine Transporter in Rat Pheochromocytoma Cells.

    PubMed

    Choi, Min Sun; Choi, Bang-sub; Kim, Sang Heon; Pak, Sok Cheon; Jang, Chul Ho; Chin, Young-Won; Kim, Young-Mi; Kim, Dong-il; Jeon, Songhee; Koo, Byung-Soo

    2015-10-01

    The dopamine transporter (DAT) protein, a component of the dopamine system, undergoes adaptive neurobiological changes from drug abuse. Prevention of relapse and reduction of withdrawal symptoms are still the major limitations in the current pharmacological treatments of drug addiction. The present study aimed to investigate the effects of essential oils extracted from Elsholtzia ciliata, Shinchim, Angelicae gigantis Radix, and Eugenia caryophyllata, well-known traditional Korean medicines for addiction, on the modulation of dopamine system in amphetamine-treated cells and to explore the possible mechanism underlying its therapeutic effect. The potential cytotoxic effect of essential oils was evaluated in PC12 rat pheochromocytoma cells using cell viability assays. Quantification of DAT, p-CREB, p-MAPK, and p-Akt was done by immunoblotting. DAT was significantly reduced in cells treated with 50 μM of amphetamine in a time-dependent manner. No significant toxicity of essential oils from Elsholtzia ciliata and Shinchim was observed at doses of 10, 25, and 50 μg/mL. However, essential oils from A. gigantis Radix at a dose of 100 μg/mL and E. caryophyllata at doses of 50 and 100 μg/mL showed cytotoxicity. Treatment with GBR 12909, a highly selective DAT inhibitor, significantly increased DAT expression compared with that of amphetamine only by enhancing phosphorylation of mitogen-activated protein kinases (MAPK) and Akt. In addition, essential oils effectively induced hyperphosphorylation of cyclic-AMP response element-binding protein (CREB), MAPK, and Akt, which resulted in DAT upregulation. Our study implies that the essential oils may rehabilitate brain dopamine function through increased DAT availability in abstinent former drug users. PMID:26295793

  10. Dopamine transporter binding in social anxiety disorder: the effect of treatment with escitalopram.

    PubMed

    Warwick, J M; Carey, P D; Cassimjee, N; Lochner, C; Hemmings, S; Moolman-Smook, H; Beetge, E; Dupont, P; Stein, D J

    2012-06-01

    Social anxiety disorder (SAD) is characterised by fear of social or performance situations where the individual is exposed to unfamiliar people or to possible scrutiny by others. The literature on dopamine ligands and dopamine genotypes in SAD is however inconsistent. In this study we measured the effects of SSRI pharmacotherapy on dopamine transporter (DAT) binding in patients with SAD, also addressing variability in DAT genotype. Adult subjects meeting DSM-IV criteria for generalised SAD were studied before and after 12 weeks of pharmacotherapy with the selective serotonin reuptake inhibitor (SSRI) escitalopram. DAT single photon emission computed tomography (SPECT) using (123)I-FP-CIT was performed at baseline, and repeated at 12 weeks. Striatal DAT binding was analysed for changes following therapy, and for correlations with clinical efficacy, in the whole group as well as for a subgroup with the A10/A10 DAT genotype. The study included 14 subjects (9 male, 5 female) with a mean (SD) age of 41 (±13) years. The subjects' Liebowitz Social Anxiety Scale (LSAS) score was significantly decreased following pharmacotherapy. In the combined group the left caudate and left putamen showed clusters of increased DAT binding after therapy. The left caudate changes were also observed in the subgroup of 9 A10/A10 homozygotes. However no correlation was found between improved symptoms and DAT binding. The changes found in DAT binding in the caudate and putamen may be due to serotonergic activation of dopamine function by SSRI therapy. This is consistent with previous work indicating decreased DAT binding in SAD, and increased DAT binding after SSRI administration. PMID:22350963

  11. N-terminal tagging of the dopamine transporter impairs protein expression and trafficking in vivo

    PubMed Central

    Vecchio, Laura M.; Bermejo, M. Kristel; Beerepoot, Pieter; Ramsey, Amy J.

    2014-01-01

    The dopamine transporter (DAT) is the primary protein responsible for the uptake of dopamine from the extracellular space back into presynaptic neurons. As such, it plays an important role in the cessation of dopaminergic neurotransmission and in the maintenance of extracellular dopamine homeostasis. Here, we report the development of a new BAC transgenic mouse line that expresses DAT with an N-terminal HA-epitope (HAD-Tg). In this line, two copies of the HA-DAT BAC are incorporated into the genome, increasing DAT mRNA levels by 47%. Despite the increase in mRNA levels, HAD-Tg mice show no significant increase in the level of DAT protein in the striatum, indicating a defect in protein trafficking or stability. By crossing HAD-Tg mice with DAT knockout mice (DAT-KO), we engineered mice that exclusively express HA-tagged DAT in the absence of endogenous DAT (DAT-KO/HAD-Tg). We show that DAT-KO/HAD-Tg mice express only 8.5% of WT DAT levels in the striatum. Importantly, the HA-tagged DAT that is present in DAT-KO/HAD-Tg mice is functional, as it is able to partially rescue the DAT-KO hyperactive phenotype. Finally, we provide evidence that the HA-tagged DAT is retained in the cell body based on a reduction in the striatum:midbrain protein ratio. These results demonstrate that the presence of the N-terminal tag leads to impaired DAT protein expression in vivo due in part to improper trafficking of the tagged transporter, and highlight the importance of the N-terminus in the transport of DAT to striatal terminals. PMID:24886986

  12. Electropolymerized molecular imprinting on glassy carbon electrode for voltammetric detection of dopamine in biological samples.

    PubMed

    Kiss, Laszlo; David, Vasile; David, Iulia Gabriela; Lazăr, Paul; Mihailciuc, Constantin; Stamatin, Ioan; Ciobanu, Adela; Ştefănescu, Cristian Dragoş; Nagy, Livia; Nagy, Géza; Ciucu, Anton Alexandru

    2016-11-01

    A simple and reliable method for preparing a selective dopamine (DA) sensor based on a molecularly imprinted polymer of ethacridine was proposed. The molecularly imprinted polymer electrode was prepared through electrodepositing polyethacridine-dopamine film on the glassy carbon electrode and then removing DA from the film via chemical induced elution. The molecular imprinted sensor was tested by cyclic voltammetry as well as by differential pulse voltammetry (DPV) to verify the changes in oxidative currents of DA. In optimized DPV conditions the oxidation peak current was well-proportional to the concentration of DA in the range from 2.0×10(-8)M up to 1×10(-6)M. The limit of detection (3σ) of DA was found to be as low as 4.4nM, by the proposed sensor that could be considered a sensitive marker of DA depletion in Parkinson's disease. Good reproducibility with relative standard deviation of 1.4% and long term stability within two weeks were also observed. The modified sensor was validated for the analysis of DA in deproteinized human serum samples using differential pulse voltammetric technique. PMID:27591643

  13. Synthesis and dopamine transporter imaging in rhesus monkeys with fluorine-18 labeled FECT

    SciTech Connect

    Keil, R.; Hoffman, J.M.; Eschima, D.

    1996-05-01

    Parkinson`s patients have been shown to suffer a 60-80% loss of dopamine transporters in the substantia nigra and striatum. Dopamine transporter ligands labeled with fluorine-18 (t {1/2}=110 min) are attractive probes for measuring the density of dopamine transporter sites n the striatum for the diagnosis and evaluation of Parkinson`s patients by PET. We have synthesized (Ki = 32 nM vs RTI-55), fluorine-18 labeled 2{beta}-carbomethoxy-3{beta}(4-chlorophenyl)-8-(3-fluoropropyl)nortropane (FECT), with favorable kinetics as a potential dopamine transporter PET imaging agent. Treatment of 2{beta}-carbomethoxy-3{beta}-(4-chlorophenyl)nortropane (1) with 1-bromo-2-fluoroethane (2) in CH3CN at 80{degrees}C gave FECT (3). [F-18]FECT (3) was prepared by treating 1,2-ditosyloxyethane (4) with NCA K[F-18]/K222 (365 mCi) for 5 min in CH3CN at 85{degrees}C to give [F-18] 1-fluoro-2-tosyloxyethane (5) (175 mCi)in 59% E.O.B. yield. Coupling of [F-18] 5 with 1 in DMF at 135 {degrees}C for 45 min gave [F-18]FECT (41 mCi) in 25% yield E.O.B. following HPLC purification in a total synthesis time of 122 min. [F-18] 5 was >99% radiochemically pure with a specific activity of 5 Ci/{mu}mole. Following intravenous administration to a rhesus monkey [F-18]FECT (8.13 mCi) showed a peak uptake at 30 min in the striatum (S) followed by a slow clearance and a rapid washout from the cerebellum to afford a high S/C ratio = 11.0 at 125 min. Radio-HPLC analysis of the ether extracts form plasma samples for radioactive metabolites detected only the presence of [F-18]FECT. These results suggest that FECT is an Research supported by DOE.

  14. A novel photoaffinity ligand for the dopamine transporter based on pyrovalerone

    PubMed Central

    Lapinsky, David J.; Aggarwal, Shaili; Huang, Yurong; Surratt, Christopher K.; Lever, John R.; Foster, James D.; Vaughan, Roxanne A.

    2009-01-01

    Non-tropane-based photoaffinity ligands for the dopamine transporter (DAT) are relatively unexplored in contrast to tropane-based compounds such as cocaine. In order to fill this knowledge gap, a ligand was synthesized in which the aromatic ring of pyrovalerone was substituted with a photoreactive azido group. The analog 1-(4-azido-3-iodophenyl)-2-pyrrolidin-1-yl-pentan-1-one demonstrated appreciable binding affinity for the DAT (Ki = 78 ± 18 nM), suggesting the potential utility of a radioiodinated version in structure-function studies of this protein. PMID:19442525

  15. Polymorphism of the dopamine transporter type 1 gene modifies the treatment response in Parkinson's disease.

    PubMed

    Moreau, Caroline; Meguig, Sayah; Corvol, Jean-Christophe; Labreuche, Julien; Vasseur, Francis; Duhamel, Alain; Delval, Arnaud; Bardyn, Thomas; Devedjian, Jean-Christophe; Rouaix, Nathalie; Petyt, Gregory; Brefel-Courbon, Christine; Ory-Magne, Fabienne; Guehl, Dominique; Eusebio, Alexandre; Fraix, Valérie; Saulnier, Pierre-Jean; Lagha-Boukbiza, Ouhaid; Durif, Frank; Faighel, Mirela; Giordana, Caroline; Drapier, Sophie; Maltête, David; Tranchant, Christine; Houeto, Jean-Luc; Debû, Bettina; Azulay, Jean-Philippe; Tison, François; Destée, Alain; Vidailhet, Marie; Rascol, Olivier; Dujardin, Kathy; Defebvre, Luc; Bordet, Régis; Sablonnière, Bernard; Devos, David

    2015-05-01

    After more than 50 years of treating Parkinson's disease with l-DOPA, there are still no guidelines on setting the optimal dose for a given patient. The dopamine transporter type 1, now known as solute carrier family 6 (neurotransmitter transporter), member 3 (SLC6A3) is the most powerful determinant of dopamine neurotransmission and might therefore influence the treatment response. We recently demonstrated that methylphenidate (a dopamine transporter inhibitor) is effective in patients with Parkinson's disease with motor and gait disorders. The objective of the present study was to determine whether genetic variants of the dopamine transporter type 1-encoding gene (SLC6A3) are associated with differences in the response to treatment of motor symptoms and gait disorders with l-DOPA and methylphenidate (with respect to the demographic, the disease and the treatment parameters and the other genes involved in the dopaminergic neurotransmission). This analysis was part of a multicentre, parallel-group, double-blind, placebo-controlled, randomized clinical trial of methylphenidate in Parkinson's disease (Protocol ID:2008-005801-20; ClinicalTrials.gov:NCT00914095). We scored the motor Unified Parkinson's Disease Rating Scale and the Stand-Walk-Sit Test before and after a standardized acute l-DOPA challenge before randomization and then after 3 months of methylphenidate treatment. Patients were screened for variants of genes involved in dopamine metabolism: rs28363170 and rs3836790 polymorphisms in the SLC6A3 gene, rs921451 and rs3837091 in the DDC gene (encoding the aromatic L-amino acid decarboxylase involved in the synthesis of dopamine from l-DOPA), rs1799836 in the MAOB gene (coding for monoamine oxidase B) and rs4680 in the COMT gene (coding for catechol-O-methyltransferase). Investigators and patients were blinded to the genotyping data throughout the study. Eighty-one subjects were genotyped and 61 were analysed for their acute motor response to l-DOPA. The SLC6A3

  16. The effect of modafinil on the rat dopamine transporter and dopamine receptors D1-D3 paralleling cognitive enhancement in the radial arm maze.

    PubMed

    Karabacak, Yasemin; Sase, Sunetra; Aher, Yogesh D; Sase, Ajinkya; Saroja, Sivaprakasam R; Cicvaric, Ana; Höger, Harald; Berger, Michael; Bakulev, Vasiliy; Sitte, Harald H; Leban, Johann; Monje, Francisco J; Lubec, Gert

    2015-01-01

    A series of drugs have been reported to increase memory performance modulating the dopaminergic system and herein modafinil was tested for its working memory (WM) enhancing properties. Reuptake inhibition of dopamine, serotonin (SERT) and norepinephrine (NET) by modafinil was tested. Sixty male Sprague-Dawley rats were divided into six groups (modafinil-treated 1-5-10 mg/kg body weight, trained and untrained and vehicle treated trained and untrained rats; daily injected intraperitoneally for a period of 10 days) and tested in a radial arm maze (RAM), a paradigm for testing spatial WM. Hippocampi were taken 6 h following the last day of training and complexes containing the unphosphorylated or phosphorylated dopamine transporter (DAT-CC and pDAT-CC) and complexes containing the D1-3 dopamine receptor subunits (D1-D3-CC) were determined. Modafinil was binding to the DAT but insignificantly to SERT or NET and dopamine reuptake was blocked specifically (IC50 = 11.11 μM; SERT 1547 μM; NET 182 μM). From day 8 (day 9 for 1 mg/kg body weight) modafinil was decreasing WM errors (WMEs) in the RAM significantly and remarkably at all doses tested as compared to the vehicle controls. WMEs were linked to the D2R-CC and the pDAT-CC. pDAT and D1-D3-CC levels were modulated significantly and modafinil was shown to enhance spatial WM in the rat in a well-documented paradigm at all the three doses and dopamine reuptake inhibition with subsequent modulation of D1-3-CC is proposed as a possible mechanism of action. PMID:26347626

  17. The effect of modafinil on the rat dopamine transporter and dopamine receptors D1–D3 paralleling cognitive enhancement in the radial arm maze

    PubMed Central

    Karabacak, Yasemin; Sase, Sunetra; Aher, Yogesh D.; Sase, Ajinkya; Saroja, Sivaprakasam R.; Cicvaric, Ana; Höger, Harald; Berger, Michael; Bakulev, Vasiliy; Sitte, Harald H.; Leban, Johann; Monje, Francisco J.; Lubec, Gert

    2015-01-01

    A series of drugs have been reported to increase memory performance modulating the dopaminergic system and herein modafinil was tested for its working memory (WM) enhancing properties. Reuptake inhibition of dopamine, serotonin (SERT) and norepinephrine (NET) by modafinil was tested. Sixty male Sprague–Dawley rats were divided into six groups (modafinil-treated 1–5–10 mg/kg body weight, trained and untrained and vehicle treated trained and untrained rats; daily injected intraperitoneally for a period of 10 days) and tested in a radial arm maze (RAM), a paradigm for testing spatial WM. Hippocampi were taken 6 h following the last day of training and complexes containing the unphosphorylated or phosphorylated dopamine transporter (DAT-CC and pDAT-CC) and complexes containing the D1–3 dopamine receptor subunits (D1–D3-CC) were determined. Modafinil was binding to the DAT but insignificantly to SERT or NET and dopamine reuptake was blocked specifically (IC50 = 11.11 μM; SERT 1547 μM; NET 182 μM). From day 8 (day 9 for 1 mg/kg body weight) modafinil was decreasing WM errors (WMEs) in the RAM significantly and remarkably at all doses tested as compared to the vehicle controls. WMEs were linked to the D2R-CC and the pDAT-CC. pDAT and D1–D3-CC levels were modulated significantly and modafinil was shown to enhance spatial WM in the rat in a well-documented paradigm at all the three doses and dopamine reuptake inhibition with subsequent modulation of D1–3-CC is proposed as a possible mechanism of action. PMID:26347626

  18. The mechanistic basis for noncompetitive ibogaine inhibition of serotonin and dopamine transporters.

    PubMed

    Bulling, Simon; Schicker, Klaus; Zhang, Yuan-Wei; Steinkellner, Thomas; Stockner, Thomas; Gruber, Christian W; Boehm, Stefan; Freissmuth, Michael; Rudnick, Gary; Sitte, Harald H; Sandtner, Walter

    2012-05-25

    Ibogaine, a hallucinogenic alkaloid proposed as a treatment for opiate withdrawal, has been shown to inhibit serotonin transporter (SERT) noncompetitively, in contrast to all other known inhibitors, which are competitive with substrate. Ibogaine binding to SERT increases accessibility in the permeation pathway connecting the substrate-binding site with the cytoplasm. Because of the structural similarity between ibogaine and serotonin, it had been suggested that ibogaine binds to the substrate site of SERT. The results presented here show that ibogaine binds to a distinct site, accessible from the cell exterior, to inhibit both serotonin transport and serotonin-induced ionic currents. Ibogaine noncompetitively inhibited transport by both SERT and the homologous dopamine transporter (DAT). Ibogaine blocked substrate-induced currents also in DAT and increased accessibility of the DAT cytoplasmic permeation pathway. When present on the cell exterior, ibogaine inhibited SERT substrate-induced currents, but not when it was introduced into the cytoplasm through the patch electrode. Similar to noncompetitive transport inhibition, the current block was not reversed by increasing substrate concentration. The kinetics of inhibitor binding and dissociation, as determined by their effect on SERT currents, indicated that ibogaine does not inhibit by forming a long-lived complex with SERT, but rather binds directly to the transporter in an inward-open conformation. A kinetic model for transport describing the noncompetitive action of ibogaine and the competitive action of cocaine accounts well for the results of the present study. PMID:22451652

  19. A surface acoustic wave sensor functionalized with a polypyrrole molecularly imprinted polymer for selective dopamine detection.

    PubMed

    Maouche, Naima; Ktari, Nadia; Bakas, Idriss; Fourati, Najla; Zerrouki, Chouki; Seydou, Mahamadou; Maurel, François; Chehimi, Mohammed Mehdi

    2015-11-01

    A surface acoustic wave sensor operating at 104 MHz and functionalized with a polypyrrole molecularly imprinted polymer has been designed for selective detection of dopamine (DA). Optimization of pyrrole/DA ratio, polymerization and immersion times permitted to obtain a highly selective sensor, which has a sensitivity of 0.55°/mM (≈ 550 Hz/mM) and a detection limit of ≈ 10 nM. Morphology and related roughness parameters of molecularly imprinted polymer surfaces, before and after extraction of DA, as well as that of the non imprinted polymer were characterized by atomic force microscopy. The developed chemosensor selectively recognized dopamine over the structurally similar compound 4-hydroxyphenethylamine (referred as tyramine), or ascorbic acid,which co-exists with DA in body fluids at a much higher concentration. Selectivity tests were also carried out with dihydroxybenzene, for which an unexpected phase variation of order of 75% of the DA one was observed. Quantum chemical calculations, based on the density functional theory, were carried out to determine the nature of interactions between each analyte and the PPy matrix and the DA imprinted PPy polypyrrole sensing layer in order to account for the important phase variation observed during dihydroxybenzene injection. PMID:26095144

  20. Recognition Properties and Competitive Assays of a Dual Dopamine/Serotonin Selective Molecularly Imprinted Polymer

    PubMed Central

    Suedee, Roongnapa; Seechamnanturakit, Vatcharee; Suksuwan, Acharee; Canyuk, Bhutorn

    2008-01-01

    A molecularly imprinted polymer (MIP) with dual dopamine/serotonin-like binding sites (DS-MIP) was synthesized for use as a receptor model of study the drug-interaction of biological mixed receptors at a molecular level. The polymer material was produced using methacrylic acid (MAA) and acrylamide (ACM) as functional monomers, N,N′-methylene bisacrylamide (MBAA) as cross-linker, methanol/water mixture (4:1, v/v) as porogen and a mixture of dopamine (D) and serotonin (S) as templates. The prepared DS-MIP exhibited the greatest rebinding of the template(s) in aqueous methanol solution with decreased recognition in acetonitrile, water and methanol solvent. The binding affinity and binding capacity of DS-MIP with S were found to be higher than those of DS-MIP with D. The selectivity profiles of DS-MIP suggest that the D binding site of DS-MIP has sufficient integrity to discriminate between species of non-optimal functional group orientation, whilst the S binding site of DS-MIP is less selective toward species having structural features and functional group orientations different from S. The ligand binding activities of a series of ergot derivatives (ergocryptine, ergocornine, ergocristine, ergonovine, agroclavine, pergolide and terguride) have been studied with the DS-MIP using a competitive ligand binding assay protocol. The binding affinities of DS-MIP were demonstrated in the micro- or submicro-molar range for a series of ergot derivatives, whereas the binding affinities were considerably greater to natural receptors derived from the rat hypothalamus. The DS-MIP afforded the same pattern of differentiation as the natural receptors, i.e. affinity for the clavines > lysergic acid derivatives > ergopeptines. The results suggest that the discrimination for the ergot derivatives by the dopamine and serotonin sites of DS-MIP is due to the structural features and functional orientation of the phenylethylamine and indolylethylamine entities at the binding sites, and the

  1. Antagonist-Induced Conformational Changes in Dopamine Transporter Extracellular Loop Two Involve Residues in a Potential Salt Bridge

    PubMed Central

    Gaffaney, Jon D.; Shetty, Madhur; Felts, Bruce; Pramod, Akula-Bala; Foster, James D.; Henry, L. Keith; Vaughan, Roxanne A.

    2014-01-01

    Ligand-induced changes in the conformation of extracellular loop (EL) 2 in the rat (r) dopamine transporter (DAT) were examined using limited proteolysis with endoproteinase Asp-N and detection of cleavage products by epitope-specific immunoblotting. The principle N-terminal fragment produced by Asp-N was a 19 kDa peptide likely derived by proteolysis of EL2 residue D174, which is present just past the extracellular end of TM3. Production of this fragment was significantly decreased by binding of cocaine and other uptake blockers, but was not affected by substrates or Zn2+, indicating the presence of a conformational change at D174 that may be related to the mechanism of transport inhibition. DA transport activity and cocaine analog binding were decreased by Asp-N treatment, suggesting a requirement for EL2 integrity in these DAT functions. In a previous study we demonstrated that ligand-induced protease resistance also occurred at R218 on the C-terminal side of rDAT EL2. Here using substituted cysteine accessibility analysis of human (h) DAT we confirm cocaine-induced alterations in reactivity of the homologous R219 and identify conformational sensitivity of V221. Focused molecular modeling of D174 and R218 based on currently available Aquifex aeolicus leucine transporter crystal structures places these residues within 2.9 Å of one another, suggesting their proximity as a structural basis for their similar conformational sensitivities and indicating their potential to form a salt bridge. These findings extend our understanding of DAT EL2 and its role in transport and binding functions. PMID:24269640

  2. Antagonist-induced conformational changes in dopamine transporter extracellular loop two involve residues in a potential salt bridge.

    PubMed

    Gaffaney, Jon D; Shetty, Madhur; Felts, Bruce; Pramod, Akula-Bala; Foster, James D; Henry, L Keith; Vaughan, Roxanne A

    2014-07-01

    Ligand-induced changes in the conformation of extracellular loop (EL) 2 in the rat (r) dopamine transporter (DAT) were examined using limited proteolysis with endoproteinase Asp-N and detection of cleavage products by epitope-specific immunoblotting. The principle N-terminal fragment produced by Asp-N was a 19kDa peptide likely derived by proteolysis of EL2 residue D174, which is present just past the extracellular end of TM3. Production of this fragment was significantly decreased by binding of cocaine and other uptake blockers, but was not affected by substrates or Zn(2+), indicating the presence of a conformational change at D174 that may be related to the mechanism of transport inhibition. DA transport activity and cocaine analog binding were decreased by Asp-N treatment, suggesting a requirement for EL2 integrity in these DAT functions. In a previous study we demonstrated that ligand-induced protease resistance also occurred at R218 on the C-terminal side of rDAT EL2. Here using substituted cysteine accessibility analysis of human (h) DAT we confirm cocaine-induced alterations in reactivity of the homologous R219 and identify conformational sensitivity of V221. Focused molecular modeling of D174 and R218 based on currently available Aquifex aeolicus leucine transporter crystal structures places these residues within 2.9Å of one another, suggesting their proximity as a structural basis for their similar conformational sensitivities and indicating their potential to form a salt bridge. These findings extend our understanding of DAT EL2 and its role in transport and binding functions. PMID:24269640

  3. Dopamine-transporter levels drive striatal responses to apomorphine in Parkinson's disease

    PubMed Central

    Passamonti, Luca; Salsone, Maria; Toschi, Nicola; Cerasa, Antonio; Giannelli, Marco; Chiriaco, Carmelina; Cascini, Giuseppe Lucio; Fera, Francesco; Quattrone, Aldo

    2013-01-01

    Dopaminergic therapy in Parkinson's disease (PD) can improve some cognitive functions while worsening others. These opposite effects might reflect different levels of residual dopamine in distinct parts of the striatum, although the underlying mechanisms remain poorly understood. We used functional magnetic resonance imaging (fMRI) to address how apomorphine, a potent dopamine agonist, influences brain activity associated with working memory in PD patients with variable levels of nigrostriatal degeneration, as assessed via dopamine-transporter (DAT) scan. Twelve PD patients underwent two fMRI sessions (Off-, On-apomorphine) and one DAT-scan session. Twelve sex-, age-, and education-matched healthy controls underwent one fMRI session. The core fMRI analyses explored: (1) the main effect of group; (2) the main effect of treatment; and (3) linear and nonlinear interactions between treatment and DAT levels. Relative to controls, PD-Off patients showed greater activations within posterior attentional regions (e.g., precuneus). PD-On versus PD-Off patients displayed reduced left superior frontal gyrus activation and enhanced striatal activation during working-memory task. The relation between DAT levels and striatal responses to apomorphine followed an inverted-U-shaped model (i.e., the apomorphine effect on striatal activity in PD patients with intermediate DAT levels was opposite to that observed in PD patients with higher and lower DAT levels). Previous research in PD demonstrated that the nigrostriatal degeneration (tracked via DAT scan) is associated with inverted-U-shaped rearrangements of postsynaptic D2-receptors sensitivity. Hence, it can be hypothesized that individual differences in DAT levels drove striatal responses to apomorphine via D2-receptor-mediated mechanisms. PMID:23785657

  4. Differential influence of dopamine transport rate on the potencies of cocaine, amphetamine, and methylphenidate.

    PubMed

    Calipari, Erin S; Ferris, Mark J; Siciliano, Cody A; Jones, Sara R

    2015-01-21

    Dopamine transporter (DAT) levels vary across brain regions and individuals, and are altered by drug history and disease states; however, the impact of altered DAT expression on psychostimulant effects in brain has not been systematically explored. Using fast scan cyclic voltammetry, we measured the effects of elevated DAT levels on presynaptic dopamine parameters as well as the uptake inhibition potency of the blockers cocaine and methylphenidate (MPH) and the releaser amphetamine (AMPH) in the nucleus accumbens core. Here we found that increases in DAT levels, resulting from either genetic overexpression or MPH self-administration, caused markedly increased maximal rates of uptake (Vmax) that were positively correlated with the uptake inhibition potency of AMPH and MPH, but not cocaine. AMPH and MPH were particularly sensitive to DAT changes, with a 100% increase in Vmax resulting in a 200% increase in potency. The relationship between Vmax and MPH potency was the same as that for AMPH, but was different from that for cocaine, indicating that MPH more closely resembles a releaser with regard to uptake inhibition. Conversely, the effects of MPH on stimulated dopamine release were similar to those of cocaine, with inverted U-shaped increases in release over a concentration-response curve. This was strikingly different from the release profile of AMPH, which showed only reductions at high concentrations, indicating that MPH is not a pure releaser. These data indicate that although MPH is a DAT blocker, its uptake-inhibitory actions are affected by DAT changes in a similar manner to releasers. Together, these data show that fluctuations in DAT levels alter the potency of releasers and MPH but not blockers and suggest an integral role of the DAT in the addictive potential of AMPH and related compounds. PMID:25474655

  5. Cognitive and olfactory deficits in Machado-Joseph disease: a dopamine transporter study.

    PubMed

    Braga-Neto, Pedro; Felicio, Andre C; Hoexter, Marcelo Q; Pedroso, José Luiz; Dutra, Lívia Almeida; Alessi, Helena; Minett, Thaís; Santos-Galduroz, Ruth F; da Rocha, Antônio José; Garcia, Lucas A L; Bertolucci, Paulo Henrique F; Bressan, Rodrigo A; Barsottini, Orlando Graziani Povoas

    2012-08-01

    Cognitive and olfactory impairments have been demonstrated in patients with Machado-Joseph disease (MJD), and a possible relationship with dopaminergic dysfunction is implicated. However, there is still controversy regarding the pattern of striatal dopaminergic dysfunction in patients with MJD. In this study, we investigated whether these patients had different dopamine transporter (DAT) densities as compared to healthy subjects, and correlated these data with cognitive performance and sense of smell. Twenty-two MJD patients and 20 control subjects were enrolled. The neuropsychological assessment comprised the spatial span, symbol search, picture completion, stroop color word test, trail making test and phonemic verbal fluency test. The 16-item Sniffin' Sticks was used to evaluate odor identification. DAT imaging was performed using the SPECT radioligand [(99m)Tc]-TRODAT-1, alongside with Magnetic Resonance imaging. Patients with MJD showed significantly lower DAT density in the caudate (1.34 ± 0.27 versus 2.02 ± 0.50, p < 0.001), posterior putamen (0.81 ± 0.32 versus 1.32 ± 0.34, p < 0.001) and anterior putamen (1.10 ± 0.31 versus 1.85 ± 0.45, p < 0.001) compared with healthy controls. The putamen/caudate ratio was also significantly lower in patients compared with controls (0.73 ± 0.038 versus 0.85 ± 0.032, p = 0.027). Even though we had only two patients with parkinsonism, we detected striatal dopaminergic deficits in those patients. No significant correlations were detected between DAT density and cognitive performance or Sniffin' Sticks scores. The data suggests that striatal dopamine deficit is not involved in cognitive or sense of smell deficits. This finding raises the possibility of extra-striatal dopamine and other neurotransmitter system involvement or of cerebellum neurodegeneration exerting a direct influence on cognitive and sensorial information processing in MJD. PMID:22575233

  6. Cocaine self-administration produces pharmacodynamic tolerance: differential effects on the potency of dopamine transporter blockers, releasers, and methylphenidate.

    PubMed

    Ferris, Mark J; Calipari, Erin S; Mateo, Yolanda; Melchior, James R; Roberts, David C S; Jones, Sara R

    2012-06-01

    The dopamine transporter (DAT) is the primary site of action for psychostimulant drugs such as cocaine, methylphenidate, and amphetamine. Our previous work demonstrated a reduced ability of cocaine to inhibit the DAT following high-dose cocaine self-administration (SA), corresponding to a reduced ability of cocaine to increase extracellular dopamine. However, this effect had only been demonstrated for cocaine. Thus, the current investigations sought to understand the extent to which cocaine SA (1.5 mg/kg/inf × 40 inf/day × 5 days) altered the ability of different dopamine uptake blockers and releasers to inhibit dopamine uptake, measured using fast-scan cyclic voltammetry in rat brain slices. We demonstrated that, similar to cocaine, the DAT blockers nomifensine and bupropion were less effective at inhibiting dopamine uptake following cocaine SA. The potencies of amphetamine-like dopamine releasers such as 3,4-methylenedioxymethamphetamine, methamphetamine, amphetamine, and phentermine, as well as a non-amphetamine releaser, 4-benzylpiperidine, were all unaffected. Finally, methylphenidate, which blocks dopamine uptake like cocaine while being structurally similar to amphetamine, shared characteristics of both, resembling an uptake blocker at low concentrations and a releaser at high concentrations. Combined, these experiments demonstrate that after high-dose cocaine SA, there is cross-tolerance of the DAT to other uptake blockers, but not releasers. The reduced ability of psychostimulants to inhibit dopamine uptake following cocaine SA appears to be contingent upon their functional interaction with the DAT as a pure blocker or releaser rather than their structural similarity to cocaine. Further, methylphenidate's interaction with the DAT is unique and concentration-dependent. PMID:22395730

  7. Computational modeling of the N-terminus of the human dopamine transporter and its interaction with PIP2 -containing membranes.

    PubMed

    Khelashvili, George; Doktorova, Milka; Sahai, Michelle A; Johner, Niklaus; Shi, Lei; Weinstein, Harel

    2015-05-01

    The dopamine transporter (DAT) is a transmembrane protein belonging to the family of neurotransmitter:sodium symporters (NSS). Members of the NSS are responsible for the clearance of neurotransmitters from the synaptic cleft, and for their translocation back into the presynaptic nerve terminal. The DAT contains long intracellular N- and C-terminal domains that are strongly implicated in the transporter function. The N-terminus (N-term), in particular, regulates the reverse transport (efflux) of the substrate through DAT. Currently, the molecular mechanisms of the efflux remain elusive in large part due to lack of structural information on the N-terminal segment. Here we report a computational model of the N-term of the human DAT (hDAT), obtained through an ab initio structure prediction, in combination with extensive atomistic molecular dynamics (MD) simulations in the context of a lipid membrane. Our analysis reveals that whereas the N-term is a highly dynamic domain, it contains secondary structure elements that remain stable in the long MD trajectories of interactions with the bilayer (totaling >2.2 μs). Combining MD simulations with continuum mean-field modeling we found that the N-term engages with lipid membranes through electrostatic interactions with the charged lipids PIP2 (phosphatidylinositol 4,5-Biphosphate) or PS (phosphatidylserine) that are present in these bilayers. We identify specific motifs along the N-term implicated in such interactions and show that differential modes of N-term/membrane association result in differential positioning of the structured segments on the membrane surface. These results will inform future structure-based studies that will elucidate the mechanistic role of the N-term in DAT function. PMID:25739722

  8. Computational modeling of the N-terminus of the human dopamine transporter and its interaction with PIP2-containing membranes

    PubMed Central

    Khelashvili, George; Doktorova, Milka; Sahai, Michelle A.; Johner, Niklaus; Shi, Lei; Weinstein, Harel

    2015-01-01

    The dopamine transporter (DAT) is a transmembrane protein belonging to the family of Neurotransmitter:Sodium Symporters (NSS). Members of the NSS are responsible for the clearance of neurotransmitters from the synaptic cleft, and for their translocation back into the presynaptic nerve terminal. The DAT contains long intracellular N- and C-terminal domains that are strongly implicated in the transporter function. The N-terminus (N-term), in particular, regulates the reverse transport (efflux) of the substrate through DAT. Currently, the molecular mechanisms of the efflux remain elusive in large part due to lack of structural information on the N-terminal segment. Here we report a computational model of the N-term of the human DAT (hDAT), obtained through an ab initio structure prediction, in combination with extensive atomistic molecular dynamics (MD) simulations in the context of a lipid membrane. Our analysis reveals that whereas the N-term is a highly dynamic domain, it contains secondary structure elements that remain stable in the long MD trajectories of interactions with the bilayer (totaling >2.2 µs). Combining MD simulations with continuum mean-field modeling we found that the N-term engages with lipid membranes through electrostatic interactions with the charged lipids PIP2 (phosphatidylinositol 4,5-Biphosphate) or PS (phosphatidylserine) that are present in these bilayers. We identify specific motifs along the N-term implicated in such interactions and show that differential modes of N-term/membrane association result in differential positioning of the structured segments on the membrane surface. These results will inform future structure-based studies that will elucidate the mechanistic role of the N-term in DAT function. PMID:25739722

  9. Predicting childhood effortful control from interactions between early parenting quality and children's dopamine transporter gene haplotypes.

    PubMed

    Li, Yi; Sulik, Michael J; Eisenberg, Nancy; Spinrad, Tracy L; Lemery-Chalfant, Kathryn; Stover, Daryn A; Verrelli, Brian C

    2016-02-01

    Children's observed effortful control (EC) at 30, 42, and 54 months (n = 145) was predicted from the interaction between mothers' observed parenting with their 30-month-olds and three variants of the solute carrier family C6, member 3 (SLC6A3) dopamine transporter gene (single nucleotide polymorphisms in intron8 and intron13, and a 40 base pair variable number tandem repeat [VNTR] in the 3'-untranslated region [UTR]), as well as haplotypes of these variants. Significant moderating effects were found. Children without the intron8-A/intron13-G, intron8-A/3'-UTR VNTR-10, or intron13-G/3'-UTR VNTR-10 haplotypes (i.e., haplotypes associated with the reduced SLC6A3 gene expression and thus lower dopamine functioning) appeared to demonstrate altered levels of EC as a function of maternal parenting quality, whereas children with these haplotypes demonstrated a similar EC level regardless of the parenting quality. Children with these haplotypes demonstrated a trade-off, such that they showed higher EC, relative to their counterparts without these haplotypes, when exposed to less supportive maternal parenting. The findings revealed a diathesis-stress pattern and suggested that different SLC6A3 haplotypes, but not single variants, might represent different levels of young children's sensitivity/responsivity to early parenting. PMID:25924976

  10. Dopamine transporter gene susceptibility to methylation is associated with impulsivity in nonhuman primates

    PubMed Central

    Rajala, Abigail Z.; Zaitoun, Ismail; Henriques, Jeffrey B.; Converse, Alexander K.; Murali, Dhanabalan; Epstein, Miles L.

    2014-01-01

    Impulsivity, the predisposition to act without regard for negative consequences, is a characteristic of several psychiatric disorders and is thought to result in part from genetic variation in the untranslated region of the dopamine transporter (DAT) gene. As the exact link between genetic mutations and impulsivity has not been established, we used oculomotor behavior to characterize rhesus monkeys as impulsive or calm and genetic/epigenetic analysis and positron emission tomography (PET) to correlate phenotype to DAT genotype, DAT gene methylation, and DAT availability. We found three single nucleotide polymorphisms (SNPs) in the 3′-UTR of the DAT gene, one of which provided a potential site for methylation in the impulsive group. Bisulfite analysis showed that the DNA of the impulsive but not the calm subjects was methylated at one SNP. Because genetic/epigenetic modifications could lead to differences in protein expression, we measured DAT availability using [18F]2β-carbomethoxy-3β-(4-chlorophenyl)-8-(2-fluoroethyl)-nortropane ([18F]FECNT) PET and found higher DAT availability in the internal globus pallidus, an output nucleus of the basal ganglia, of the impulsive group. Higher DAT availability lowers dopamine levels, potentially altering neuronal circuits involved in the initiation of action, thus contributing to the impulsive phenotype. The association between increased methylation in the DAT gene and greater DAT availability suggests that mutations to the regulatory portion of the DAT gene lead to a susceptibility to epigenetic modification resulting in a discrete behavioral phenotype. PMID:25122707

  11. Interaction of Dopamine Transporter Gene and Observed Parenting Behaviors on Attention-Deficit/Hyperactivity Disorder: A Structural Equation Modeling Approach

    ERIC Educational Resources Information Center

    Li, James J.; Lee, Steve S.

    2013-01-01

    Emerging evidence suggests that some individuals may be simultaneously more responsive to the effects from environmental adversity "and" enrichment (i.e., differential susceptibility). Given that parenting behavior and a variable number tandem repeat polymorphism in the 3'untranslated region of the dopamine transporter (DAT1) gene…

  12. Membrane potential shapes regulation of dopamine transporter trafficking at the plasma membrane

    PubMed Central

    Richardson, Ben D.; Saha, Kaustuv; Krout, Danielle; Cabrera, Elizabeth; Felts, Bruce; Henry, L. Keith; Swant, Jarod; Zou, Mu-Fa; Newman, Amy Hauck; Khoshbouei, Habibeh

    2016-01-01

    The dopaminergic system is essential for cognitive processes, including reward, attention and motor control. In addition to DA release and availability of synaptic DA receptors, timing and magnitude of DA neurotransmission depend on extracellular DA-level regulation by the dopamine transporter (DAT), the membrane expression and trafficking of which are highly dynamic. Data presented here from real-time TIRF (TIRFM) and confocal microscopy coupled with surface biotinylation and electrophysiology suggest that changes in the membrane potential alone, a universal yet dynamic cellular property, rapidly alter trafficking of DAT to and from the surface membrane. Broadly, these findings suggest that cell-surface DAT levels are sensitive to membrane potential changes, which can rapidly drive DAT internalization from and insertion into the cell membrane, thus having an impact on the capacity for DAT to regulate extracellular DA levels. PMID:26804245

  13. Dopamine transporters, D2 receptors, and glucose metabolism in corticobasal degeneration.

    PubMed

    Klaffke, Stefanie; Kuhn, Andrea A; Plotkin, Michail; Amthauer, Holger; Harnack, Daniel; Felix, Roland; Kupsch, Andreas

    2006-10-01

    Alterations in presynaptic and postsynaptic dopaminergic system and cerebral glucose metabolism in corticobasal degeneration (CBD) were assessed to evaluate the potential usefulness of different imaging methods for CBD. (123)I-FP-CIT/(123)I-beta-CIT SPECT and (123)I-IBZM SPECT as well as (18)F-FDG PET were performed in eight CBD patients. Decreased presynaptic dopamine transporter binding was found in all CBD patients while D2 receptor binding was reduced in only one patient. (18)F-FDG PET displayed a contralateral hypometabolism in cortical and subcortical areas in seven out of eight patients. Our results demonstrate that glucose metabolism and DAT are reduced, while D2 receptors may be frequently preserved in CBD. PMID:16773621

  14. Synthesis and evaluation of novel tropane derivatives as potential PET imaging agents for the dopamine transporter

    PubMed Central

    Qiao, Hongwen; Zhu, Lin; Lieberman, Brian P.; Zha, Zhihao; Plössl, Karl; Kung, Hank F.

    2012-01-01

    A novel series of tropane derivatives containing a fluorinated tertiary amino or amide at the 2β position was synthesized, labeled with the positron-emitter fluorine-18 (T1/2 = 109.8 min), and tested as potential in vivo dopamine transporter (DAT) imaging agents. The corresponding chlorinated analogs were prepared and employed as precursors for radiolabeling leading to the fluorine-18-labeled derivatives via a one-step nucleophilic aliphatic substitution reaction. In vitro binding results showed that the 2β-amino compounds 6b, 6d and 7b displayed moderately high affinities to DAT (Ki < 10 nM). Biodistribution studies of [18F]6b and [18F]6d showed that the brain uptakes in rats were low. This is likely due to their low lipophilicities. Further structural modifications of these tropane derivatives will be needed to improve their in vivo properties as DAT imaging agents. PMID:22658558

  15. Dopamine transporter gene polymorphism and psychiatric symptoms seen in schizophrenic patients at their first episode

    SciTech Connect

    Inada, Toshiya; Sugita, Tetsuyoshi; Dobashi, Izumi

    1996-07-26

    To investigate the possible role of the dopamine transporter (DAT) gene in determining the phenotype in human subjects, allele frequencies for the 40-bp variable number of tandem repeats (VNTR) polymorphism at this site were compared between 117 Japanese normal controls and 118 schizophrenic patients, including six subgroups: early-onset, those with a family history, and those suffering from one of the following psychiatric symptoms at their first episode: delusion and hallucination; disorganization; bizarre behavior; and negative symptoms. No significant differences were observed between the group as a whole or any subgroup of schizophrenic patients and controls. The results indicate that VNTR polymorphism in the DAT gene is unlikely to be a major contributor to any of the psychiatric parameters examined in the present population of schizophrenic subjects. 12 refs., 1 fig., 2 tabs.

  16. Polymer brushes for molecular transport

    NASA Astrophysics Data System (ADS)

    Tu, Huilin

    Polymer brushes and self-assembled monolayers (SAMs) are used as solid-state, 2-dimensional transport media to confine molecular diffusion. Microcontact printing and photolithography combined with surface-initiated atom transfer radical polymerization (ATRP) are the major techniques to construct patterned polymer brushes. Fluorescence recovery after photobleaching (FRAP) is used to determine diffusion coefficients of fluorescent dye, Prodan, in the polymer brushes and SAMs. Similar diffusion coefficients of Prodan are found for the SAMs formed by chlorotrimethyl silane (CTS) and silanated poly(ethylene glycol) (PEG). No fluorescence recovery is observed on the octadecyltrichlorosilane (OTS) SAM or on the clean silica surface. Patterned poly(N-isopropylacrylamide) (PNIPAAm) brushes were fabricated on silica substrates by surface-initiated ATRP of N-isopropylacrylamide from a micropatterned initiator. Variable temperature ellipsometry indicated that the lower critical solution temperature (LCST) of the hydrated PNIPAAm brush was broad, occurring over the range of 20-35°C. FRAP results of Prodan in PNIPAAm layers indicate that bulk translational diffusion is very slow relative to other diffusion mechanisms. Poly(oligoethylene glycol acrylate) (POEGA) chains are grafted onto silica substrates by surface-initiated ATRP. The diffusion of Prodan in dry POEGA is very fast, which is attributable to the low glass transition temperature of POEGA. The diffusion of Prodan in POEGA under variable humidity can be described reasonably well using WLF equation. Patterned POEGA brushes are not able to confine Prodan to diffuse exclusively inside the polymer regions, and possible explanations are rotational diffusion and a combination of fast surface diffusion and slow bulk diffusion. PNIPAAm chains are tethered onto silica particles via surface-initiated polymerization. Thermal annealing slightly improved the crystallinity of the colloidal assembly. Video microscopy of the two

  17. Altering behavioral responses and dopamine transporter protein with antisense peptide nucleic acids.

    PubMed

    Tyler-McMahon, B M; Stewart, J A; Jackson, J; Bitner, M D; Fauq, A; McCormick, D J; Richelson, E

    2001-10-01

    The dopamine transporter (DAT) plays a role in locomotion and is an obligatory target for amphetamines. We designed and synthesized an antisense peptide nucleic acid (PNA) to rat DAT to examine the effect of this antisense molecule on locomotion and on responsiveness to amphetamines. Rats were injected intraperitoneally daily for 9 days with either saline, an antisense DAT PNA, a scrambled DAT PNA, or a mismatch DAT PNA. On days 7 and 9 after initial motility measurements were taken, the animals were challenged with 10 mg/kg of amphetamine and scored for motility. On day 7, there was no significant difference between the baseline levels of activity of any of the groups or their responses to amphetamine. On day 9, the antisense PNA-treated rats showed a statistically significant increase in their resting motility (P < 0.01). When these rats were challenged with amphetamine, motility of the saline-, scrambled PNA-, and mismatch PNA-treated animals showed increases of 31-, 36-, and 20-fold, respectively, while the antisense PNA-treated animals showed increases of only 3.4-fold (P < 0.01). ELISA results revealed a 32% decrease in striatal DAT in antisense PNA-treated rats compared with the saline, scrambled PNA, and mismatch PNA controls (P < 0.001). These results extend our previous findings that brain proteins can be knocked down in a specific manner by antisense molecules administered extracranially. Additionally, these results suggest some novel approaches for the treatment of diseases dependent upon the function of the dopamine transporter. PMID:11543728

  18. Putamen–midbrain functional connectivity is related to striatal dopamine transporter availability in patients with Lewy body diseases

    PubMed Central

    Rieckmann, A.; Gomperts, S.N.; Johnson, K.A.; Growdon, J.H.; Van Dijk, K.R.A.

    2015-01-01

    Prior work has shown that functional connectivity between the midbrain and putamen is altered in patients with impairments in the dopamine system. This study examines whether individual differences in midbrain–striatal connectivity are proportional to the integrity of the dopamine system in patients with nigrostriatal dopamine loss (Parkinson's disease and dementia with Lewy bodies). We assessed functional connectivity of the putamen during resting state fMRI and dopamine transporter (DAT) availability in the striatum using 11C-Altropane PET in twenty patients. In line with the hypothesis that functional connectivity between the midbrain and the putamen reflects the integrity of the dopaminergic neurotransmitter system, putamen–midbrain functional connectivity was significantly correlated with striatal DAT availability even after stringent control for effects of head motion. DAT availability did not relate to functional connectivity between the caudate and thalamus/prefrontal areas. As such, resting state functional connectivity in the midbrain–striatal pathway may provide a useful indicator of underlying pathology in patients with nigrostriatal dopamine loss. PMID:26137443

  19. Putamen-midbrain functional connectivity is related to striatal dopamine transporter availability in patients with Lewy body diseases.

    PubMed

    Rieckmann, A; Gomperts, S N; Johnson, K A; Growdon, J H; Van Dijk, K R A

    2015-01-01

    Prior work has shown that functional connectivity between the midbrain and putamen is altered in patients with impairments in the dopamine system. This study examines whether individual differences in midbrain-striatal connectivity are proportional to the integrity of the dopamine system in patients with nigrostriatal dopamine loss (Parkinson's disease and dementia with Lewy bodies). We assessed functional connectivity of the putamen during resting state fMRI and dopamine transporter (DAT) availability in the striatum using 11C-Altropane PET in twenty patients. In line with the hypothesis that functional connectivity between the midbrain and the putamen reflects the integrity of the dopaminergic neurotransmitter system, putamen-midbrain functional connectivity was significantly correlated with striatal DAT availability even after stringent control for effects of head motion. DAT availability did not relate to functional connectivity between the caudate and thalamus/prefrontal areas. As such, resting state functional connectivity in the midbrain-striatal pathway may provide a useful indicator of underlying pathology in patients with nigrostriatal dopamine loss. PMID:26137443

  20. The association between heroin expenditure and dopamine transporter availability--a single-photon emission computed tomography study.

    PubMed

    Lin, Shih-Hsien; Chen, Kao Chin; Lee, Sheng-Yu; Chiu, Nan Tsing; Lee, I Hui; Chen, Po See; Yeh, Tzung Lieh; Lu, Ru-Band; Chen, Chia-Chieh; Liao, Mei-Hsiu; Yang, Yen Kuang

    2015-03-30

    One of the consequences of heroin dependency is a huge expenditure on drugs. This underlying economic expense may be a grave burden for heroin users and may lead to criminal behavior, which is a huge cost to society. The neuropsychological mechanism related to heroin purchase remains unclear. Based on recent findings and the established dopamine hypothesis of addiction, we speculated that expenditure on heroin and central dopamine activity may be associated. A total of 21 heroin users were enrolled in this study. The annual expenditure on heroin was assessed, and the availability of the dopamine transporter (DAT) was assessed by single-photon emission computed tomography (SPECT) using [(99m)TC]TRODAT-1. Parametric and nonparametric correlation analyses indicated that annual expenditure on heroin was significantly and negatively correlated with the availability of striatal DAT. After adjustment for potential confounders, the predictive power of DAT availability was significant. Striatal dopamine function may be associated with opioid purchasing behavior among heroin users, and the cycle of spiraling dysfunction in the dopamine reward system could play a role in this association. PMID:25659472

  1. A Biochemical and Functional Protein Complex Involving Dopamine Synthesis and Transport into Synaptic Vesicles

    PubMed Central

    Cartier, Etienne A.; Parra, Leonardo A.; Baust, Tracy B.; Quiroz, Marisol; Salazar, Gloria; Faundez, Victor; Egaña, Loreto; Torres, Gonzalo E.

    2010-01-01

    Synaptic transmission depends on neurotransmitter pools stored within vesicles that undergo regulated exocytosis. In the brain, the vesicular monoamine transporter-2 (VMAT2) is responsible for the loading of dopamine (DA) and other monoamines into synaptic vesicles. Prior to storage within vesicles, DA synthesis occurs at the synaptic terminal in a two-step enzymatic process. First, the rate-limiting enzyme tyrosine hydroxylase (TH) converts tyrosine to di-OH-phenylalanine. Aromatic amino acid decarboxylase (AADC) then converts di-OH-phenylalanine into DA. Here, we provide evidence that VMAT2 physically and functionally interacts with the enzymes responsible for DA synthesis. In rat striata, TH and AADC co-immunoprecipitate with VMAT2, whereas in PC 12 cells, TH co-immunoprecipitates with the closely related VMAT1 and with overexpressed VMAT2. GST pull-down assays further identified three cytosolic domains of VMAT2 involved in the interaction with TH and AADC. Furthermore, in vitro binding assays demonstrated that TH directly interacts with VMAT2. Additionally, using fractionation and immunoisolation approaches, we demonstrate that TH and AADC associate with VMAT2-containing synaptic vesicles from rat brain. These vesicles exhibited specific TH activity. Finally, the coupling between synthesis and transport of DA into vesicles was impaired in the presence of fragments involved in the VMAT2/TH/AADC interaction. Taken together, our results indicate that DA synthesis can occur at the synaptic vesicle membrane, where it is physically and functionally coupled to VMAT2-mediated transport into vesicles. PMID:19903816

  2. Rare Autism-Associated Variants Implicate Syntaxin 1 (STX1 R26Q) Phosphorylation and the Dopamine Transporter (hDAT R51W) in Dopamine Neurotransmission and Behaviors

    PubMed Central

    Cartier, Etienne; Hamilton, Peter J.; Belovich, Andrea N.; Shekar, Aparna; Campbell, Nicholas G.; Saunders, Christine; Andreassen, Thorvald F.; Gether, Ulrik; Veenstra-Vanderweele, Jeremy; Sutcliffe, James S.; Ulery-Reynolds, Paula G.; Erreger, Kevin; Matthies, Heinrich J.G.; Galli, Aurelio

    2015-01-01

    Background Syntaxin 1 (STX1) is a presynaptic plasma membrane protein that coordinates synaptic vesicle fusion. STX1 also regulates the function of neurotransmitter transporters, including the dopamine (DA) transporter (DAT). The DAT is a membrane protein that controls DA homeostasis through the high-affinity re-uptake of synaptically released DA. Methods We adopt newly developed animal models and state-of-the-art biophysical techniques to determine the contribution of the identified gene variants to impairments in DA neurotransmission observed in autism spectrum disorder (ASD). Outcomes Here, we characterize two independent autism-associated variants in the genes that encode STX1 and the DAT. We demonstrate that each variant dramatically alters DAT function. We identify molecular mechanisms that converge to inhibit reverse transport of DA and DA-associated behaviors. These mechanisms involve decreased phosphorylation of STX1 at Ser14 mediated by casein kinase 2 as well as a reduction in STX1/DAT interaction. These findings point to STX1/DAT interactions and STX1 phosphorylation as key regulators of DA homeostasis. Interpretation We determine the molecular identity and the impact of these variants with the intent of defining DA dysfunction and associated behaviors as possible complications of ASD. PMID:25774383

  3. Molecular transport: Catch the carbon dioxide

    NASA Astrophysics Data System (ADS)

    Kirchner, Barbara; Intemann, Barbara

    2016-05-01

    Understanding the minute details of CO2 transport is key to finding new technologies that reduce the hazardous levels of CO2 in our atmosphere. Now, the observation that the transport of CO2 in molten calcium carbonate occurs faster than standard molecular diffusion brings us one step closer.

  4. Regulation of ethanol intake under chronic mild stress: roles of dopamine receptors and transporters

    PubMed Central

    Delis, Foteini; Rombola, Christina; Bellezza, Robert; Rosko, Lauren; Grandy, David K.; Volkow, Nora D.; Thanos, Panayotis K.

    2015-01-01

    Studies have shown that exposure to chronic mild stress decreases ethanol intake and preference in dopamine D2 receptor wild-type mice (Drd2+/+), while it increases intake in heterozygous (Drd2+/−) and knockout (Drd2−/−) mice. Dopaminergic neurotransmission in the basal forebrain plays a major role in the reinforcing actions of ethanol as well as in brain responses to stress. In order to identify neurochemical changes associated with the regulation of ethanol intake, we used in vitro receptor autoradiography to measure the levels and distribution of dopamine D1 and D2 receptors and dopamine transporters (DAT). Receptor levels were measured in the basal forebrain of Drd2+/+, Drd2+/−, and Drd2−/− mice belonging to one of four groups: control (C), ethanol intake (E), chronic mild stress exposure (S), and ethanol intake under chronic mild stress (ES). D2 receptor levels were higher in the lateral and medial striatum of Drd2+/+ ES mice, compared with Drd2+/+ E mice. Ethanol intake in Drd2+/+ mice was negatively correlated with striatal D2 receptor levels. D2 receptor levels in Drd2+/− mice were the same among the four treatment groups. DAT levels were lower in Drd2+/− C and Drd2−/− C mice, compared with Drd2+/+ C mice. Among Drd2+/− mice, S and ES groups had higher DAT levels compared with C and E groups in most regions examined. In Drd2−/− mice, ethanol intake was positively correlated with DAT levels in all regions studied. D1 receptor levels were lower in Drd2+/− and Drd2−/− mice, compared with Drd2+/+, in all regions examined and remained unaffected by all treatments. The results suggest that in normal mice, ethanol intake is associated with D2 receptor-mediated neurotransmission, which exerts a protective effect against ethanol overconsumption under stress. In mice with low Drd2 expression, where DRD2 levels are not further modulated, ethanol intake is associated with DAT function which is upregulated under stress leading to ethanol

  5. Understanding charge transport in molecular electronics.

    PubMed

    Kushmerick, J J; Pollack, S K; Yang, J C; Naciri, J; Holt, D B; Ratner, M A; Shashidhar, R

    2003-12-01

    For molecular electronics to become a viable technology the factors that control charge transport across a metal-molecule-metal junction need to be elucidated. We use an experimentally simple crossed-wire tunnel junction to interrogate how factors such as metal-molecule coupling, molecular structure, and the choice of metal electrode influence the current-voltage characteristics of a molecular junction. PMID:14976024

  6. Regulation of Tyrosine Hydroxylase Expression and Phosphorylation in Dopamine Transporter-Deficient Mice.

    PubMed

    Salvatore, Michael F; Calipari, Erin S; Jones, Sara R

    2016-07-20

    Tyrosine hydroxylase (TH) and dopamine transporters (DATs) regulate dopamine (DA) neurotransmission at the biosynthesis and reuptake steps, respectively. Dysfunction or loss of these proteins occurs in impaired locomotor or addictive behavior, but little is known about the influence of DAT expression on TH function. Differences in TH phosphorylation, DA tissue content, l-DOPA biosynthesis, and DA turnover exist between the somatodendritic and terminal field compartments of nigrostriatal and mesoaccumbens pathways. We examined whether differential DAT expression affects these compartmental differences in DA regulation by comparing TH expression and phosphorylation at ser31 and ser40. In heterozygous DAT knockout (KO) (+/-) mice, DA tissue content and DA turnover were unchanged relative to wild-type mice, despite a 40% reduction in DAT protein expression. In DAT KO (-/-) mice, DA turnover increased in all DA compartments, but DA tissue content decreased (90-96%) only in terminal fields. TH protein expression and phosphorylation were differentially affected within DA pathway compartments by relative expression of DAT. TH protein decreased (∼74%), though to a significantly lesser extent than DA, in striatum and nucleus accumbens (NAc) in DAT -/- mice, with no decrease in substantia nigra or ventral tegmental area. Striatal ser31 TH phosphorylation and recovery of DA relative to TH protein expression in DAT +/- and DAT -/- mice decreased, whereas ser40 TH phosphorylation increased ∼2- to 3-fold in striatum and NAc of DAT -/- mice. These results suggest that DAT expression affects TH expression and phosphorylation largely in DA terminal field compartments, further corroborating evidence for dichotomous regulation of TH between somatodendritic and terminal field compartments of the nigrostriatal and mesoaccumbens pathways. PMID:27124386

  7. Temporal pattern of cocaine intake determines tolerance vs sensitization of cocaine effects at the dopamine transporter.

    PubMed

    Calipari, Erin S; Ferris, Mark J; Zimmer, Benjamin A; Roberts, David C S; Jones, Sara R

    2013-11-01

    The dopamine transporter (DAT) is responsible for terminating dopamine (DA) signaling and is the primary site of cocaine's reinforcing actions. Cocaine self-administration has been shown previously to result in changes in cocaine potency at the DAT. To determine whether the DAT changes associated with self-administration are due to differences in intake levels or temporal patterns of cocaine-induced DAT inhibition, we manipulated cocaine access to produce either continuous or intermittent elevations in cocaine brain levels. Long-access (LgA, 6 h) and short-access (ShA, 2 h) continuous self-administration produced similar temporal profiles of cocaine intake that were sustained throughout the session; however, LgA had greater intake. ShA and intermittent-access (IntA, 6 h) produced the same intake, but different temporal profiles, with 'spiking' brain levels in IntA compared with constant levels in ShA. IntA consisted of 5-min access periods alternating with 25-min timeouts, which resulted in bursts of high responding followed by periods of no responding. DA release and uptake, as well as the potency of cocaine for DAT inhibition, were assessed by voltammetry in the nucleus accumbens slices following control, IntA, ShA, and LgA self-administration. Continuous-access protocols (LgA and ShA) did not change DA parameters, but the 'spiking' protocol (IntA) increased both release and uptake of DA. In addition, high continuous intake (LgA) produced tolerance to cocaine, while 'spiking' (IntA) produced sensitization, relative to ShA and naive controls. Thus, intake and pattern can both influence cocaine potency, and tolerance seems to be produced by high intake, while sensitization is produced by intermittent temporal patterns of intake. PMID:23719505

  8. Membrane Cholesterol Modulates the Outward Facing Conformation of the Dopamine Transporter and Alters Cocaine Binding*

    PubMed Central

    Hong, Weimin C.; Amara, Susan G.

    2010-01-01

    Clearance of synaptically released dopamine is regulated by the plasmalemmal dopamine transporter (DAT), an integral membrane protein that resides within a complex lipid milieu. Here we demonstrate that cholesterol, a major component of the lipid bilayer, can modulate the conformation of DAT and alter cocaine binding to DAT. In striatal synaptosomes and transfected cells, DAT was in cholesterol-rich membrane fractions after mild detergent extraction. After increasing the membrane cholesterol content by treatment of water-soluble cholesterol (cholesterol mixed with methyl-β-cyclodextrin), we observed an increase in DAT binding Bmax values for cocaine analogs [3H]WIN35428 and [125I]RTI-55, but similar levels of DAT proteins on the cell surface were shown by surface biotinylation assays. Membrane cholesterol addition also markedly enhanced the accessibility of cysteine sulfhydryl moieties in DAT as probed by a membrane-impermeable maleimide-biotin conjugate. We identified cysteine 306, a juxtamembrane residue on transmembrane domain 6 (TM6) of DAT, as the intrinsic residue exhibiting enhanced reactivity. Similar effects on DAT cysteine accessibility and radioligand binding were observed with addition of zinc, a reagent known to promote the outward facing conformation of DAT. Using substituted cysteine mutants on various positions likely to be extracellular, we identified additional residues located on TM1, TM6, TM7, and TM12 of DAT that are sensitive to alterations in the membrane cholesterol content. Our findings in transfected cells and native tissues support the hypothesis that DAT adopts an outward facing conformation in a cholesterol-rich membrane environment, suggesting a novel modulatory role of the surrounding membrane lipid milieu on DAT function. PMID:20688912

  9. Gender and environmental enrichment impact dopamine transporter expression after experimental traumatic brain injury.

    PubMed

    Wagner, Amy K; Chen, Xiangbai; Kline, Anthony E; Li, Youming; Zafonte, Ross D; Dixon, C Edward

    2005-10-01

    Dopamine (DA) systems are implicated in cognitive deficits following traumatic brain injury (TBI). Rodent studies have demonstrated that both environmental enrichment (EE) and sex hormones can influence DA systems. The dopamine transporter (DAT) plays a crucial role in regulating DA transmission, and previous work shows that DAT is decreased after TBI in males. Therefore, the purpose of this study was to examine the effects of gender and EE on frontal cortex and striatal DAT expression after TBI. Sprague-Dawley male (n = 24) and cycling female rats (n = 24) were placed into EE or standard housing after controlled cortical impact (2.7 mm, 4.0 m/s) injury or sham surgery (eight groups, n = 6/group). Four weeks post-surgery, bilateral frontal cortex and striatal DAT expression was examined via Western blot. Results demonstrated that there was a significant effect of injury, EE, and region on DAT expression (P < 0.05 all comparisons) on female groups. There were no significant DAT decreases in any region as a result of injury, however, EE did promote significant post-injury DAT decreases in the striatum and ipsilateral frontal cortex (P < 0.05 all comparisons) compared to female shams housed in the standard environment. For males, there was a significant effect of injury, EE, and region for male groups (P < 0.05 all comparisons). There were decreases in DAT expression in three regions studied for injured males housed in the standard environment compared to sham males in the standard environment (P < 0.05 all comparisons), however, EE did not add significantly to post-injury DAT decreases in these regions. These results suggest that CCI causes larger relative decreases in DAT expression for males compared to females and that treatment with EE has larger effects on post-injury DAT expression for females than males. These findings may have some relevance to treatment paradigms using dopaminergic neurostimulants after TBI. PMID:16023635

  10. Membrane cholesterol modulates the outward facing conformation of the dopamine transporter and alters cocaine binding.

    PubMed

    Hong, Weimin C; Amara, Susan G

    2010-10-15

    Clearance of synaptically released dopamine is regulated by the plasmalemmal dopamine transporter (DAT), an integral membrane protein that resides within a complex lipid milieu. Here we demonstrate that cholesterol, a major component of the lipid bilayer, can modulate the conformation of DAT and alter cocaine binding to DAT. In striatal synaptosomes and transfected cells, DAT was in cholesterol-rich membrane fractions after mild detergent extraction. After increasing the membrane cholesterol content by treatment of water-soluble cholesterol (cholesterol mixed with methyl-β-cyclodextrin), we observed an increase in DAT binding B(max) values for cocaine analogs [(3)H]WIN35428 and [(125)I]RTI-55, but similar levels of DAT proteins on the cell surface were shown by surface biotinylation assays. Membrane cholesterol addition also markedly enhanced the accessibility of cysteine sulfhydryl moieties in DAT as probed by a membrane-impermeable maleimide-biotin conjugate. We identified cysteine 306, a juxtamembrane residue on transmembrane domain 6 (TM6) of DAT, as the intrinsic residue exhibiting enhanced reactivity. Similar effects on DAT cysteine accessibility and radioligand binding were observed with addition of zinc, a reagent known to promote the outward facing conformation of DAT. Using substituted cysteine mutants on various positions likely to be extracellular, we identified additional residues located on TM1, TM6, TM7, and TM12 of DAT that are sensitive to alterations in the membrane cholesterol content. Our findings in transfected cells and native tissues support the hypothesis that DAT adopts an outward facing conformation in a cholesterol-rich membrane environment, suggesting a novel modulatory role of the surrounding membrane lipid milieu on DAT function. PMID:20688912

  11. PET evaluation of the dopamine system of the human brain

    SciTech Connect

    Volkow, N.D.; Fowler, J.S.; Gatley, S. |

    1996-07-01

    Dopamine plays a pivotal role in the regulation and control of movement, motivation and cognition. It also is closely linked to reward, reinforcement and addiction. Abnormalities in brain dopamine are associated with many neurological and psychiatric disorders including Parkinson`s disease, schizophrenia and substance abuse. This close association between dopamine and neurological and psychiatric diseases and with substance abuse make it an important topic in research in the neurosciences and an important molecular target in drug development. PET enables the direct measurement of components of the dopamine system in the living human brain. It relies on radiotracers which label dopamine receptors, dopamine transporters, precursors of dopamine or compounds which have specificity for the enzymes which degrade dopamine. Additionally, by using tracers that provide information on regional brain metabolism or blood flow as well as neurochemically specific pharmacological interventions, PET can be used to assess the functional consequences of change in brain dopamine activity. PET dopamine measurements have been used to investigate the normal human brain and its involvement in psychiatric and neurological diseases. It has also been used in psychopharmacological research to investigate dopamine drugs used in the treatment of Parkinson`s disease and of schizophrenia as well as to investigate the effects of drugs of abuse on the dopamine system. Since various functional and neurochemical parameters can be studied in the same subject, PET enables investigation of the functional integrity of the dopamine system in the human brain and investigation of the interactions of dopamine with other neurotransmitters. This paper summarizes the different tracers and experimental strategies developed to evaluate the various elements of the dopamine system in the human brain with PET and their applications to clinical research. 254 refs., 7 figs., 3 tabs.

  12. The effects of benzofury (5-APB) on the dopamine transporter and 5-HT2-dependent vasoconstriction in the rat.

    PubMed

    Dawson, Patrick; Opacka-Juffry, Jolanta; Moffatt, James D; Daniju, Yusuf; Dutta, Neelakshi; Ramsey, John; Davidson, Colin

    2014-01-01

    5-APB, commonly marketed as 'benzofury' is a new psychoactive substance and erstwhile 'legal high' which has been implicated in 10 recent drug-related deaths in the UK. This drug was available on the internet and in 'head shops' and was one of the most commonly sold legal highs up until its recent UK temporary ban (UK Home Office). Despite its prominence, very little is known about its pharmacology. This study was undertaken to examine the pharmacology of 5-APB in vitro. We hypothesised that 5-APB would activate the dopamine and 5-HT systems which may underlie its putative stimulant and hallucinogenic effects. Autoradiographic studies showed that 5-APB displaced both [(125)I] RTI-121 and [(3)H] ketanserin from rat brain tissue suggesting affinity at the dopamine transporter and 5-HT2 receptor sites respectively. Voltammetric studies in rat accumbens brain slices revealed that 5-APB slowed dopamine reuptake, and at high concentrations caused reverse transport of dopamine. 5-APB also caused vasoconstriction of rat aorta, an effect antagonised by the 5-HT2A receptor antagonist ketanserin, and caused contraction of rat stomach fundus, which was reversed by the 5-HT2B receptor antagonist RS-127445. These data show that 5-APB interacts with the dopamine transporter and is an agonist at the 5-HT2A and 5-HT2B receptors in the rat. Thus 5-APB's pharmacology is consistent with it having both stimulant and hallucinogenic properties. In addition, 5-APB's activity at the 5-HT2B receptor may cause cardiotoxicity. PMID:24012617

  13. Electrochemical sensor for dopamine based on a novel graphene-molecular imprinted polymers composite recognition element.

    PubMed

    Mao, Yan; Bao, Yu; Gan, Shiyu; Li, Fenghua; Niu, Li

    2011-10-15

    A novel composite of graphene sheets/Congo red-molecular imprinted polymers (GSCR-MIPs) was synthesized through free radical polymerization (FRP) and applied as a molecular recognition element to construct dopamine (DA) electrochemical sensor. The template molecules (DA) were firstly absorbed at the GSCR surface due to their excellent affinity, and subsequently, selective copolymerization of methacrylic acid (MAA) and ethylene glycol dimethacrylate (EGDMA) was further achieved at the GSCR surface. Potential scanning was presented to extract DA molecules from the imprinted polymers film, and as a result, DA could be rapidly and completely removed by this way. With regard to the traditional MIPs, the GSCR-MIPs not only possessed a faster desorption and adsorption dynamics, but also exhibited a higher selectivity and binding capacity toward DA molecule. As a consequence, an electrochemical sensor for highly sensitive and selective detection of DA was successfully constructed as demonstration based on the synthesized GSCR-MIPs nanocomposites. Under experimental conditions, selective detection of DA in a linear concentration range of 1.0 × 10(-7)-8.3 × 10(-4)M was obtained, which revealed a lower limit of detection and wider linear response compared to some previously reported DA electrochemical MIPs sensors. The new DA electrochemical sensor based on GSCR-MIPs composites also exhibited excellent repeatability, which expressed as relative standard deviation (RSD) was about 2.50% for 30 repeated analyses of 20 μM DA. PMID:21824760

  14. Brominated and radioiodinated derivatives of methylphenidate (MP): Potential imaging agents for the dopamine (DA) transporter

    SciTech Connect

    Pan, D.; Gatley, S.J.; Dewey, S.L.

    1994-05-01

    MP (Ritalin) is a psychomotor stimulant used in the treatment of attention-deficit hyperactivity disorder. The therapeutic properties of MP are thought to be mediated by its binding to a site on the DA transporter, resulting in inhibition of DA reuptake and enhanced levels of synaptic dopamine. MP also inhibits reuptake of norepinephrine (NE) in vitro. MP has two chiral centers, but its pharmacological activity is believed due solely to the d-threo isomer. We have found that d,l-threo-C-11 MP has favorable properties for PET studies, and therefore examined the effects of incorporating halogen atoms into the phenyl ring of MP, with a view to preparing C-11 and I-123 MP analogs as potential PET/SPECT tracers. We synthesized the 2-, 3- and 4-bromo MP analogs from the corresponding bromophenylacetonitriles by modification of the original synthesis of MP. In in vitro binding assays all three d,l-threo bromo compounds had higher affinities than MP for DA transporter sites labeled with tritiated WIN 35,428 (3->4-, 2->MP). They also showed high activity with NE reuptake sites labeled with tritiated nisoxetine. They were active in vivo as demonstrated by inhibition of heart uptake of tritiated NE in the mouse, and elevation of striatal extracellular DA (microdialysis) and stimulation of locomotor activity in the rat.

  15. Molecular Imaging of Transporters with Positron Emission Tomography

    NASA Astrophysics Data System (ADS)

    Antoni, Gunnar; Sörensen, Jens; Hall, Håkan

    Positron emission tomography (PET) visualization of brain components in vivo is a rapidly growing field. Molecular imaging with PET is also increasingly used in drug development, especially for the determination of drug receptor interaction for CNS-active drugs. This gives the opportunity to relate clinical efficacy to per cent receptor occupancy of a drug on a certain targeted receptor and to relate drug pharmacokinetics in plasma to interaction with target protein. In the present review we will focus on the study of transporters, such as the monoamine transporters, the P-glycoprotein (Pgp) transporter, the vesicular monoamine transporter type 2, and the glucose transporter using PET radioligands. Neurotransmitter transporters are presynaptically located and in vivo imaging using PET can therefore be used for the determination of the density of afferent neurons. Several promising PET ligands for the noradrenaline transporter (NET) have been labeled and evaluated in vivo including in man, but a really useful PET ligand for NET still remains to be identified. The most promising tracer to date is (S,S)-[18F]FMeNER-D2. The in vivo visualization of the dopamine transporter (DAT) may give clues in the evaluation of conditions related to dopamine, such as Parkinson's disease and drug abuse. The first PET radioligands based on cocaine were not selective, but more recently several selective tracers such as [11C]PE2I have been characterized and shown to be suitable as PET radioligands. Although there are a large number of serotonin transporter inhibitors used today as SSRIs, it was not until very recently, when [11C]McN5652 was synthesized, that this transporter was studied using PET. New candidates as PET radioligands for the SERT have subsequently been developed and [11C]DASB and [11C]MADAM and their analogues are today the most promising ligands. The existing radioligands for Pgp transporters seem to be suitable tools for the study of both peripheral and central drug

  16. Postnatal manganese exposure alters dopamine transporter function in adult rats: Potential impact on nonassociative and associative processes.

    PubMed

    McDougall, S A; Reichel, C M; Farley, C M; Flesher, M M; Der-Ghazarian, T; Cortez, A M; Wacan, J J; Martinez, C E; Varela, F A; Butt, A E; Crawford, C A

    2008-06-23

    In the present study, we examined whether exposing rats to a high-dose regimen of manganese chloride (Mn) during the postnatal period would depress presynaptic dopamine functioning and alter nonassociative and associative behaviors. To this end, rats were given oral supplements of Mn (750 microg/day) on postnatal days (PD) 1-21. On PD 90, dopamine transporter (DAT) immunoreactivity and [3H]dopamine uptake were assayed in the striatum and nucleus accumbens, while in vivo microdialysis was used to measure dopamine efflux in the same brain regions. The effects of postnatal Mn exposure on nigrostriatal functioning were evaluated by assessing rotorod performance and amphetamine-induced stereotypy in adulthood. In terms of associative processes, both cocaine-induced conditioned place preference (CPP) and sucrose-reinforced operant responding were examined. Results showed that postnatal Mn exposure caused persistent declines in DAT protein expression and [3H]dopamine uptake in the striatum and nucleus accumbens, as well as long-term reductions in striatal dopamine efflux. Rotorod performance did not differ according to exposure condition, however Mn-exposed rats did exhibit substantially more amphetamine-induced stereotypy than vehicle controls. Mn exposure did not alter performance on any aspect of the CPP task (preference, extinction, or reinstatement testing), nor did Mn affect progressive ratio responding (a measure of motivation). Interestingly, acquisition of a fixed ratio task was impaired in Mn-exposed rats, suggesting a deficit in procedural learning. In sum, these results indicate that postnatal Mn exposure causes persistent declines in various indices of presynaptic dopaminergic functioning. Mn-induced alterations in striatal functioning may have long-term impact on associative and nonassociative behavior. PMID:18485605

  17. 2-Substituted 3β-Aryltropane Cocaine Analogs Produce Atypical Effects without Inducing Inward-Facing Dopamine Transporter Conformations

    PubMed Central

    Hong, Weimin C.; Kopajtic, Theresa A.; Xu, Lifen; Lomenzo, Stacey A.; Jean, Bernandie; Madura, Jeffry D.; Surratt, Christopher K.; Trudell, Mark L.

    2016-01-01

    Previous structure-activity relationship studies indicate that a series of cocaine analogs, 3β-aryltropanes with 2β-diarylmethoxy substituents, selectively bind to the dopamine transporter (DAT) with nanomolar affinities that are 10-fold greater than the affinities of their corresponding 2α-enantiomers. The present study compared these compounds to cocaine with respect to locomotor effects in mice, and assessed their ability to substitute for cocaine (10 mg/kg, i.p.) in rats trained to discriminate cocaine from saline. Despite nanomolar DAT affinity, only the 2β-Ph2COCH2-3β-4-Cl-Ph analog fully substituted for cocaine-like discriminative effects. Whereas all of the 2β compounds increased locomotion, only the 2β-(4-ClPh)PhCOCH2-3β-4-Cl-Ph analog had cocaine-like efficacy. None of the 2α-substituted compounds produced either of these cocaine-like effects. To explore the molecular mechanisms of these drugs, their effects on DAT conformation were probed using a cysteine-accessibility assay. Previous reports indicate that cocaine binds with substantially higher affinity to the DAT in its outward (extracellular)- compared with inward-facing conformation, whereas atypical DAT inhibitors, such as benztropine, have greater similarity in affinity to these conformations, and this is postulated to explain their divergent behavioral effects. All of the 2β- and 2α-substituted compounds tested altered cysteine accessibility of DAT in a manner similar to cocaine. Furthermore, molecular dynamics of in silico inhibitor-DAT complexes suggested that the 2-substituted compounds reach equilibrium in the binding pocket in a cocaine-like fashion. These behavioral, biochemical, and computational results show that aryltropane analogs can bind to the DAT and stabilize outward-facing DAT conformations like cocaine, yet produce effects that differ from those of cocaine. PMID:26769919

  18. Hypoxia-inducible factor-1α upregulates tyrosine hydroxylase and dopamine transporter by nuclear receptor ERRγ in SH-SY5Y cells.

    PubMed

    Lim, Juhee; Kim, Hyo-In; Bang, Yeojin; Seol, Wongi; Choi, Hueng-Sik; Choi, Hyun Jin

    2015-04-15

    Hypoxia-inducible factor-1α (HIF-1α) is a transcription factor relevant to the development of many mammalian organs including the brain. However, the molecular mechanisms by which signaling events mediate neuronal differentiation have not been fully elucidated. In the present study, we show for the first time that the orphan nuclear receptor estrogen-related receptor γ (ERRγ) is upregulated by HIF-1α and plays essential roles in HIF-1α-induced upregulation of dopaminergic marker molecules such as tyrosine hydroxylase and dopamine transporter. We found that deferoxamine upregulated HIF-1α and enhanced the dopaminergic phenotype and neurite outgrowth of SH-SY5Y cells. Deferoxamine activated transcription and protein expression of ERRγ, and deferoxamine-induced upregulation of tyrosine hydroxylase and dopamine transporter was attenuated by using the ERRγ inverse agonist or silencing ERRγ. Altogether, these results suggest that HIF-1α can positively regulate the dopaminergic phenotype through ERRγ. This study could provide new perspectives for understanding the mechanisms underlying the promotion of dopaminergic neuronal differentiation by hypoxia. PMID:25807177

  19. Methamphetamine-Induced Dopamine Transporter Complex Formation and Dopaminergic Deficits: The Role of D2 Receptor Activation

    PubMed Central

    Hadlock, Gregory C.; Chu, Pei-Wen; Walters, Elliot T.; Hanson, Glen R.

    2010-01-01

    Methamphetamine (METH) abuse is a serious public health issue. Of particular concern are findings that repeated high-dose administrations of METH cause persistent dopaminergic deficits in rodents, nonhuman primates, and humans. Previous studies have also revealed that METH treatment causes alterations in the dopamine transporter (DAT), including the formation of higher molecular mass DAT-associated complexes. The current study extends these findings by examining mechanisms underlying DAT complex formation. The association among DAT complex formation and other METH-induced phenomena, including alterations in vesicular monoamine transporter 2 (VMAT2) immunoreactivity, astrocytic activation [as assessed by increased glial fibrillary acidic protein (GFAP) immunoreactivity], and persistent dopaminergic deficits was also explored. Results revealed that METH-induced DAT complex formation and reductions in VMAT2 immunoreactivity precede increases in GFAP immunoreactivity. Furthermore, and as reported previously for DAT complexes, pretreatment with the D2 receptor antagonist eticlopride [S-(−)-3-chloro-5-ethyl-N-[(1-ethyl-2-pyrrolidinyl)methyl]-6-hydroxy-2-methoxybenzamide hydrochloride] attenuated the decrease in VMAT2 immunoreactivity as assessed 24 h after METH treatment. DAT complexes distinct from those present 24 h after METH treatment, decreases in VMAT2 immunoreactivity, and increased GFAP immunoreactivity were present 48 to 72 h after METH treatment. Pretreatment with eticlopride attenuated each of these phenomena. Finally, DAT complexes were present 7 days after METH treatment, a time point at which VMAT2 and DAT monomer immunoreactivity were also reduced. Eticlopride pretreatment attenuated each of these phenomena. These findings provide novel insight into not only receptor-mediated mechanisms underlying the effects of METH but also the interaction among factors that probably are associated with the persistent dopaminergic deficits caused by the stimulant. PMID

  20. Ack1 is a dopamine transporter endocytic brake that rescues a trafficking-dysregulated ADHD coding variant

    PubMed Central

    Wu, Sijia; Bellve, Karl D.; Fogarty, Kevin E.; Melikian, Haley E.

    2015-01-01

    The dopamine (DA) transporter (DAT) facilitates high-affinity presynaptic DA reuptake that temporally and spatially constrains DA neurotransmission. Aberrant DAT function is implicated in attention-deficit/hyperactivity disorder and autism spectrum disorder. DAT is a major psychostimulant target, and psychostimulant reward strictly requires binding to DAT. DAT function is acutely modulated by dynamic membrane trafficking at the presynaptic terminal and a PKC-sensitive negative endocytic mechanism, or “endocytic brake,” controls DAT plasma membrane stability. However, the molecular basis for the DAT endocytic brake is unknown, and it is unknown whether this braking mechanism is unique to DAT or common to monoamine transporters. Here, we report that the cdc42-activated, nonreceptor tyrosine kinase, Ack1, is a DAT endocytic brake that stabilizes DAT at the plasma membrane and is released in response to PKC activation. Pharmacologic and shRNA-mediated Ack1 silencing enhanced basal DAT internalization and blocked PKC-stimulated DAT internalization, but had no effects on SERT endocytosis. Both cdc42 activation and PKC stimulation converge on Ack1 to control Ack1 activity and DAT endocytic capacity, and Ack1 inactivation is required for stimulated DAT internalization downstream of PKC activation. Moreover, constitutive Ack1 activation is sufficient to rescue the gain-of-function endocytic phenotype exhibited by the ADHD DAT coding variant, R615C. These findings reveal a unique endocytic control switch that is highly specific for DAT. Moreover, the ability to rescue the DAT(R615C) coding variant suggests that manipulating DAT trafficking mechanisms may be a potential therapeutic approach to correct DAT coding variants that exhibit trafficking dysregulation. PMID:26621748

  1. Dosimetry of an iodine-123-labeled tropane to image dopamine transporters

    SciTech Connect

    Mozley, P.D.; Stubbs, J.B.; Kim, H.J.

    1996-01-01

    N-(3-iodopropen-2-yl)-2{beta}-carbomethoxy-3{beta}(4-chlorophenyl)tropane (IPT) is an analog of cocaine that selectively binds the presynaptic dopamine transporter. The present study sought to measure the radiation dosimetry of IPT in seven healthy human volunteers. Dynamic renal scans were acquired immediately after the intravenous administration of 165 {+-} 16 MBq (4.45 {+-} 0.42 mCi) of [{sup 123}I]IPT. Between 7 and 12 sets of whole-body scans were acquired over the next 24 hr. The 24-hr renal excretion fractions were measured from conjugate emission scans of 7-11 discreet voided urine specimens. The fraction of the administered dose in 11 organs and each urine specimen was quantified from the attenuation-corrected geometric mean counts in opposing views. Subject-specific residence times were evaluated for each subject independently by fitting the time-activity curves to a multicompartmental model. The radiation doses were estimated with the MIRD technique from the residence times for each subject individually before any results were averaged. The findings showed that IPT was excreted rapidly by the renal system. There were no reservoirs of retained activity outside the basal ganglia, where SPECT images in these subjects showed that the mean ratio of caudate to calcarine cortex averaged 25:1 at 3 hr after injection (range 19.6-32 hr). The basal ganglia received a radiation dose of 0.028 mGy/MBq (0.10 rad/mCi). The dose-limiting organ in men was the stomach, which received an estimated 0.11 mGy/MBq (0.37 rad/mCi). In women, the critical organ was the urinary bladder at 0.14 mGy/MBq (0.51 rad/mCi). Relatively high-contrast images of the presynaptic dopamine transporters in the basal ganglia can be acquired with 185 MBq (5 mCi) of [{sup 123}I]IPT. The radiation exposure that results is significantly less than the maximum allowed by current safety guidelines for research volunteers. 33 refs., 4 figs., 3 tabs.

  2. Selective toxicity of L-DOPA to dopamine transporter-expressing neurons and locomotor behavior in zebrafish larvae.

    PubMed

    Stednitz, Sarah J; Freshner, Briana; Shelton, Samantha; Shen, Tori; Black, Donovan; Gahtan, Ethan

    2015-01-01

    Dopamine signaling is conserved across all animal species and has been implicated in the disease process of many neurological disorders, including Parkinson's disease (PD). The primary neuropathology in PD involves the death of dopaminergic cells in the substantia nigra (SN), an anatomical region of the brain implicated in dopamine production and voluntary motor control. Increasing evidence suggests that the neurotransmitter dopamine may have a neurotoxic metabolic product (DOPAL) that selectively damages dopaminergic cells. This study was designed to test this theory of oxidative damage in an animal model of Parkinson's disease, using a transgenic strain of zebrafish with fluorescent labeling of cells that express the dopamine transporter. The pretectum and ventral diencephalon exhibited reductions in cell numbers due to L-DOPA treatment while reticulospinal neurons that do not express the DAT were unaffected, and this was partially rescued by monoamine oxidase inhibition. Consistent with the MPTP model of PD in zebrafish larvae, spontaneous locomotor behavior in L-DOPA treated animals was depressed following a 24-h recovery period, while visually-evoked startle response rates and latencies were unaffected. PMID:26546233

  3. Vibrational Heat Transport in Molecular Junctions

    NASA Astrophysics Data System (ADS)

    Segal, Dvira; Agarwalla, Bijay Kumar

    2016-05-01

    We review studies of vibrational energy transfer in a molecular junction geometry, consisting of a molecule bridging two heat reservoirs, solids or large chemical compounds. This setup is of interest for applications in molecular electronics, thermoelectrics, and nanophononics, and for addressing basic questions in the theory of classical and quantum transport. Calculations show that system size, disorder, structure, dimensionality, internal anharmonicities, contact interaction, and quantum coherent effects are factors that combine to determine the predominant mechanism (ballistic/diffusive), effectiveness (poor/good), and functionality (linear/nonlinear) of thermal conduction at the nanoscale. We review recent experiments and relevant calculations of quantum heat transfer in molecular junctions. We recount the Landauer approach, appropriate for the study of elastic (harmonic) phononic transport, and outline techniques that incorporate molecular anharmonicities. Theoretical methods are described along with examples illustrating the challenge of reaching control over vibrational heat conduction in molecules.

  4. Vibrational Heat Transport in Molecular Junctions.

    PubMed

    Segal, Dvira; Agarwalla, Bijay Kumar

    2016-05-27

    We review studies of vibrational energy transfer in a molecular junction geometry, consisting of a molecule bridging two heat reservoirs, solids or large chemical compounds. This setup is of interest for applications in molecular electronics, thermoelectrics, and nanophononics, and for addressing basic questions in the theory of classical and quantum transport. Calculations show that system size, disorder, structure, dimensionality, internal anharmonicities, contact interaction, and quantum coherent effects are factors that combine to determine the predominant mechanism (ballistic/diffusive), effectiveness (poor/good), and functionality (linear/nonlinear) of thermal conduction at the nanoscale. We review recent experiments and relevant calculations of quantum heat transfer in molecular junctions. We recount the Landauer approach, appropriate for the study of elastic (harmonic) phononic transport, and outline techniques that incorporate molecular anharmonicities. Theoretical methods are described along with examples illustrating the challenge of reaching control over vibrational heat conduction in molecules. PMID:27215814

  5. Nitric oxide inhibits uptake of dopamine and N-methyl-4-phenylpyridinium (MPP+) but not release of MPP+ in rat C6 glioma cells expressing human dopamine transporter

    PubMed Central

    Cao, Bo-Jin; Reith, Maarten E A

    2002-01-01

    Conflicting results have been reported regarding the influence of nitric oxide (NO) and peroxynitrite on dopamine (DA) uptake and release. In the present study, effects of NO donors were studied in rat C6 glioma cells expressing human DA transporter. [3H]-DA uptake was inhibited by S-nitroso-thiol S-nitroso-N-acetylpenicillamine, spermine/NO, diethylamine/NO (DEA/NO), (Z)-1-[N-(3-ammoniopropyl)-N-(n-propyl)-amino]/NO (PAPA/NO), and 3-morphosynodiomine (SIN-1) in a rank order correlating with their half lives as NO donors, whereas no effect was observed for diethylenetriamine/NO and dipropylenetriamine/NO, which release NO very slowly. Hydroxycobalamin, a NO scavenger, but not superoxide dismutase and catalase, enzymes that metabolize superoxide and hydrogen peroxide, respectively, abolished the inhibitory effect of DEA/NO and SIN-1, indicating that they inhibit DA uptake through a mechanism related to the production of NO but unrelated to the formation of peroxynitrite. In consonance, peroxynitrite did not alter DA uptake in the present system. DEA/NO and PAPA/NO reduced [3H]-MPP+ uptake, whereas the release of [3H]-MPP+ was not modified, demonstrating that NO can inhibit uptake of DA transporter substrate without accelerating DA transporter-mediated reverse transport of substrate under the same conditions. PMID:12466224

  6. Molecular Mechanism of Biological Proton Transport

    SciTech Connect

    Pomes, R.

    1998-09-01

    Proton transport across lipid membranes is a fundamental aspect of biological energy transduction (metabolism). This function is mediated by a Grotthuss mechanism involving proton hopping along hydrogen-bonded networks embedded in membrane-spanning proteins. Using molecular simulations, the authors have explored the structural, dynamic, and thermodynamic properties giving rise to long-range proton translocation in hydrogen-bonded networks involving water molecules, or water wires, which are emerging as ubiquitous H{sup +}-transport devices in biological systems.

  7. Increased dopamine transporter function as a mechanism for dopamine hypoactivity in the adult infralimbic medial prefrontal cortex following adolescent social stress.

    PubMed

    Novick, Andrew M; Forster, Gina L; Hassell, James E; Davies, Daniel R; Scholl, Jamie L; Renner, Kenneth J; Watt, Michael J

    2015-10-01

    Being bullied during adolescence is associated with later mental illnesses characterized by deficits in cognitive tasks mediated by prefrontal cortex (PFC) dopamine (DA). Social defeat of adolescent male rats, as a model of teenage bullying victimization, results in medial PFC (mPFC) dopamine (DA) hypofunction in adulthood that is associated with increased drug seeking and working memory deficits. Increased expression of the DA transporter (DAT) is also seen in the adult infralimbic mPFC following adolescent defeat. We propose the functional consequence of this increased DAT expression is enhanced DA clearance and subsequently decreased infralimbic mPFC DA availability. To test this, in vivo chronoamperometry was used to measure changes in accumulation of the DA signal following DAT blockade, with increased DAT-mediated clearance being reflected by lower DA signal accumulation. Previously defeated rats and controls were pre-treated with the norepinephrine transporter (NET) inhibitor desipramine (20 mg/kg, ip.) to isolate infralimbic mPFC DA clearance to DAT, then administered the selective DAT inhibitor GBR-12909 (20 or 40 mg/kg, sc.). Sole NET inhibition with desipramine produced no differences in DA signal accumulation between defeated rats and controls. However, rats exposed to adolescent social defeat demonstrated decreased DA signal accumulation compared to controls in response to both doses of GBR-12909, indicating greater DAT-mediated clearance of infralimbic mPFC DA. These results suggest that protracted increases in infralimbic mPFC DAT function represent a mechanism by which adolescent social defeat stress produces deficits in adult mPFC DA activity and corresponding behavioral and cognitive dysfunction. PMID:26056032

  8. Tissue Specific Expression of Cre in Rat Tyrosine Hydroxylase and Dopamine Active Transporter-Positive Neurons.

    PubMed

    Liu, Zhenyi; Brown, Andrew; Fisher, Dan; Wu, Yumei; Warren, Joe; Cui, Xiaoxia

    2016-01-01

    The rat is a preferred model system over the mouse for neurological studies, and cell type-specific Cre expression in the rat enables precise ablation of gene function in neurons of interest, which is especially valuable for neurodegenerative disease modeling and optogenetics. Yet, few such Cre rats are available. Here we report the characterization of two Cre rats, tyrosine hydroxylase (TH)-Cre and dopamine active transporter (DAT or Slc6a3)-Cre, by using a combination of immunohistochemistry (IHC) and mRNA fluorescence in situ hybridization (FISH) as well as a fluorescent reporter for Cre activity. We detected Cre expression in expected neurons in both Cre lines. Interestingly, we also found that in Th-Cre rats, but not DAT-Cre rats, Cre is expressed in female germ cells, allowing germline excision of the floxed allele and hence the generation of whole-body knockout rats. In summary, our data demonstrate that targeted integration of Cre cassette lead to faithful recapitulation of expression pattern of the endogenous promoter, and mRNA FISH, in addition to IHC, is an effective method for the analysis of the spatiotemporal gene expression patterns in the rat brain, alleviating the dependence on high quality antibodies that are often not available against rat proteins. The Th-Cre and the DAT-Cre rat lines express Cre in selective subsets of dopaminergic neurons and should be particularly useful for researches on Parkinson's disease. PMID:26886559

  9. Longitudinal changes in the dopamine transporter and cognition in suicide attempters with charcoal burning.

    PubMed

    Yang, Kai-Chun; Wang, Shyh-Jen; Hsieh, Wen-Chi; Lirng, Jiing-Feng; Yang, Chen-Chang; Deng, Jou-Fang; Lin, Chun-Lung; Chou, Yuan-Hwa

    2015-02-28

    Suicide with charcoal burning, which results in carbon monoxide (CO) poisoning, is common in Asia. This study was designed to elucidate associations between changes in the dopamine transporter (DAT) and cognitive function in patients following CO poisoning during a follow-up period of 6 months. Participants comprised 31 healthy controls (HCs) and 21 CO poisoning patients. Each subject underwent single photon emission computed tomography with [(99m)Tc] TRODAT-1 to measure DAT availability and completed a cognitive battery assessing attention, memory, and executive function. For CO poisoning patients, a second DAT measurement and repeated cognitive evaluations were performed 6 months later. At baseline, DAT availability over bilateral striatum in CO poisoning subjects was significantly lower than in HCs. After 6 months, there was no significant change of DAT availability in CO poisoning patients. CO poisoning patients also had worse cognitive performance in all domains compared with HCs at baseline. After 6 months, most cognitive functions were significantly improved, except for the Wisconsin Card Sorting Test (WCST), a measure of executive function. Interestingly, changes in the WCST were significantly correlated with changes in DAT availability during the 6-month follow-up period. The persistence of reduced DAT availability and its association with impaired performance on the WCST indicate a crucial role of DAT in the recovery of executive function following CO poisoning. PMID:25572798

  10. Individual differences in impulsive action and dopamine transporter function in rat orbitofrontal cortex.

    PubMed

    Yates, J R; Darna, M; Beckmann, J S; Dwoskin, L P; Bardo, M T

    2016-01-28

    Impulsivity, which can be subdivided into impulsive action and impulsive choice, is implicated as a factor underlying drug abuse vulnerability. Although previous research has shown that dopamine (DA) systems in prefrontal cortex are involved in impulsivity and substance abuse, it is not known if inherent variation in DA transporter (DAT) function contributes to impulsivity. The current study determined if individual differences in either impulsive action or impulsive choice are related to DAT function in orbitofrontal (OFC) and/or medial prefrontal cortex (mPFC). Rats were first tested both for impulsive action in a cued go/no-go task and for impulsive choice in a delay-discounting task. Following behavioral evaluation, in vitro [(3)H]DA uptake assays were performed in OFC and mPFC isolated from individual rats. Vmax in OFC, but not mPFC, was correlated with performance in the cued go/no-go task, with decreased OFC DAT function being associated with high impulsive action. In contrast, Vmax in OFC and mPFC was not correlated with performance in the delay-discounting task. The current results demonstrate that impulsive behavior in cued go/no-go performance is associated with decreased DAT function in OFC, suggesting that hyperdopaminergic tone in this prefrontal subregion mediates, at least in part, increased impulsive action. PMID:26608122

  11. Intermittent cocaine self-administration produces sensitization of stimulant effects at the dopamine transporter.

    PubMed

    Calipari, Erin S; Ferris, Mark J; Siciliano, Cody A; Zimmer, Benjamin A; Jones, Sara R

    2014-05-01

    Previous literature investigating neurobiological adaptations following cocaine self-administration has shown that high, continuous levels of cocaine intake (long access; LgA) results in reduced potency of cocaine at the dopamine transporter (DAT), whereas an intermittent pattern of cocaine administration (intermittent access; IntA) results in sensitization of cocaine potency at the DAT. Here, we aimed to determine whether these changes are specific to cocaine or translate to other psychostimulants. Psychostimulant potency was assessed by fast-scan cyclic voltammetry in brain slices containing the nucleus accumbens following IntA, short access, and LgA cocaine self-administration, as well as in brain slices from naive animals. We assessed the potency of amphetamine (a releaser), and methylphenidate (a DAT blocker, MPH). MPH was selected because it is functionally similar to cocaine and structurally related to amphetamine. We found that MPH and amphetamine potencies were increased following IntA, whereas neither was changed following LgA or short access cocaine self-administration. Therefore, whereas LgA-induced tolerance at the DAT is specific to cocaine as shown in previous work, the sensitizing effects of IntA apply to cocaine, MPH, and amphetamine. This demonstrates that the pattern with which cocaine is administered is important in determining the neurochemical consequences of not only cocaine effects but potential cross-sensitization/cross-tolerance effects of other psychostimulants as well. PMID:24566123

  12. Dopamine transporter availability in motor subtypes of de novo drug-naïve Parkinson's disease.

    PubMed

    Moccia, Marcello; Pappatà, Sabina; Picillo, Marina; Erro, Roberto; Coda, Anna Rita Daniela; Longo, Katia; Vitale, Carmine; Amboni, Marianna; Brunetti, Arturo; Capo, Giuseppe; Salvatore, Marco; Barone, Paolo; Pellecchia, Maria Teresa

    2014-11-01

    Tremor dominant (TD) and akinetic-rigid type (ART) are two motor subtypes of Parkinson's disease associated with different disease progression and neurochemical/neuropathological features. The role of presynaptic nigrostriatal dopaminergic damage is still controversial, poorly explored, and only assessed in medicated patients. In this study, we investigated with FP-CIT SPECT the striatal dopamine transporter (DAT) availability in drug-naïve PD patients with ART and TD phenotypes. Fifty-one de novo, drug-naïve patients with PD underwent FP-CIT SPECT studies. Patients were evaluated with Unified Parkinson's Disease Rating Scale (UPDRS) part III and Hoehn and Yahr scale (H&Y) and divided into ART (24/51) and TD (27/51) according to UPDRS part III. ART and TD patients were not different with regard to age, gender, and disease duration. However, compared to TD, ART patients presented higher UPDRS part III (p = 0.01) and H&Y (p = 0.02) and lower DAT availability in affected and unaffected putamen (p = 0.008 and p = 0.007, respectively), whereas no differences were found in caudate. Moreover, in the whole group of patients, rigidity and bradykinesia, but not tremor scores of UPDRS part III were significantly related to FP-CIT binding in the putamen. These results suggest that in newly diagnosed drug-naïve PD patients DAT availability might be different between ART and TD in relation to different disease severity. PMID:25119838

  13. Tissue Specific Expression of Cre in Rat Tyrosine Hydroxylase and Dopamine Active Transporter-Positive Neurons

    PubMed Central

    Liu, Zhenyi; Brown, Andrew; Fisher, Dan; Wu, Yumei; Warren, Joe; Cui, Xiaoxia

    2016-01-01

    The rat is a preferred model system over the mouse for neurological studies, and cell type-specific Cre expression in the rat enables precise ablation of gene function in neurons of interest, which is especially valuable for neurodegenerative disease modeling and optogenetics. Yet, few such Cre rats are available. Here we report the characterization of two Cre rats, tyrosine hydroxylase (TH)-Cre and dopamine active transporter (DAT or Slc6a3)-Cre, by using a combination of immunohistochemistry (IHC) and mRNA fluorescence in situ hybridization (FISH) as well as a fluorescent reporter for Cre activity. We detected Cre expression in expected neurons in both Cre lines. Interestingly, we also found that in Th-Cre rats, but not DAT-Cre rats, Cre is expressed in female germ cells, allowing germline excision of the floxed allele and hence the generation of whole-body knockout rats. In summary, our data demonstrate that targeted integration of Cre cassette lead to faithful recapitulation of expression pattern of the endogenous promoter, and mRNA FISH, in addition to IHC, is an effective method for the analysis of the spatiotemporal gene expression patterns in the rat brain, alleviating the dependence on high quality antibodies that are often not available against rat proteins. The Th-Cre and the DAT-Cre rat lines express Cre in selective subsets of dopaminergic neurons and should be particularly useful for researches on Parkinson’s disease. PMID:26886559

  14. Epigenetic Regulation of Dopamine Transporter mRNA Expression in Human Neuroblastoma Cells.

    PubMed

    Green, Ashley L; Hossain, Muhammad M; Tee, Siew C; Zarbl, Helmut; Guo, Grace L; Richardson, Jason R

    2015-07-01

    The dopamine transporter (DAT) is a key regulator of dopaminergic neurotransmission. As such, proper regulation of DAT expression is important to maintain homeostasis, and disruption of DAT expression can lead to neurobehavioral dysfunction. Based on genomic features within the promoter of the DAT gene, there is potential for DAT expression to be regulated through epigenetic mechanisms, including DNA methylation and histone acetylation. However, the relative contribution of these mechanisms to DAT expression has not been empirically determined. Using pharmacologic and genetic approaches, we demonstrate that inhibition of DNA methyltransferase (DNMT) activity increased DAT mRNA approximately 1.5-2 fold. This effect was confirmed by siRNA knockdown of DNMT1. Likewise, the histone deacetylase (HDAC) inhibitors valproate and butyrate also increased DAT mRNA expression, but the response was much more robust with expression increasing over tenfold. Genetic knockdown of HDAC1 by siRNA also increased DAT expression, but not to the extent seen with pharmacological inhibition, suggesting additional isoforms of HDAC or other targets may contribute to the observed effect. Together, these data identify the relative contribution of DNMTs and HDACs in regulating expression. These finding may aid in understanding the mechanistic basis for changes in DAT expression in normal and pathophysiological states. PMID:25963949

  15. Visualization of the Cocaine-Sensitive Dopamine Transporter with Ligand-Conjugated Quantum Dots

    PubMed Central

    2011-01-01

    The presynaptic dopamine (DA) transporter is responsible for DA inactivation following release and is a major target for the psychostimulants cocaine and amphetamine. Dysfunction and/or polymorphisms in human DAT (SLC6A3) have been associated with schizophrenia, bipolar disorder, Parkinson’s disease, and attention-deficit hyperactivity disorder (ADHD). Despite the clinical importance of DAT, many uncertainties remain regarding the transporter’s regulation, in part due to the poor spatiotemporal resolution of conventional methodologies and the relative lack of efficient DAT-specific fluorescent probes. We developed a quantum dot-based labeling approach that uses a DAT-specific, biotinylated ligand, 2-β-carbomethoxy-3-β-(4-fluorophenyl)tropane (IDT444), that can be bound by streptavidin-conjugated quantum dots. Flow cytometry and confocal microscopy were used to detect DAT in stably and transiently transfected mammalian cells. IDT444 is useful for quantum-dot-based fluorescent assays to monitor DAT expression, function, and plasma membrane trafficking in living cells as evidenced by the visualization of acute, protein-kinase-C (PKC)-dependent DAT internalization. PMID:22816024

  16. Persistent Drug-Induced Parkinsonism in Patients with Normal Dopamine Transporter Imaging

    PubMed Central

    Sunwoo, Mun Kyung; Oh, Jungsu S.; Kim, Jae Seung; Sohn, Young H.; Lee, Phil Hyu

    2016-01-01

    Functional neuroimaging for the dopamine transporter (DAT) is used to distinguish drug-induced parkinsonism (DIP) from subclinical Parkinson’s disease (PD). Although DIP patients who show a normal DAT image are expected to recover completely, some do not. We investigated whether these patients showed changes in striatal DAT activity using semi-quantitative analysis of 18F-FP-CIT PET data. DIP patients with visually normal DAT images were selected from medical records. The subjects were classified as patients who recovered partially (PR) or completely within 12 months (CR). The 18F-FP-CIT uptake in each striatal subregion was compared between the CR and the PR groups. In total, 41 and 9 patients of the CR and PR groups were assessed, respectively. The two patient groups were comparable in terms of clinical characteristics including age, sex, and severity of parkinsonism. From semi-quantitative analysis of the PET image, the PR patients showed a relatively lower ligand uptake in the ventral striatum, the anterior putamen and the posterior putamen compared with the CR patients. This result suggests that persistent DIP in patients with visually normal DAT imaging may be associated with subtle decrement of DAT activity. PMID:27294367

  17. Dopamine Transporter Genotype and Stimulant Dose-Response in Youth with Attention-Deficit/Hyperactivity Disorder

    PubMed Central

    Waldman, Irwin; Newcorn, Jeffrey; Bishop, Jeffrey; Kittles, Rick; Cook, Edwin H.

    2014-01-01

    Abstract Objectives: This study seeks to determine if variation in the dopamine transporter gene (SLC6A3/DAT1) moderates the dose-response effects of long-acting dexmethylphenidate (D-MPH) and mixed amphetamine salts (MAS) in children with attention-deficit/hyperactivity disorder (ADHD). Methods: Fifty-six children and adolescents (mean age=11.7±2.2) participated in a double-blind, two period crossover, dose-response study with a randomized placebo week in each 4 week drug period. Each period consisted of sequential week-long exposures to three dose levels (10, 20, 25–30 mg, depending upon weight) of D-MPH or MAS. Results: Doses of 10–20 mg of either D-MPH or MAS had little to no effect on hyperactivity-impulsivity and total ADHD symptom scores in subjects with the 9/9 genotype; this was in contrast to the dose-response curves of subjects with either the 10/10 or 10/9 genotype. Conclusions: ADHD youth with the 9/9 genotype may require higher stimulant doses to achieve adequate symptom control. PMID:24813374

  18. The influence of genetic variants on striatal dopamine transporter and D2 receptor binding after TBI.

    PubMed

    Wagner, Amy K; Scanlon, Joelle M; Becker, Carl R; Ritter, Anne C; Niyonkuru, Christian; Dixon, Clifton E; Conley, Yvette P; Price, Julie C

    2014-08-01

    Dopamine (DA) neurotransmission influences cognition and recovery after traumatic brain injury (TBI). We explored whether functional genetic variants affecting the DA transporter (DAT) and D2 receptor (DRD2) impacted in vivo dopaminergic binding with positron emission tomography (PET) using [(11)C]βCFT and [(11)C]raclopride. We examined subjects with moderate/severe TBI (N=12) ∼1 year post injury and similarly matched healthy controls (N=13). The variable number of tandem repeat polymorphism within the DAT gene and the TaqI restriction fragment length polymorphism near the DRD2 gene were assessed. TBI subjects had age-adjusted DAT-binding reductions in the caudate, putamen, and ventral striatum, and modestly increased D2 binding in ventral striatum versus controls. Despite small sample sizes, multivariate analysis showed lower caudate and putamen DAT binding among DAT 9-allele carriers and DRD2 A2/A2 homozygotes with TBI versus controls with the same genotype. Among TBI subjects, 9-allele carriers had lower caudate and putamen binding than 10/10 homozygotes. This PET study suggests a hypodopaminergic environment and altered DRD2 autoreceptor DAT interactions that may influence DA transmission after TBI. Future work will relate these findings to cognitive performance; future studies are required to determine how DRD2/DAT1 genotype and DA-ligand binding are associated with neurostimulant response and TBI recovery. PMID:24849661

  19. Molecular pharmacology of renal organic anion transporters.

    PubMed

    Van Aubel, R A; Masereeuw, R; Russel, F G

    2000-08-01

    Renal organic anion transport systems play an important role in the elimination of drugs, toxic compounds, and their metabolites, many of which are potentially harmful to the body. The renal proximal tubule is the primary site of carrier-mediated transport from blood to urine of a wide variety of anionic substrates. Recent studies have shown that organic anion secretion in renal proximal tubule is mediated by distinct sodium-dependent and sodium-independent transport systems. Knowledge of the molecular identity of these transporters and their substrate specificity has increased considerably in the past few years by cloning of various carrier proteins. However, a number of fundamental questions still have to be answered to elucidate the participation of the cloned transporters in the overall tubular secretion of anionic xenobiotics. This review summarizes the latest knowledge on molecular and pharmacological properties of renal organic anion transporters and homologs, with special reference to their nephron and plasma membrane localization, transport characteristics, and substrate and inhibitor specificity. A number of the recently cloned transporters, such as the p-aminohippurate/dicarboxylate exchanger OAT1, the anion/sulfate exchanger SAT1, the peptide transporters PEPT1 and PEPT2, and the nucleoside transporters CNT1 and CNT2, are key proteins in organic anion handling that possess the same characteristics as has been predicted from previous physiological studies. The role of other cloned transporters, such as MRP1, MRP2, OATP1, OAT-K1, and OAT-K2, is still poorly characterized, whereas the only information that is available on the homologs OAT2, OAT3, OATP3, and MRP3-6 is that they are expressed in the kidney, but their localization, not to mention their function, remains to be elucidated. PMID:10919840

  20. Charge Redistribution and Transport in Molecular Contacts

    NASA Astrophysics Data System (ADS)

    Corso, Martina; Ondráček, Martin; Lotze, Christian; Hapala, Prokop; Franke, Katharina J.; Jelínek, Pavel; Pascual, J. Ignacio

    2015-09-01

    The forces between two single molecules brought into contact, and their connection with charge transport through the molecular junction, are studied here using non contact AFM, STM, and density functional theory simulations. A carbon monoxide molecule approaching an acetylene molecule (C2 H2 ) initially feels weak attractive electrostatic forces, partly arising from charge reorganization in the presence of molecular . We find that the molecular contact is chemically passive, and protects the electron tunneling barrier from collapsing, even in the limit of repulsive forces. However, we find subtle conductance and force variations at different contacting sites along the C2 H2 molecule attributed to a weak overlap of their respective frontier orbitals.

  1. Charge transport network dynamics in molecular aggregates.

    PubMed

    Jackson, Nicholas E; Chen, Lin X; Ratner, Mark A

    2016-08-01

    Due to the nonperiodic nature of charge transport in disordered systems, generating insight into static charge transport networks, as well as analyzing the network dynamics, can be challenging. Here, we apply time-dependent network analysis to scrutinize the charge transport networks of two representative molecular semiconductors: a rigid n-type molecule, perylenediimide, and a flexible p-type molecule, [Formula: see text] Simulations reveal the relevant timescale for local transfer integral decorrelation to be [Formula: see text]100 fs, which is shown to be faster than that of a crystalline morphology of the same molecule. Using a simple graph metric, global network changes are observed over timescales competitive with charge carrier lifetimes. These insights demonstrate that static charge transport networks are qualitatively inadequate, whereas average networks often overestimate network connectivity. Finally, a simple methodology for tracking dynamic charge transport properties is proposed. PMID:27439871

  2. Dopamine transporters are markedly reduced in Lesch-Nyhan disease in vivo.

    PubMed Central

    Wong, D F; Harris, J C; Naidu, S; Yokoi, F; Marenco, S; Dannals, R F; Ravert, H T; Yaster, M; Evans, A; Rousset, O; Bryan, R N; Gjedde, A; Kuhar, M J; Breese, G R

    1996-01-01

    Dopamine (DA) deficiency has been implicated in Lesch-Nyhan disease (LND), a genetic disorder that is characterized by hyperuricemia, choreoathetosis, dystonia, and compulsive self-injury. To establish that DA deficiency is present in LND, the ligand WIN-35,428, which binds to DA transporters, was used to estimate the density of DA-containing neurons in the caudate and putamen of six patients with classic LND. Comparisons were made with 10 control subjects and 3 patients with Rett syndrome. Three methods were used to quantify the binding of the DA transporter so that its density could be estimated by a single dynamic positron emission tomography study. These approaches included the caudate- or putamen-to-cerebellum ratio of ligand at 80-90 min postinjection, kinetic analysis of the binding potential [Bmax/(Kd x Vd)] using the assumption of equal partition coefficients in the striatum and the cerebellum, and graphical analysis of the binding potential. Depending on the method of analysis, a 50-63% reduction of the binding to DA transporters in the caudate, and a 64-75% reduction in the putamen of the LND patients was observed compared to the normal control group. When LND patients were compared to Rett syndrome patients, similar reductions were found in the caudate (53-61%) and putamen (67-72%) in LND patients. Transporter binding in Rett syndrome patients was not significantly different from the normal controls. Finally, volumetric magnetic resonance imaging studies detected a 30% reduction in the caudate volume of LND patients. To ensure that a reduction in the caudate volume would not confound the results, a rigorous partial volume correction of the caudate time activity curve was performed. This correction resulted in an even greater decrease in the caudate-cerebellar ratio in LND patients when contrasted to controls. To our knowledge, these findings provide the first in vivo documentation of a dopaminergic reduction in LND and illustrate the role of positron

  3. Controlling molecular transport in minimal emulsions

    NASA Astrophysics Data System (ADS)

    Gruner, Philipp; Riechers, Birte; Semin, Benoît; Lim, Jiseok; Johnston, Abigail; Short, Kathleen; Baret, Jean-Christophe

    2016-01-01

    Emulsions are metastable dispersions in which molecular transport is a major mechanism driving the system towards its state of minimal energy. Determining the underlying mechanisms of molecular transport between droplets is challenging due to the complexity of a typical emulsion system. Here we introduce the concept of `minimal emulsions', which are controlled emulsions produced using microfluidic tools, simplifying an emulsion down to its minimal set of relevant parameters. We use these minimal emulsions to unravel the fundamentals of transport of small organic molecules in water-in-fluorinated-oil emulsions, a system of great interest for biotechnological applications. Our results are of practical relevance to guarantee a sustainable compartmentalization of compounds in droplet microreactors and to design new strategies for the dynamic control of droplet compositions.

  4. Controlling molecular transport in minimal emulsions

    PubMed Central

    Gruner, Philipp; Riechers, Birte; Semin, Benoît; Lim, Jiseok; Johnston, Abigail; Short, Kathleen; Baret, Jean-Christophe

    2016-01-01

    Emulsions are metastable dispersions in which molecular transport is a major mechanism driving the system towards its state of minimal energy. Determining the underlying mechanisms of molecular transport between droplets is challenging due to the complexity of a typical emulsion system. Here we introduce the concept of ‘minimal emulsions', which are controlled emulsions produced using microfluidic tools, simplifying an emulsion down to its minimal set of relevant parameters. We use these minimal emulsions to unravel the fundamentals of transport of small organic molecules in water-in-fluorinated-oil emulsions, a system of great interest for biotechnological applications. Our results are of practical relevance to guarantee a sustainable compartmentalization of compounds in droplet microreactors and to design new strategies for the dynamic control of droplet compositions. PMID:26797564

  5. Controlling molecular transport in minimal emulsions.

    PubMed

    Gruner, Philipp; Riechers, Birte; Semin, Benoît; Lim, Jiseok; Johnston, Abigail; Short, Kathleen; Baret, Jean-Christophe

    2016-01-01

    Emulsions are metastable dispersions in which molecular transport is a major mechanism driving the system towards its state of minimal energy. Determining the underlying mechanisms of molecular transport between droplets is challenging due to the complexity of a typical emulsion system. Here we introduce the concept of 'minimal emulsions', which are controlled emulsions produced using microfluidic tools, simplifying an emulsion down to its minimal set of relevant parameters. We use these minimal emulsions to unravel the fundamentals of transport of small organic molecules in water-in-fluorinated-oil emulsions, a system of great interest for biotechnological applications. Our results are of practical relevance to guarantee a sustainable compartmentalization of compounds in droplet microreactors and to design new strategies for the dynamic control of droplet compositions. PMID:26797564

  6. Loss of striatal cannabinoid CB1 receptor function in attention-deficit / hyperactivity disorder mice with point-mutation of the dopamine transporter.

    PubMed

    Castelli, Maura; Federici, Mauro; Rossi, Silvia; De Chiara, Valentina; Napolitano, Francesco; Studer, Valeria; Motta, Caterina; Sacchetti, Lucia; Romano, Rosaria; Musella, Alessandra; Bernardi, Giorgio; Siracusano, Alberto; Gu, Howard H; Mercuri, Nicola B; Usiello, Alessandro; Centonze, Diego

    2011-11-01

    Abnormal dopamine (DA) transmission in the striatum plays a pivotal role in attention-deficit/hyperactivity disorder (ADHD). As striatal DA signalling modulates the endocannabinoid system (ECS), the present study was aimed at investigating cannabinoid CB1 receptor (CB1R) function in a model of ADHD obtained by triple point-mutation in the dopamine transporter (DAT) gene in mice, making them insensitive to cocaine [DAT cocaine-insensitive (DAT-CI) mice]. DAT-CI mice had a marked hyperactive phenotype, and neurophysiological recordings revealed that the sensitivity of CB1Rs controlling GABA-mediated synaptic currents [CB1Rs((GABA)) ] in the striatum was completely lost. In contrast, CB1Rs modulating glutamate transmission [CB1Rs((Glu)) ], and GABA(B) receptors were not affected in this model of ADHD. In DAT-CI mice, the blockade of CB1R((GABA)) function was complete even after cocaine or environmental manipulations activating the endogenous DA-dependent reward system, which are known to sensitize these receptors in control animals. Conversely, the hedonic property of sucrose was intact in DAT-CI mice, indicating normal sweet perception in these animals. Our results point to CB1Rs as novel molecular players in ADHD, and suggest that therapeutic strategies aimed at interfering with the ECS might prove effective in this disorder. PMID:22034972

  7. Age-related changes in dopamine transporters and accumulation of 3-nitrotyrosine in rhesus monkey midbrain dopamine neurons: Relevance in selective neuronal vulnerability to degeneration

    PubMed Central

    Kanaan, N. M.; Kordower, J. H.; Collier, T. J.

    2012-01-01

    Aging is the strongest risk factor for developing Parkinson’s disease (PD). There is a preferential loss of dopamine (DA) neurons in the ventral tier of the substantia nigra (vtSN) compared to the dorsal tier and ventral tegmental area (VTA) in PD. Examining age-related and region-specific differences in DA neurons represents a means of identifying factors potentially involved in vulnerability or resistance to degeneration. Nitrative stress is among the factors potentially underlying DA neuron degeneration. We studied the relationship between 3-nitrotyrosine (3NT; a marker of nitrative damage) and DA transporters [DA transporter (DAT) and vesicular monoamine transporter-2 (VMAT)] during aging in DA subregions of rhesus monkeys. The percentage of DA neurons containing 3NT increased significantly only in the vtSN with advancing age, and the vtSN had a greater percentage of 3NT-positive neurons when compared to the VTA. The relationship between 3NT and DA transporters was determined by measuring fluorescence intensity of 3NT, DAT and VMAT staining. 3NT intensity increased with advancing age in the vtSN. Increased DAT, VMAT and DAT/VMAT ratios were associated with increased 3NT in individual DA neurons. These results suggest nitrative damage accumulates in midbrain DA neurons with advancing age, an effect exacerbated in the vulnerable vtSN. The capacity of a DA neuron to accumulate more cytosolic DA, as inferred from DA transporter expression, is related to accumulation of nitrative damage. These findings are consistent with a role for aging-related accrual of nitrative damage in the selective vulnerability of vtSN neurons to degeneration in PD. PMID:18598263

  8. Pharmacological and Behavioral Characterization of D-473, an Orally Active Triple Reuptake Inhibitor Targeting Dopamine, Serotonin and Norepinephrine Transporters

    PubMed Central

    Dutta, Aloke K.; Santra, Soumava; Sharma, Horrick; Voshavar, Chandrashekhar; Xu, Liping; Mabrouk, Omar; Antonio, Tamara; Reith, Maarten E. A.

    2014-01-01

    Major depressive disorder (MDD) is a debilitating disease affecting a wide cross section of people around the world. The current therapy for depression is less than adequate and there is a considerable unmet need for more efficacious treatment. Dopamine has been shown to play a significant role in depression including production of anhedonia which has been one of the untreated symptoms in MDD. It has been hypothesized that drugs acting at all three monoamine transporters including dopamine transporter should provide more efficacious antidepressants activity. This has led to the development of triple reuptake inhibitor D-473 which is a novel pyran based molecule and interacts with all three monoamine transporters. The monoamine uptake inhibition activity in the cloned human transporters expressed in HEK-293 cells (70.4, 9.18 and 39.7 for DAT, SERT and NET, respectively) indicates a serotonin preferring triple reuptake inhibition profile for this drug. The drug D-473 exhibited good brain penetration and produced efficacious activity in rat forced swim test under oral administration. The optimal efficacy dose did not produce any locomotor activation. Microdialysis experiment demonstrated that systemic administration of D-473 elevated extracellular level of the three monoamines DA, 5-HT, and NE efficaciously in the dorsal lateral striatum (DLS) and the medial prefrontal cortex (mPFC) area, indicating in vivo blockade of all three monoamine transporters by D-473. Thus, the current biological data from D-473 indicate potent antidepressant activity of the molecule. PMID:25427177

  9. Ex vivo identification of protein-protein interactions involving the dopamine transporter.

    PubMed

    Hadlock, Gregory C; Nelson, Chad C; Baucum, Anthony J; Hanson, Glen R; Fleckenstein, Annette E

    2011-03-30

    The dopamine (DA) transporter (DAT) is a key regulator of dopaminergic signaling as it mediates the reuptake of extrasynaptic DA and thereby terminates dopaminergic signaling. Emerging evidence indicates that DAT function is influenced through interactions with other proteins. The current report describes a method to identify such interactions following DAT immunoprecipitation from a rat striatal synaptosomal preparation. This subcellular fraction was selected since DAT function is often determined ex vivo by measuring DA uptake in this preparation and few reports investigating DAT-protein interactions have utilized this preparation. Following SDS-PAGE and colloidal Coomassie staining, selected protein bands from a DAT-immunoprecipitate were excised, digested with trypsin, extracted, and analyzed by liquid chromatography tandem mass spectrometry (LC/MS/MS). From the analysis of the tryptic peptides, several proteins were identified including DAT, Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) β, CaMKII δ, protein kinase C (PKC) β, and PKC γ. Co-immunoprecipitation of PKC, CaMKII, and protein interacting with C kinase-1 with DAT was confirmed by Western blotting. Thus, the present study highlights a method to immunoprecipitate DAT and to identify co-immunoprecipitating proteins using LC/MS/MS and Western blotting. This method can be utilized to evaluate DAT protein-protein interactions but also to assess interactions involving other synaptic proteins. Ex vivo identification of protein-protein interactions will provide new insight into the function and regulation of a variety of synaptic, membrane-associated proteins, including DAT. PMID:21291912

  10. In vivo comparison of the reinforcing and dopamine transporter effects of local anesthetics in rhesus monkeys.

    PubMed

    Wilcox, Kristin M; Kimmel, Heather L; Lindsey, Kimberly P; Votaw, John R; Goodman, Mark M; Howell, Leonard L

    2005-12-15

    Dopaminergic mechanisms are thought to play a central role in the reinforcing effects of cocaine. Similar to cocaine, other local anesthetics bind to the dopamine transporter (DAT) and inhibit DA uptake in rodent and monkey brain. Additionally, local anesthetics are self-administered in rhesus monkeys, indicative of abuse liability. The present study examined the reinforcing and DAT effects of the local anesthetics dimethocaine, procaine and cocaine using in vivo techniques. Monkeys were trained to respond under a second-order schedule for i.v. cocaine administration (0.10 or 0.30 mg/kg/infusion). When responding was stable, dimethocaine (0.030-1.7 mg/kg/ infusion) or procaine (0.10-10 mg/kg/ infusion) was substituted for the cocaine training dose. Dimethocaine administration produced higher response rates compared with that of procaine, and was a more potent reinforcer. Drug effects on behavior were related to DAT occupancy in monkey striatum during neuroimaging with positron emission tomography (PET). DAT occupancy was determined by displacement of 8-(2-[(18)F]fluroethyl)2beta-carbomethoxy-3beta-(4-chlorophenyl)nortropane (FECNT). DAT occupancy was between 66 and 82% and <10-41% for doses of dimethocaine and procaine that maintained maximum response rates, respectively. Finally, in vivo microdialysis in awake subjects determined drug-induced changes in extracellular DA in the caudate nucleus. There was close correspondence between peak increases in DA and DAT occupancy. Overall, reinforcing effects were consistent with DAT effects determined with in vivo techniques. The results further support a role for the DAT in the abuse liability of local anesthetics. PMID:16206183

  11. Cocaine occupancy of sigma1 receptors and dopamine transporters in mice.

    PubMed

    Lever, John R; Fergason-Cantrell, Emily A; Watkinson, Lisa D; Carmack, Terry L; Lord, Sarah A; Xu, Rong; Miller, Dennis K; Lever, Susan Z

    2016-03-01

    Activation of sigma1 (σ1) receptors contributes to the behavioral and toxic effects of (-)-cocaine. We studied a key step, the ability of (-)-cocaine to occupy σ1 receptors in vivo, using CD-1(®) mice and the novel radioligand [(125) I]E-N-1-(3'-iodoallyl)-N'-4-(3",4"-dimethoxyphenethyl)-piperazine ([(125) I]E-IA-DM-PE-PIPZE). (-)-Cocaine displayed an ED50 of 68 μmol/kg for inhibition of specific radioligand binding in whole brain, with values between 73 and 80 μmol/kg for heart, lung, and spleen. For comparison, an ED50 of 26 μmol/kg for (-)-cocaine occupancy of striatal dopamine transporters (DAT) was determined by inhibition of [(125) I]3β-(4-iodophenyl)tropan-2β-carboxylic acid isopropyl ester ([(125) I]RTI-121) binding. A chief finding is the relatively small potency difference between (-)-cocaine occupancy of σ1 receptors and the DAT, although the DAT occupancy is likely underestimated. Interactions of (-)-cocaine with σ1 receptors were assessed further using [(125) I]E-IA-DM-PE-PIPZE for regional cerebral biodistribution studies and quantitative ex vivo autoradiography of brain sections. (-)-Cocaine binding to cerebral σ1 receptors proved directly proportional to the relative site densities known for the brain regions. Nonradioactive E-IA-DM-PE-PIPZE gave an ED50 of 0.23 μmol/kg for occupancy of cerebral σ1 receptors, and a 3.16 μmol/kg (i.p.) dose attenuated (-)-cocaine-induced locomotor hyperactivity by 30%. This effect did not reach statistical significance, but suggests that E-IA-DM-PE-PIPZE is a probable σ1 receptor antagonist. As groundwork for the in vivo studies, we used standard techniques in vitro to determine ligand affinities, site densities, and pharmacological profiles for the σ1 and σ2 receptors expressed in CD-1(®) mouse brain. PMID:26618331

  12. Methylone-induced hyperthermia and lethal toxicity: role of the dopamine and serotonin transporters.

    PubMed

    Piao, Ying-Shan; Hall, Frank Scott; Moriya, Yuki; Ito, Miki; Ohara, Arihisa; Kikura-Hanajiri, Ruri; Goda, Yukihiro; Lesch, Klaus-Peter; Murphy, Dennis L; Uhl, George R; Sora, Ichiro

    2015-06-01

    Methylone (2-methylamino-1-[3,4-methylenedioxy-phenyl]propan-1-one), an amphetamine analog, has emerged as a popular drug of abuse worldwide. Methylone induces hyperthermia, which is thought to contribute toward the lethal consequences of methylone overdose. Methylone has been assumed to induce hyperthermic effects through inhibition of serotonin and/or dopamine transporters (SERT and DAT, respectively). To examine the roles of each of these proteins in methylone-induced toxic effects, we used SERT and DAT knockout (KO) mice and assessed the hyperthermic and lethal effects caused by a single administration of methylone. Methylone produced higher rates of lethal toxicity compared with other amphetamine analogs in wild-type mice. Compared with wild-type mice, lethality was significantly lower in DAT KO mice, but not in SERT KO mice. By contrast, only a slight diminution in the hyperthermic effects of methylone was observed in DAT KO mice, whereas a slight enhancement of these effects was observed in SERT KO mice. Administration of the selective D1 receptor antagonist SCH 23390 and the D2 receptor antagonist raclopride reduced methylone-induced hyperthermia, but these drugs also had hypothermic effects in saline-treated mice, albeit to a smaller extent than the effects observed in methylone-treated mice. In contradistinction to 3,4-methylenedioxymethamphetamine, which induces its toxicity through SERT and DAT, these data indicate that DAT, but not SERT, is strongly associated with the lethal toxicity produced by methylone, which did not seem to be dependent on the hyperthermic effects of methylone. DAT is therefore a strong candidate molecule for interventions aimed at preventing acute neurotoxic and lethal effects of methylone. PMID:25794333

  13. Differential effects of dopamine transporter inhibitors in the rodent Iowa Gambling Task: Relevance to mania

    PubMed Central

    van Enkhuizen, Jordy; Geyer, Mark A.; Young, Jared W.

    2012-01-01

    Rationale The Iowa Gambling Task (IGT) can be used to quantify impulsive and risky choice behaviors in psychiatric patients, e.g. Bipolar Disorder (BD) sufferers. Although developing treatments for these behaviors is important, few predictive animal models exist. Inhibition of the dopamine transporter (DAT) can model profiles of altered motor activity and exploration seen in patients with BD. The effect of DAT inhibition on impulsive choices related to BD has received limited study however. We used a rodent IGT to elucidate the effects of similarly acting drugs on risky choice behavior. Objectives We hypothesized that 1) C57BL/6 mice could adopt the ‘safe’ choice options in the IGT and 2) DAT inhibition would alter risk preference. Methods Mice were trained in the IGT to a stable risk-preference and then administered the norepinephrine/DAT inhibitor amphetamine, or the more selective DAT inhibitors modafinil or GBR12909. Results Mice developed a preference for the ‘safe’ option, which was potentiated by amphetamine administration. GBR12909 or modafinil administration increased motor impulsivity, motivation significantly, and risk preference subtly. Conclusions The rodent IGT can measure different impulse-related behaviors and differentiate similarly acting BD-related drugs. The contrasting effects of amphetamine and modafinil in mice are similar to effects in rats and humans in corresponding IGT tasks, supporting the translational validity of the task. GBR12909 and modafinil elicited similar behaviors in the IGT, likely through a shared mechanism. Future studies using a within-session IGT are warranted to confirm the suitability of DAT inhibitors to model risk-preference in BD. PMID:22945515

  14. Absence of age-related dopamine transporter loss in current cocaine abusers

    SciTech Connect

    Wang, G.J.; Volkow, N.D.; Fischman, M.

    1997-05-01

    The brain dopamine (DA) system appears to play a crucial role in the reinforcing properties of cocaine. Using PET we had previously shown significant decreases in DA D2 receptors but no changes in DA transporters (DAT) in detoxified cocaine abusers (>1 month after last cocaine use). This study evaluates DAT availability in current cocaine abusers (15 male and 5 female; age = 36.2{+-}5.3 years old) using PET and [C-11]cocaine, as a DAT ligand, and compares it to that in 18 male and 2 female age matched normal controls. Cocaine abusers had a history of abusing 4.2{+-}2.8 gm /week of cocaine for an average of 11.0{+-}4.9 years and their last use of cocaine was 5.4{+-}8 days prior to PET study. DAT availability was obtained using the ratio of the distribution volume in the region of interest (caudate, pulamen) to that in cerebellum which is a function of Bmax./Kd.+1. DAT availability in cocaine abusers did not differ to that in normals (N) (C= 1.78{+-}0.14, N= 1.77{+-}0.13). In addition, there were no differences between the groups in the distribution volume or the Kl (plasma to brain transfer constant) measures for [C-11]cocaine. However, in the normals but not in the abusers striatal DAT availability decreased with age (C: r = -0.07, p = 0.76; N: r = -0.55, p < 0.01). Though this study fails to show group differences in DAT availability between normals and current cocaine abusers it indicates a blunting of the age-related decline in DAT availability in the cocaine abusers. Future studies in older cocaine abusers at different time after detoxification arc required in order to assess if cocaine slows the loss of DAT with age or whether these changes reflect compensation to increased DAT blockade and recover with detoxification.

  15. Locomotor hyperactivity in 14-3-3ζ KO mice is associated with dopamine transporter dysfunction

    PubMed Central

    Ramshaw, H; Xu, X; Jaehne, E J; McCarthy, P; Greenberg, Z; Saleh, E; McClure, B; Woodcock, J; Kabbara, S; Wiszniak, S; Wang, Ting-Yi; Parish, C; van den Buuse, M; Baune, B T; Lopez, A; Schwarz, Q

    2013-01-01

    Dopamine (DA) neurotransmission requires a complex series of enzymatic reactions that are tightly linked to catecholamine exocytosis and receptor interactions on pre- and postsynaptic neurons. Regulation of dopaminergic signalling is primarily achieved through reuptake of extracellular DA by the DA transporter (DAT) on presynaptic neurons. Aberrant regulation of DA signalling, and in particular hyperactivation, has been proposed as a key insult in the presentation of schizophrenia and related neuropsychiatric disorders. We recently identified 14-3-3ζ as an essential component of neurodevelopment and a central risk factor in the schizophrenia protein interaction network. Our analysis of 14-3-3ζ-deficient mice now shows that baseline hyperactivity of knockout (KO) mice is rescued by the antipsychotic drug clozapine. 14-3-3ζ KO mice displayed enhanced locomotor hyperactivity induced by the DA releaser amphetamine. Consistent with 14-3-3ζ having a role in DA signalling, we found increased levels of DA in the striatum of 14-3-3ζ KO mice. Although 14-3-3ζ is proposed to modulate activity of the rate-limiting DA biosynthesis enzyme, tyrosine hydroxylase (TH), we were unable to identify any differences in total TH levels, TH localization or TH activation in 14-3-3ζ KO mice. Rather, our analysis identified significantly reduced levels of DAT in the absence of notable differences in RNA or protein levels of DA receptors D1–D5. Providing insight into the mechanisms by which 14-3-3ζ controls DAT stability, we found a physical association between 14-3-3ζ and DAT by co-immunoprecipitation. Taken together, our results identify a novel role for 14-3-3ζ in DA neurotransmission and provide support to the hyperdopaminergic basis of pathologies associated with schizophrenia and related disorders. PMID:24301645

  16. Altered corticostriatal neurotransmission and modulation in dopamine transporter knock-down mice.

    PubMed

    Wu, Nanping; Cepeda, Carlos; Zhuang, Xiaoxi; Levine, Michael S

    2007-07-01

    Dopamine (DA) modulates glutamate neurotransmission in the striatum. Abnormal DA modulation has been implicated in neurological and psychiatric disorders. The development of DA transporter knock-down (DAT-KD) mice has permitted modeling of these disorders and has shed new light on DA modulation. DAT-KD mice exhibit increased extracellular DA, hyperactivity, and alterations in habituation. We used whole cell patch-clamp recordings from visually identified striatal neurons in slices to examine the effects of DAT-KD on corticostriatal transmission. Electrophysiological recordings from medium-sized spiny neurons in the dorsal striatum revealed alterations in both amplitude and frequency, of spontaneous glutamate receptor-mediated synaptic currents in cells from DAT-KD mice. Furthermore, kinetic analyses revealed that these currents had shorter half-amplitude durations and faster decay times. In contrast, GABA-receptor-mediated synaptic currents were not altered. Striatal neurons from DAT-KD mice also responded differently to amphetamine, cocaine, and DA D2-receptor agonists or antagonists compared with wildtype (WT) littermate controls. In WTs amphetamine and cocaine reduced the frequency of spontaneous glutamate currents and these effects appeared to be mediated by activation of D2 receptors. In contrast, in DAT-KD mice either no changes or only small increases in frequency occurred. D2-receptor agonists or antagonists also had opposing effects in WT and DAT-KD mice. Together, these results indicate that chronically increased extracellular DA produces long-lasting changes in corticostriatal communication that may be mediated by changes in D2-receptor function. These findings have implications for understanding mechanisms underlying attention deficit hyperactivity disorder and Tourette's syndrome and may provide insights into novel therapeutic approaches. PMID:17522168

  17. GM1 Ganglioside in Parkinson’s Disease: Pilot Study of Effects on Dopamine Transporter Binding

    PubMed Central

    Schneider, Jay S.; Cambi, Franca; Gollomp, Stephen M.; Kuwabara, Hiroto; Brašić, James R.; Leiby, Benjamin; Sendek, Stephanie; Wong, Dean F.

    2015-01-01

    Objective GM1 ganglioside has been suggested as a treatment for Parkinson’s disease (PD), potentially having symptomatic and disease modifying effects. The current pilot imaging study was performed to examine effects of GM1 on dopamine transporter binding, as a surrogate measure of disease progression, studied longitudinally. Methods Positron emission tomography (PET) imaging data were obtained from a subset of subjects enrolled in a delayed start clinical trial of GM1 in PD1: 15 Early-start (ES) subjects, 14 Delayed-start (DS) subjects, and 11 Comparison (standard-of-care) subjects. Treatment subjects were studied over a 2.5 year period while Comparison subjects were studied over 2 years. Dynamic PET scans were performed over 90 minutes following injection of [11C]methylphenidate. Regional values of binding potential (BPND) were analyzed for several striatal volumes of interest. Results Clinical results for this subset of subjects were similar to those previously reported for the larger study group. ES subjects showed early symptomatic improvement and slow symptom progression over the study period. DS and Comparison subjects were initially on the same symptom progression trajectory but diverged once DS subjects received GM1 treatment. Imaging results showed significant slowing of BPND loss in several striatal regions in GM1-treated subjects and in some cases, an increased BPND in some striatal regions was detected after GM1 use. Interpretation Results of this pilot imaging study provide additional data to suggest a potential disease modifying effect of GM1 on PD. These results need to be confirmed in a larger number of subjects. PMID:26099170

  18. Reduced Dopamine Transporter Functioning Induces High-Reward Risk-Preference Consistent with Bipolar Disorder

    PubMed Central

    van Enkhuizen, Jordy; Henry, Brook L; Minassian, Arpi; Perry, William; Milienne-Petiot, Morgane; Higa, Kerin K; Geyer, Mark A; Young, Jared W

    2014-01-01

    Individuals with bipolar disorder (BD) exhibit deleterious decision making, negatively impacting their lives. Such aberrant decision making can be quantified using the Iowa Gambling Task (IGT), which requires choosing between advantageous and disadvantageous options based on different reward/punishment schedules. The mechanisms underlying this behavioral deficit are unknown, but may include the reduced dopamine transporter (DAT) functioning reported in BD patients. Using both human and mouse IGTs, we tested whether reduced DAT functioning would recreate patterns of deficient decision making of BD patients. We assessed the IGT performance of 16 BD subjects (7 female) and 17 healthy control (HC) subjects (12 female). We recorded standard IGT performance measures and novel post-reward and post-punishment decision-making strategies. We characterized a novel single-session mouse IGT using C57BL/6J mice (n=44). The BD and HC IGT performances were compared with the effects of chronic (genetic knockdown (KD; n=31) and wild-type (n=28) mice) and acute (C57BL/6J mice (n=89) treated with the DAT inhibitor GBR12909) reductions of DAT functioning in mice performing this novel IGT. BD patients exhibited impaired decision making compared with HC subjects. Both the good-performing DAT KD and GBR12909-treated mice exhibited poor decision making in the mouse IGT. The deficit of each population was driven by high-reward sensitivity. The single-session mouse IGT measures dynamic risk-based decision making similar to humans. Chronic and acute reductions of DAT functioning in mice impaired decision-making consistent with poor IGT performance of BD patients. Hyperdopaminergia caused by reduced DAT may impact poor decision making in BD patients, which should be confirmed in future studies. PMID:25005251

  19. Neural correlates of attentional bias for smoking cues: modulation by variance in the dopamine transporter gene

    PubMed Central

    Wetherill, Reagan R.; Jagannathan, Kanchana; Lohoff, Falk W.; Ehrman, Ronald; O’Brien, Charles P.; Childress, Anna Rose; Franklin, Teresa R.

    2014-01-01

    Cigarette-dependent smokers automatically and involuntarily orient attention towards smoking cues (SCs). This attentional bias is clinically significant, as it may contribute to relapse. Thus, identifying neural and genetic correlates of attentional bias is critical for improving interventions. Our previous studies show that the dopamine transporter (DAT) SLC6A3 genotype exerts profound effects on limbic responses to SCs. One potential mechanism underlying these effects is greater attentional bias for SCs. Here, we explored associations between attentional bias for SCs and neural responses to SCs among ‘sated’ smokers genotyped for the SLC6A3 polymorphism. Pseudo-Continuous arterial spin-labeled (pCASL) perfusion fMR images were acquired during SC exposure in 35 smokers genotyped for the SLC6A3 variable number of tandem repeats (VNTR) polymorphism (n=16, 9-repeats; n=19,10/10-repeats). Participants completed a visual dot-probe attentional bias task, which contained pictures of smoking and non-smoking pictures, to examine whether genetic variation in DAT influences attentional bias and to investigate relationships between attentional bias and neural responses to SCs. Although attentional bias to smoking pictures was not significantly different between 9-repeats and 10/10-repeats, 9-repeats showed a positive correlation between attentional bias and increased SC-induced brain activity in the amygdala; whereas, 10/10-repeats showed an inverse correlation in the medial orbitofrontal cortex (mOFC). In group comparisons, 9-repeats exhibited positive correlations between attentional bias and SCs in the mOFC and amygdala, relative to 10/10-repeats. Findings suggest that genetic variation in the DAT gene influences brain responses associated with attentional bias; thus, providing additional support for a SC-vulnerable endophenotype. PMID:23061530

  20. 3,4-Methylenedioxypyrovalerone prevents while methylone enhances methamphetamine-induced damage to dopamine nerve endings: β-ketoamphetamine modulation of neurotoxicity by the dopamine transporter

    PubMed Central

    Anneken, John H.; Angoa-Pérez, Mariana; Kuhn, Donald M.

    2016-01-01

    Methylone, 3,4-methylenedioxypyrovalerone (MDPV), and mephedrone are psychoactive ingredients of ‘bath salts’ and their abuse represents a growing public health care concern. These drugs are cathinone derivatives and are classified chemically as β-ketoamphetamines. Because of their close structural similarity to the amphetamines, methylone, MDPV, and mephedrone share most of their pharmacological, neurochemical, and behavioral properties. One point of divergence in their actions is the ability to cause damage to the CNS. Unlike methamphetamine, the β-ketoamphetamines do not damage dopamine (DA) nerve endings. However, mephedrone has been shown to significantly accentuate methamphetamine neurotoxicity. Bath salt formulations contain numerous different psychoactive ingredients, and individuals who abuse bath salts also coabuse other illicit drugs. Therefore, we have evaluated the effects of methylone, MDPV, mephedrone, and methamphetamine on DA nerve endings. The β-ketoamphetamines alone or in all possible two-drug combinations do not result in damage to DA nerve endings but do cause hyperthermia. MDPV completely protects against the neurotoxic effects of methamphetamine while methylone accentuates it. Neither MDPV nor methylone attenuates the hyperthermic effects of methamphetamine. The potent neuroprotective effects of MDPV extend to amphetamine-, 3,4-methylenedioxymethamphetamine-, and MPTP-induced neurotoxicity. These results indicate that β-ketoamphetamine drugs that are non-substrate blockers of the DA transporter (i.e., MDPV) protect against methamphetamine neurotoxicity, whereas those that are substrates for uptake by the DA transporter and which cause DA release (i.e., methylone, mephedrone) accentuate neurotoxicity. PMID:25626880

  1. Effects of combined dopamine and serotonin transporter inhibitors on cocaine self-administration in rhesus monkeys.

    PubMed

    Howell, Leonard L; Carroll, F Ivy; Votaw, John R; Goodman, Mark M; Kimmel, Heather L

    2007-02-01

    Dopamine transporter (DAT) inhibitors may represent a promising class of drugs in the development of cocaine pharmacotherapies. In the present study, the effects of pretreatments with the selective DAT inhibitor 3beta-(4-chlorophenyl)tropane-2beta-[3-(4'-methylphenyl)isoxazol-5-yl] hydrochloride (RTI-336) (0.3-1.7 mg/kg) were characterized in rhesus monkeys trained to self-administer cocaine (0.1 and 0.3 mg/kg/injection) under a multiple second-order schedule of i.v. drug or food delivery. In addition, RTI-336 (0.01-1.0 mg/kg/injection) was substituted for cocaine to characterize its reinforcing effects. Last, the dose of RTI-336 that reduced cocaine-maintained behavior by 50% (ED(50)) was coadministered with the selective serotonin transporter (SERT) inhibitors fluoxetine (3.0 mg/kg) and citalopram (3.0 mg/kg) to characterize their combined effects on cocaine self-administration. PET neuroimaging with the selective DAT ligand [(18)F]8-(2-[(18)F]fluoroethyl)-2beta-carbomethoxy-3beta-(4-chlorophenyl)nortropane quantified DAT occupancy at behaviorally relevant doses of RTI-336. Pretreatments of RTI-336 produced dose-related reductions in cocaine self-administration, and the ED(50) dose resulted in approximately 90% DAT occupancy. RTI-336 was reliably self-administered, but responding maintained by RTI-336 was lower than responding maintained by cocaine. Doses of RTI-336 that maintained peak rates of responding resulted in approximately 62% DAT occupancy. Co-administration of the ED(50) dose of RTI-336 in combination with either SERT inhibitor completely suppressed cocaine self-administration without affecting DAT occupancy. Hence, at comparable levels of DAT occupancy, coadministration of SERT inhibitors with RTI-336 produced more robust reductions in cocaine self-administration compared with RTI-336 alone. Collectively, the results indicate that combined inhibition of DAT and SERT warrants consideration as a viable approach in the development of cocaine medications

  2. Methylphenidate Treatment in Adolescent Rats with an Attention Deficit/Hyperactivity Disorder Phenotype: Cocaine Addiction Vulnerability and Dopamine Transporter Function

    PubMed Central

    Harvey, Roxann C; Sen, Sucharita; Deaciuc, Agripina; Dwoskin, Linda P; Kantak, Kathleen M

    2011-01-01

    Appropriate animal models of attention deficit/hyperactivity disorder (ADHD) and drug reinforcement allow investigation of possible underlying biological bases of ADHD and its comorbidity with cocaine addiction. Toward this end, spontaneously hypertensive rats (SHRs) exhibiting an ADHD phenotype were compared with Wistar-Kyoto (WKY) and Wistar (WIS) rats. Initially, 1.5 mg/kg oral methylphenidate or vehicle was administered between postnatal days 28 and 55, and acquisition of visual discrimination learning was examined. After discontinuing adolescent treatments, adult rats were evaluated for cocaine self-administration and dopamine transporter (DAT) function in the prefrontal cortex (PFC) and striatum. During adolescence, SHRs showed deficits in visual discrimination relative to WKY and WIS rats when non-medicated. Methylphenidate improved visual discrimination only in SHRs. Compared with WKY and WIS rats, SHRs with previous methylphenidate treatment acquired cocaine self-administration faster, identified cocaine as a highly efficacious reinforcer by displaying an upward shift in the cocaine dose–response function, and showed the greatest motivation to self-administer cocaine by exhibiting the highest progressive ratio breakpoints. In the PFC, the maximal dopamine uptake (Vmax) at DAT was decreased in SHRs and increased in WKY and WIS rats by previous methylphenidate treatment. The affinity (Km) for dopamine at DAT in the PFC was not different between strains, nor was Vmax or Km altered in the striatum by previous methylphenidate treatment in any strain. Methylphenidate-induced decreases in dopamine clearance by DAT in the PFC may underlie increased cocaine self-administration in SHRs. These preclinical findings suggest that caution should be exercised when methylphenidate is prescribed for first-time treatment of ADHD in adolescent patients, as cocaine addiction vulnerability may be augmented. PMID:21150910

  3. Charge Redistribution and Transport in Molecular Contacts.

    PubMed

    Corso, Martina; Ondráček, Martin; Lotze, Christian; Hapala, Prokop; Franke, Katharina J; Jelínek, Pavel; Pascual, J Ignacio

    2015-09-25

    The forces between two single molecules brought into contact, and their connection with charge transport through the molecular junction, are studied here using non contact AFM, STM, and density functional theory simulations. A carbon monoxide molecule approaching an acetylene molecule (C_{2}H_{2}) initially feels weak attractive electrostatic forces, partly arising from charge reorganization in the presence of molecular . We find that the molecular contact is chemically passive, and protects the electron tunneling barrier from collapsing, even in the limit of repulsive forces. However, we find subtle conductance and force variations at different contacting sites along the C_{2}H_{2} molecule attributed to a weak overlap of their respective frontier orbitals. PMID:26451568

  4. Quantitation of dopamine transporter mRNA in the rat brain: mapping, effects of "binge" cocaine administration and withdrawal.

    PubMed

    Maggos, C E; Spangler, R; Zhou, Y; Schlussman, S D; Ho, A; Kreek, M J

    1997-05-01

    Dopamine transporter (DAT) mRNA from selected brain regions of individual male Fischer rats was quantitated utilizing a sensitive solution hybridization assay in which the levels of RNase-protected 32P-labeled mRNA:cRNA hybrids were measured. DAT mRNA was detected in whole brain regions known to contain abundant DAT mRNA (mean picogram of DAT mRNA/microgram of total RNA +/- SEM): substantia nigra, 7.17 +/- 0.47; ventral tegmentum, 4.71 +/- 0.38. In regions known to contain low levels of DAT mRNA, these levels were detected: central grey, 0.39 +/- 0.06; hypothalamus, 0.14 +/- 0.03. In addition, DAT mRNA was detected in areas where it had not previously been identified: amygdala, 0.19 +/- 0.03; caudate-putamen, 0.15 +/- 0.03; nucleus accumbens, 0.13 +/- 0.01; pons/medulla, 0.12 +/- 0.02; globus pallidus, 0.09 +/- 0.04; pituitary 0.07 +/- 0.01; frontal cortex, 0.05 +/- 0.01. No DAT mRNA was detected in 150 micrograms of rat liver RNA. As cocaine binds to and inhibits the activity of the dopamine transporter, we sought to determine if there were differences in dopamine transporter mRNA levels between saline- and cocaine-injected rats or rats withdrawn from a chronic "binge" pattern (15 mg/kg per dose i.p.; three doses at 1 h intervals each day) cocaine injection. Using trichloroacetic acid precipitation of mRNA:cRNA hybrids from RNA extracted from whole brain regions, we found no significant differences in the substantia nigra or the ventral tegmentum following subacute (3 days) binge, chronic (14 days) binge or 10 days withdrawal from a chronic binge pattern cocaine or saline administration. PMID:9097405

  5. Relationship between dopamine transporter occupancy and methylphenidate induced high in humans

    SciTech Connect

    Volkow, N.D.; Wang, G.J.; Fowler, J.S. |

    1996-05-01

    The inhibition of the dopamine transporter (DAT) by cocaine has been shown to be indispensable for its reinforcing properties. The development of drugs that inibit the DAT has become a major target to prevent cocaine`s effects. However prevention of the {open_quotes}high{close_quotes} by DAT inhibitors has never been demonstrated. This study evaluates the ability to block methylphenidate (MP), a DAT inhibitor drug with similar reinforcing properties to cocaine, induced {open_quotes}high{close_quotes} by prior DAT inhibition. It uses PET and [{sup 11}C]d-threo-methylphenidate to measure the relationship between DAT occupancy prior to administration of MP and the intensity of the subjective perception of the {open_quotes}high{close_quotes} in 8 controls. MP (0.375 mg/kg iv) which was administered as a single injection and also as two sequential doses given 60 minutes apart significantly reduced the ratio of the distribution volume for [{sup 11}C]d-threo-methylphenidate in striatum to that in cerebellum from a baseline of 2.83 {plus_minus} 0.2 to 1.29 {plus_minus} 0.1 at 7 minutes and to 1.37 {plus_minus} 0.2 at 60 minutes after a single injection of MP and to 1.14 {plus_minus} 0.1 at 7 minutes after the second of two sequential MP doses. This corresponds to a DAT occupancy by MP of 84% {plus_minus} 7 at 7 minutes and of 77% {plus_minus} 6 at 60 minutes after a single injection of MP and of 93% {plus_minus} 7 at 7 after the second of two sequential MP doses. The subjective perception of {open_quotes}high{close_quotes} experienced after the second injection of MP was of a similar magnitude to that experienced after the first injection of MP was of a similar magnitude to that experienced after the first injection, in spite of the very different starting DAT occupancies (0 and 77%, respectively). DAT occupancy was not correlated with the {open_quotes}high{close_quotes}; and one subject with 100% DAT occupancy did not perceive the {open_quotes}high{close_quotes}.

  6. Quantification of dopamine transporter density with [18F]FECNT PET in healthy humans

    PubMed Central

    Nye, Jonathon A.; Votaw, John R.; Bremner, J. Douglas; Davis, Margaret R.; Voll, Ronald J.; Camp, Vernon M.; Goodman, Mark M.

    2015-01-01

    Introduction Fluorine-18 labeled 2β-carbomethoxy-3β-(4-chlorophenyl)-8-(2-fluoroethyl)nortropane ([18 F]FECNT) binds reversibly to the dopamine transporter (DAT) with high selectivity. [18 F]FECNT has been used extensively in the quantification of DAT occupancy in non-human primate brain and can distinguish between Parkinson's and healthy controls in humans. The purpose of this work was to develop a compartment model to characterize the kinetics of [18 F]FECNT for quantification of DAT density in healthy human brain. Methods Twelve healthy volunteers underwent 180 min dynamic [18 F]FECNT PET imaging including sampling of arterial blood. Regional time-activity curves were extracted from the caudate, putamen and midbrain including a reference region placed in the cerebellum. Binding potential, BPND, was calculated for all regions using kinetic parameters estimated from compartmental and Logan graphical model fits to the time-activity data. Simulations were performed to determine whether the compartment model could reliably fit time-activity data over a range of BPND values. Results The kinetics of [18 F]FECNT were well-described by the reversible 2-tissue arterial input and full reference tissue compartment models. Calculated binding potentials in the caudate, putamen and midbrain were in good agreement between the arterial input model, reference tissue model and the Logan graphical model. The distribution volume in the cerebellum did not reach a plateau over the duration of the study, which may be a result of non-specific binding in the cerebellum. Simulations that included non-specific binding show that the reference and arterial input models are able to estimate BPND for DAT densities well below that observed in normal volunteers. Conclusion The kinetics of [18 F]FECNT in human brain are well-described by arterial input and reference tissue compartment models. Measured and simulated data show that BPND calculated with reference tissue model is proportional to

  7. Differential action of methamphetamine on tyrosine hydroxylase and dopamine transport in the nigrostriatal pathway of μ-opioid receptor knockout mice.

    PubMed

    Park, Sang Won; He, Zhi; Shen, Xine; Roman, Richard J; Ma, Tangeng

    2012-06-01

    Extensive anatomical and functional interactions exist between central dopaminergic and opioidergic systems and both systems are proposed to be targets for amphetamine-like drugs. We have previously reported that μ-opioid receptor (μ-OR) knockout mice are resistant to the loss of dopamine in the striatum and the development of behavioral sensitization induced by repeated methamphetamine (METH) treatment. The present study assessed whether METH-treated μ-OR knockout mice exhibit a differential response of the expression of dopamine transporter and tyrosine hydroxylase (TH), the rate-limiting enzyme for dopamine synthesis and maintaining dopamine levels. Mice daily received intraperitoneal injection of METH (0, 0.6, 2.5, or 10 mg/kg) for 7 days and sacrificed on day 11 (4 days after the last injection). The expression of TH protein in the striatum and the levels of TH mRNA and number of TH positive neurons in the substantia nigra were reduced in wild-type mice treated with METH (2.5 and 10 mg/kg), but not in the μ-OR knockout mice. In contrast, METH exposure at the highest dose (10 mg/kg) reduced dopamine transporter levels in both strains of mice. These results suggest that the μ-OR contributes to METH-induced loss of dopamine and behavioral sensitization by decreasing the expression of TH. PMID:22329540

  8. Ascorbic acid and striatal transport of (/sup 3/H)1-methyl-4-phenylpyridine (MPP/sup +/) and (/sup 3/H)dopamine

    SciTech Connect

    Debler, E.A.; Hashim, A.; Lajtha, A.; Sershen, H.

    1988-01-01

    The inhibition of uptake of (/sup 3/H)dopamine and (/sup 3/H)1-methyl-4-phenylpyridine (MPP/sup +/) was examined in mouse striatal synaptosomal preparations. Kinetic analysis indicated that ascorbic acid is a noncompetitive inhibitor of (/sup 3/H)MPP/sup +/ uptake. No inhibition of (/sup 3/H)dopamine uptake is observed. The dopamine uptake blockers, GBR-12909, cocaine, and mazindol strongly inhibit (IC/sub 50/ < 1 ..mu..M) both (/sup 3/H)dopamine and (/sup 3/H)MPP/sup +/ transport. Nicotine, its metabolites, and other tobacco alkaloids are weak inhibitors except 4-phenylpyridine and lobeline, which are moderate inhibitors of both (/sup 3/H)dopamine and (/sup 3/H)MPP/sup +/ uptake. These similarities in potencies are in agreement with the suggestion that (/sup 3/H)MPP/sup +/ and (/sup 3/H) are transported by the same carrier. The differences observed in the alteration of dopaminergic transport and mazindol binding by ascorbic acid suggest that ascorbic acid's effects on (/sup 3/H)MPP/sup +/ transport are related to translocation and/or dissociation processes occurring subsequent to the initial binding event.

  9. Single Cell Measurement of Dopamine Release with Simultaneous Voltage-clamp and Amperometry

    PubMed Central

    Saha, Kaustuv; Swant, Jarod; Khoshbouei, Habibeh

    2012-01-01

    After its release into the synaptic cleft, dopamine exerts its biological properties via its pre- and post-synaptic targets1. The dopamine signal is terminated by diffusion2-3, extracellular enzymes4, and membrane transporters5. The dopamine transporter, located in the peri-synaptic cleft of dopamine neurons clears the released amines through an inward dopamine flux (uptake). The dopamine transporter can also work in reverse direction to release amines from inside to outside in a process called outward transport or efflux of dopamine5. More than 20 years ago Sulzer et al. reported the dopamine transporter can operate in two modes of activity: forward (uptake) and reverse (efflux)5. The neurotransmitter released via efflux through the transporter can move a large amount of dopamine to the extracellular space, and has been shown to play a major regulatory role in extracellular dopamine homeostasis6. Here we describe how simultaneous patch clamp and amperometry recording can be used to measure released dopamine via the efflux mechanism with millisecond time resolution when the membrane potential is controlled. For this, whole-cell current and oxidative (amperometric) signals are measured simultaneously using an Axopatch 200B amplifier (Molecular Devices, with a low-pass Bessel filter set at 1,000 Hz for whole-cell current recording). For amperometry recording a carbon fiber electrode is connected to a second amplifier (Axopatch 200B) and is placed adjacent to the plasma membrane and held at +700 mV. The whole-cell and oxidative (amperometric) currents can be recorded and the current-voltage relationship can be generated using a voltage step protocol. Unlike the usual amperometric calibration, which requires conversion to concentration, the current is reported directly without considering the effective volume7. Thus, the resulting data represent a lower limit to dopamine efflux because some transmitter is lost to the bulk solution. PMID:23207721

  10. Molecularly imprinted nanohybrids based on dopamine-modified poly(γ-glutamic acid) for electrochemical sensing of melamine.

    PubMed

    Zhang, Rongli; Xu, Sheng; Zhu, Ye; Zhao, Wei; Luo, Jing; Liu, Xiaoya; Tang, Dingxing

    2016-11-15

    A voltammetric sensor for melamine (MEL) was prepared from molecularly imprinted nanohybrids (MINBs). A dopamine modified poly-γ-glutamic acid copolymer (γ-PGA-DA) and MEL were self-assembled into MEL/γ-PGA-DA nanoparticles (NPs) in aqueous solution via weak interactions, followed by adding an aqueous AgNO3 solution into the mixture. The Ag(+) was adsorbed in the MEL/γ-PGA-DA NPs and spontaneously reduced to Ag NPs by the dopamine moieties of γ-PGA-DA, forming Ag/MEL/γ-PGA-DA MINBs, which were then cast on a gold electrode to form a MINBs film. The MEL was removed by electrolysis via catalysis of Ag NPs at a constant potential of 1.4V in phosphate buffer saline solution, to obtain a voltammetric sensor for MEL. The sensor responded linearly to MEL in the concentration range of 5×10(-18) to 5×10(-7)molL(-1). Compared to other published molecularly imprinted polymer sensors for sensing MEL, the prepared MINBs sensor had much wider detection range with lower detection limit. PMID:27196255

  11. Molecular modeling of auxin transport inhibitors

    SciTech Connect

    Gardner, G.; Black-Schaefer, C.; Bures, M.G. )

    1990-05-01

    Molecular modeling techniques have been used to study the chemical and steric properties of auxin transport inhibitors. These bind to a specific site on the plant plasma membrane characterized by its affinity for N-1-naphthylphthalamic acid (NPA). A three-dimensional model was derived from critical features of ligands for the NPA receptor, and a suggested binding conformation is proposed. This model, along with three-dimensional structural searching techniques, was then used to search the Abbott corporate database of chemical structures. Of the 467 compounds that satisfied the search criteria, 77 representative molecules were evaluated for their ability to compete for ({sup 3}H)NPA binding to corn microsomal membranes. Nineteen showed activity that ranged from 16 to 85% of the maximum NPA binding. Four of the most active of these, from chemical classes not included in the original compound set, also inhibited polar auxin transport through corn coleoptile sections.

  12. Vibrationally mediated transport in molecular transistors

    NASA Astrophysics Data System (ADS)

    Santamore, D. H.; Lambert, Neill; Nori, Franco

    2013-02-01

    We investigate the steady-state electronic transport through a suspended dimer molecule coupled to leads. When strongly coupled to a vibrational mode, the electron transport is enhanced at the phonon resonant frequency and higher order resonances. The temperature and bias determine the nature of the phonon-assisted resonances, with clear absorption and emission peaks. The strong coupling also induces a Frank-Condon-like blockade, suppressing the current between the resonances. We compare an analytical polaron transformation method to two exact numerical methods: the Hierarchy equations of motion and an exact diagonalization in the Fock basis. In the steady state, our two numerical results are an exact match and qualitatively reflect the main features of the polaron treatment. Our results indicate the possibility of a new type of molecular transistor or sensor where the current can be extremely sensitive to small changes in the energies of the electronic states in the dimer.

  13. Molecular motor driven transportation on microtubule loops

    NASA Astrophysics Data System (ADS)

    Sikora, Aurelien; Federici, Filippo; Kim, Kyongwan; Nakazawa, Hikaru; Umetsu, Mitsuo; Hwang, Wonmuk; Teizer, Winfried

    2015-03-01

    Molecular motors such as kinesin are naturally fitted for the transport of cargo. By offering an unlimited path, microtubule loops allow the study of kinesin motility on distances exceeding that offered by a single microtubule. Moreover, the periodicity of the path allows the comparisons of trajectories between laps. Here we study the motility of quantum dot labeled kinesin on microtubule loops. Motility of kinesins over multiple laps is observed and their trajectories are extracted from kymograph using a custom algorithm. Distribution of velocities at given locations do not vary randomly but show a correlation with the presence of obstacles. Possible mechanisms responsible for the long range transport are discussed in the context of available theories.

  14. Synthesis and evaluation of novel N-fluoropyridyl derivatives of tropane as potential PET imaging agents for the dopamine transporter

    PubMed Central

    Liu, Jingying; Zhu, Lin; Plössl, Karl; Lieberman, Brian P.; Kung, Hank F.

    2011-01-01

    A series of novel N-fluoropyridyl-containing tropane derivatives were synthesized and their binding affinities for the dopamine transporter (DAT), serotonin transporter (SERT) and norepinephrine (NET) were determined via competitive radioligand binding assays. Among these derivatives, compound 6d showed the highest binding affinity to DAT (Ki = 4.1 nM), and selectivity for DAT over SERT (5 fold) and NET (16 fold). Compound 6d was radiolabeled with Fluorine-18 in two steps. Regional brain distribution and ex vivo autoradiography studies of [18F]6d demonstrated that the ligand was selectively localized in the striatum region, where DAT binding sites are highly expressed. [18F]6d may be useful as a potential radioligand for imaging DATs with PET. PMID:21458259

  15. 3,4-Methylenedioxypyrovalerone prevents while methylone enhances methamphetamine-induced damage to dopamine nerve endings: β-ketoamphetamine modulation of neurotoxicity by the dopamine transporter.

    PubMed

    Anneken, John H; Angoa-Pérez, Mariana; Kuhn, Donald M

    2015-04-01

    Methylone, 3,4-methylenedioxypyrovalerone (MDPV), and mephedrone are psychoactive ingredients of 'bath salts' and their abuse represents a growing public health care concern. These drugs are cathinone derivatives and are classified chemically as β-ketoamphetamines. Because of their close structural similarity to the amphetamines, methylone, MDPV, and mephedrone share most of their pharmacological, neurochemical, and behavioral properties. One point of divergence in their actions is the ability to cause damage to the CNS. Unlike methamphetamine, the β-ketoamphetamines do not damage dopamine (DA) nerve endings. However, mephedrone has been shown to significantly accentuate methamphetamine neurotoxicity. Bath salt formulations contain numerous different psychoactive ingredients, and individuals who abuse bath salts also coabuse other illicit drugs. Therefore, we have evaluated the effects of methylone, MDPV, mephedrone, and methamphetamine on DA nerve endings. The β-ketoamphetamines alone or in all possible two-drug combinations do not result in damage to DA nerve endings but do cause hyperthermia. MDPV completely protects against the neurotoxic effects of methamphetamine while methylone accentuates it. Neither MDPV nor methylone attenuates the hyperthermic effects of methamphetamine. The potent neuroprotective effects of MDPV extend to amphetamine-, 3,4-methylenedioxymethamphetamine-, and MPTP-induced neurotoxicity. These results indicate that β-ketoamphetamine drugs that are non-substrate blockers of the DA transporter (i.e., MDPV) protect against methamphetamine neurotoxicity, whereas those that are substrates for uptake by the DA transporter and which cause DA release (i.e., methylone, mephedrone) accentuate neurotoxicity. METH (a) enters DA nerve endings via the DAT, causes leakage of DA into the cytoplasm and then into the synapse via DAT-mediated reverse transport. Methylone (METHY) and mephedrone (MEPH; b), like METH, are substrates for the DAT but release

  16. Changes in dopamine transporter binding in nucleus accumbens following chronic self-administration cocaine: heroin combinations.

    PubMed

    Pattison, Lindsey P; McIntosh, Scot; Sexton, Tammy; Childers, Steven R; Hemby, Scott E

    2014-10-01

    Concurrent use of cocaine and heroin (speedball) has been shown to exert synergistic effects on dopamine neurotransmission in the nucleus accumbens (NAc), as observed by significant increases in extracellular dopamine levels and compensatory elevations in the maximal reuptake rate of dopamine. The present studies were undertaken to determine whether chronic self-administration of cocaine, heroin or a combination of cocaine:heroin led to compensatory changes in the abundance and/or affinity of high- and low-affinity DAT binding sites. Saturation binding of the cocaine analog [(125) I] 3β-(4-iodophenyl)tropan-2β-carboxylic acid methyl ester ([(125) I]RTI-55) in rat NAc membranes resulted in binding curves that were best fit to two-site binding models, allowing calculation of dissociation constant (Kd ) and binding density (Bmax ) values corresponding to high- and low-affinity DAT binding sites. Scatchard analysis of the saturation binding curves clearly demonstrate the presence of high- and low- affinity binding sites in the NAc, with low-affinity sites comprising 85 to 94% of the binding sites. DAT binding analyses revealed that self-administration of cocaine and a cocaine:heroin combination increased the affinity of the low-affinity site for the cocaine congener RTI-55 compared to saline. These results indicate that the alterations observed following chronic speedball self-administration are likely due to the cocaine component alone; thus further studies are necessary to elaborate upon the synergistic effect of cocaine:heroin combinations on the dopamine system in the NAc. PMID:24916769

  17. Molecular cloning and characterization of a novel dopamine receptor (D3) as a target for neuroleptics

    NASA Astrophysics Data System (ADS)

    Sokoloff, Pierre; Giros, Bruno; Martres, Marie-Pascale; Bouthenet, Marie-Louise; Schwartz, Jean-Charles

    1990-09-01

    A dopamine receptor has been characterized which differs in its pharmacology and signalling system from the D1 or D2 receptor and represents both an autoreceptor and a postsynaptic receptor. The D3 receptor is localized to limbic areas of the brain, which are associated with cognitive, emotional and endocrine functions. It seems to mediate some of the effects of antipsychotic drugs and drugs used against Parkinson's disease, that were previously thought to interact only with D2 receptors.

  18. Serotonin2C receptors modulate dopamine transmission in the nucleus accumbens independently of dopamine release: behavioral, neurochemical and molecular studies with cocaine.

    PubMed

    Cathala, Adeline; Devroye, Céline; Maitre, Marlène; Piazza, Pier Vincenzo; Abrous, Djoher Nora; Revest, Jean-Michel; Spampinato, Umberto

    2015-05-01

    In keeping with its ability to control the mesoaccumbens dopamine (DA) pathway, the serotonin2C receptor (5-HT2C R) plays a key role in mediating the behavioral and neurochemical effects of drugs of abuse. Studies assessing the influence of 5-HT2C R agonists on cocaine-induced responses have suggested that 5-HT2C Rs can modulate mesoaccumbens DA pathway activity independently of accumbal DA release, thereby controlling DA transmission in the nucleus accumbens (NAc). In the present study, we assessed this hypothesis by studying the influence of the 5-HT2C R agonist Ro 60-0175 on cocaine-induced behavioral, neurochemical and molecular responses. The i.p. administration of 1 mg/kg Ro 60-0175 inhibited hyperlocomotion induced by cocaine (15 mg/kg, i.p.), had no effect on cocaine-induced DA outflow in the shell, and increased it in the core subregion of the NAc. Furthermore, Ro 60-0175 inhibited the late-onset locomotion induced by the subcutaneous administration of the DA-D2 R agonist quinpirole (0.5 mg/kg), as well as cocaine-induced increase in c-Fos immunoreactivity in NAc subregions. Finally, Ro 60-0175 inhibited cocaine-induced phosphorylation of the DA and c-AMP regulated phosphoprotein of Mr 32 kDa (DARPP-32) at threonine residues in the NAc core, this effect being reversed by the selective 5-HT2C R antagonist SB 242084 (0.5 mg/kg, i.p.). Altogether, these findings demonstrate that 5-HT2C Rs are capable of modulating mesoaccumbens DA pathway activity at post-synaptic level by specifically controlling DA signaling in the NAc core subregion. In keeping with the tight relationship between locomotor activity and NAc DA function, this interaction could participate in the inhibitory control of cocaine-induced locomotor activity. PMID:24661380

  19. No association between schizophrenia and polymorphisms within the genes for debrisoquine 4-hydroxylase (CYP2D6) and the dopamine transporter (DAT)

    SciTech Connect

    Daniels, J.; Williams, J.; Asherson, P.; McGuffin, P.; Owen, M.

    1995-02-27

    It has been suggested that the cytochrome P450 mono-oxygenase, debrisoquine 4-hydroxylase, is involved in the catabolism and processing of neurotransmitters subsequent to their reuptake into target cells. It is also thought to be related to the dopamine transporter that acts to take released dopamine back up into presynaptic terminals. The present study used the association approach to test the hypothesis that mutations in the genes for debrisoquine 4-hydroxylase (CYP2D6) and the dopamine transporter (DAT) confer susceptibility to schizophrenia. There were no differences in allele or genotype frequencies between patients and controls in the mutations causing the poor metaboliser phenotype in CYP2D6. In addition there was no association found between schizophrenia and a 48 bp repeat within the 3{prime} untranslated region of DAT. 18 refs., 2 tabs.

  20. Molecular approach to intracellular cargo transport

    NASA Astrophysics Data System (ADS)

    Yildiz, Ahmet

    2010-03-01

    Landmark discoveries in the study of cytoplasmic motors have been made through advances in single molecule biophysics and detailed mechanistic models exist for kinesin and dynein. However, the function of motors in physiological conditions has not been carefully tested. In cells, more than few dyneins can attach to the same cargo and interact with the opposite polarity motors of kinesin. To study the molecular crosstalk between the motors, we have used intraflagellar transport (IFT) in Chlamydomonas reinhardtii as a model system. Ultrahigh spatio-temporal tracking of single cargo movement showed that IFT particles move for long distances unidirectionally with 8 nm increments, agreeing with measured step sizes of kinesin and dynein. To measure how many motors transport each cargo, we have linked large polystyrene beads to internal IFT particles through a transmembrane protein. Force measurements indicated that, on average, 3-4 motors transport cargoes in each direction. The results showed that IFT motors are tightly coordinated and might be involved in recycling each other to the appropriate end of the flagellum.

  1. Influence of chronic dopamine transporter inhibition by RTI-336 on motor behavior, sleep, and hormone levels in rhesus monkeys.

    PubMed

    Andersen, Monica L; Sawyer, Eileen K; Carroll, F Ivy; Howell, Leonard L

    2012-04-01

    Dopamine transporter (DAT) inhibitors have been developed as a promising treatment approach for cocaine dependence. However, the stimulant effects of DAT inhibitors have the potential to disrupt sleep patterns, and the influence of long-term treatment on dopamine neurochemistry is still unknown. The objectives of this study were to (1) explore the stimulant-related effects of chronic DAT inhibitor (RTI-336) treatment on motor activity and sleep-like measures in male rhesus monkeys (Macaca mulatta; n = 4) and (2) to determine the effect of drug treatment on prolactin and cortisol levels. Subjects were fitted with a collar-mounted activity monitor to evaluate their motor activity, with 4 days of baseline recording preceding 21 days of daily saline or RTI-336 (1 mg/kg/day; intramuscular) injections. Blood samples were collected immediately prior to and following chronic treatment to assess hormone levels. RTI-336 produced a significant increase in locomotor activity at the end of the daytime period compared to saline administration. During the 3-week treatment period, sleep efficiency was decreased and the fragmentation index and latency to sleep onset were significantly increased. Hormone levels were not changed throughout the study. Chronic treatment with RTI-336 has a mild but significant stimulant effect, as evidenced by the significant increase in activity during the evening period which may cause minor disruptions in sleep measures. PMID:22023668

  2. Impaired spatial working memory and decreased frontal cortex BDNF protein level in dopamine transporter knock out mice

    PubMed Central

    Li, BingJin; Arime, Yosefu; Hall, F. Scott; Uhl, George R.; Sora, Ichiro

    2010-01-01

    Brain-derived neurotrophic factor (BDNF), one of the key brain neurotrophins, has been implicated in neuronal plasticity and memory. Recent studies document the importance of BDNF for normal long-term memory functions. However, there are few studies of the roles of BDNF in short term memory. Dopamine is likely to play important roles in BDNF gene expression in specific brain regions, including frontal cortical regions that are implicated in short term working memory processes that include spontaneous alternation. We have thus tested spatial working memory in dopamine transporter knockout (DAT KO) and wild-type mice. Spontaneous alternation in the Y-maze, an index of short-term spatial working memory in mice, was significantly decreased in DAT KO mice compared to wildtype mice. BDNF protein was significantly decreased in frontal cortex, though not in striatum or hippocampus, of the DAT KO mice. The data support the hypothesis that impaired spatial working memory in DAT KO mice may be related to decreased frontal cortical BDNF in these animals, and document apparent roles for BDNF in a short term memory process. PMID:19932884

  3. Predicting childhood effortful control from interactions between early parenting quality and children’s dopamine transporter gene haplotypes

    PubMed Central

    LI, YI; SULIK, MICHAEL J.; EISENBERG, NANCY; SPINRAD, TRACY L.; LEMERY-CHALFANT, KATHRYN; STOVER, DARYN A.; VERRELLI, BRIAN C.

    2015-01-01

    Children’s observed effortful control (EC) at 30, 42, and 54 months (n = 145) was predicted from the interaction between mothers’ observed parenting with their 30-month-olds and three variants of the solute carrier family C6, member 3 (SLC6A3) dopamine transporter gene (single nucleotide polymorphisms in intron8 and intron13, and a 40 base pair variable number tandem repeat [VNTR] in the 3′-untranslated region [UTR]), as well as haplotypes of these variants. Significant moderating effects were found. Children without the intron8-A/intron13-G, intron8-A/3′-UTR VNTR-10, or intron13-G/3′-UTR VNTR-10 haplotypes (i.e., haplotypes associated with the reduced SLC6A3 gene expression and thus lower dopamine functioning) appeared to demonstrate altered levels of EC as a function of maternal parenting quality, whereas children with these haplotypes demonstrated a similar EC level regardless of the parenting quality. Children with these haplotypes demonstrated a trade-off, such that they showed higher EC, relative to their counterparts without these haplotypes, when exposed to less supportive maternal parenting. The findings revealed a diathesis–stress pattern and suggested that different SLC6A3 haplotypes, but not single variants, might represent different levels of young children’s sensitivity/responsivity to early parenting. PMID:25924976

  4. Progression of changes in dopamine transporter binding site density as a result of cocaine self-administration in rhesus monkeys.

    PubMed

    Letchworth, S R; Nader, M A; Smith, H R; Friedman, D P; Porrino, L J

    2001-04-15

    The present study examined the time course of alterations in levels of dopamine transporter (DAT) binding sites that accompany cocaine self-administration using quantitative in vitro receptor autoradiography with [(3)H]WIN 35,428. The density of dopamine transporter binding sites in the striatum of rhesus monkeys with 5 d, 3.3 months, or 1.5 years of cocaine self-administration experience was compared with DAT levels in cocaine-naive control monkeys. Animals in the long-term (1.5 years) exposure group self-administered cocaine at 0.03 mg/kg per injection, whereas the initial (5 d) and chronic (3.3 months) treatment groups were each divided into lower dose (0.03 mg/kg per injection) and higher dose (0.3 mg/kg per injection) groups. Initial cocaine exposure led to moderate decreases in [(3)H]WIN 35,428 binding sites, with significant changes in the dorsolateral caudate (-25%) and central putamen (-19%) at the lower dose. Longer exposure, in contrast, resulted in elevated levels of striatal binding sites. The increases were most pronounced in the ventral striatum at the level of the nucleus accumbens shell. At the lower dose of the chronic phase, for example, significant increases of 21-42% were measured at the caudal level of the ventral caudate, ventral putamen, olfactory tubercle, and accumbens core and shell. Systematic variation of cocaine dose and drug exposure time demonstrated the importance of these factors in determining the intensity of increased DAT levels. With self-administration of higher doses especially, increases were more intense and included dorsal portions of the striatum so that every region at the caudal level exhibited a significant increase in DAT binding sites (20-54%). The similarity of these findings to previous studies in human cocaine addicts strongly suggest that the increased density of dopamine transporters observed in studies of human drug abusers are the result of the neurobiological effects of cocaine, ruling out confounds such as

  5. Transport of toxic metals by molecular mimicry.

    PubMed Central

    Ballatori, Nazzareno

    2002-01-01

    Intracellular concentrations of essential metals are normally maintained within a narrow range, whereas the nonessential metals generally lack homeostatic controls. Some of the factors that contribute to metal homeostasis have recently been identified at the molecular level and include proteins that mediate import of essential metals from the extracellular environment, those that regulate delivery to specific intracellular proteins or compartments, and those that mediate metal export from the cell. Some of these proteins appear highly selective for a given essential metal; however, others are less specific and interact with multiple metals, including toxic metals. For example, DCT1 (divalent cation transporter-1; also known as NRAMP2 or DMT1) is considered to be a major cellular uptake mechanism for Fe(2+) and other essential divalent metals, but this protein also mediates uptake of Cd(2+), Pb(2+), and possibly of other toxic divalent metals. The ability of nonessential metals to interact with binding sites for essential metals is critical for their ability to gain access to specific cellular compartments and for their ability to disrupt normal biochemical or physiological functions. Another major mechanism by which metals traverse cell membranes and produce cell injury is by forming complexes whose overall structures mimic those of endogenous molecules. For example, it has long been known that arsenate and vanadate can compete with phosphate for transport and metabolism, thereby disrupting normal cellular functions. Similarly, cromate and molybdate can mimic sulfate in biological systems. Studies in our laboratory have focused on the transport and toxicity of methylmercury (MeHg) and inorganic mercury. Mercury has a high affinity for reduced sulfhydryl groups, including those of cysteine and glutathione (GSH). MeHg-l-cysteine is structurally similar to the amino acid methionine, and this complex is a substrate for transport systems that carry methionine across

  6. Dopamine transporter-dependent and -independent striatal binding of the benztropine analog JHW 007, a cocaine antagonist with low abuse liability

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The benztropine analog JHW 007 displays high affinity for the dopamine transporter (DAT), but unlike typical DAT ligands, has relatively low abuse liability and blocks effects of cocaine,including its self-administration. To determine sites responsible for the cocaine-antagonist effects of JHW 007, ...

  7. Lack of Association between a 3'UTR VNTR Polymorphism of Dopamine Transporter Gene (SLC6A3) and ADHD in a Brazilian Sample of Adult Patients

    ERIC Educational Resources Information Center

    Aperecida da Silva, Maria; Cordeiro, Quirino; Louza, Mario; Vallada, Homero

    2011-01-01

    Objective: To investigate a possible association between a 3'UTR VNTR polymorphism of the dopamine transporter gene (SLC6A3) and ADHD in a Brazilian sample of adult patients. Method: Study Case-control with 102 ADHD adult outpatients ("DSM-IV" criteria) and 479 healthy controls. The primers' sequence used were: 3'UTR-Forward: 5' TGT GGT GAT GGG…

  8. How Addictive Drugs Disrupt Presynaptic Dopamine Neurotransmission

    PubMed Central

    Sulzer, David

    2011-01-01

    The fundamental principle that unites addictive drugs appears to be that each enhances synaptic dopamine by means that dissociate it from normal behavioral control, so that they act to reinforce their own acquisition. This occurs via the modulation of synaptic mechanisms involved in learning, including enhanced excitation or disinhibition of dopamine neuron activity, blockade of dopamine reuptake, and altering the state of the presynaptic terminal to enhance evoked over basal transmission. Amphetamines offer an exception to such modulation in that they combine multiple effects to produce non-exocytic stimulation-independent release of neurotransmitter via reverse transport independent from normal presynaptic function. Questions on the molecular actions of addictive drugs, prominently including the actions of alcohol and solvents, remain unresolved, but their ability to co-opt normal presynaptic functions helps to explain why treatment for addiction has been challenging. PMID:21338876

  9. Molecular dynamics simulations of microscale fluid transport

    SciTech Connect

    Wong, C.C.; Lopez, A.R.; Stevens, M.J.; Plimpton, S.J.

    1998-02-01

    Recent advances in micro-science and technology, like Micro-Electro-Mechanical Systems (MEMS), have generated a group of unique liquid flow problems that involve characteristic length scales of a Micron. Also, in manufacturing processes such as coatings, current continuum models are unable to predict microscale physical phenomena that appear in these non-equilibrium systems. It is suspected that in these systems, molecular-level processes can control the interfacial energy and viscoelastic properties at the liquid/solid boundary. A massively parallel molecular dynamics (MD) code has been developed to better understand microscale transport mechanisms, fluid-structure interactions, and scale effects in micro-domains. Specifically, this MD code has been used to analyze liquid channel flow problems for a variety of channel widths, e.g. 0.005-0.05 microns. This report presents results from MD simulations of Poiseuille flow and Couette flow problems and addresses both scaling and modeling issues. For Poiseuille flow, the numerical predictions are compared with existing data to investigate the variation of the friction factor with channel width. For Couette flow, the numerical predictions are used to determine the degree of slip at the liquid/solid boundary. Finally, the results also indicate that shear direction with respect to the wall lattice orientation can be very important. Simulation results of microscale Couette flow and microscale Poiseuille flow for two different surface structures and two different shear directions will be presented.

  10. Cooperative cargo transport by several molecular motors

    PubMed Central

    Klumpp, Stefan; Lipowsky, Reinhard

    2005-01-01

    The transport of cargo particles that are pulled by several molecular motors in a cooperative manner is studied theoretically in this article. The transport properties depend primarily on the maximal number N of motor molecules that may pull simultaneously on the cargo particle. Because each motor must unbind from the filament after a finite number of steps but can also rebind to it again, the actual number of pulling motors is not constant but varies with time between zero and N. An increase in the maximal number N leads to a strong increase of the average walking distance (or run length) of the cargo particle. If the cargo is pulled by up to N kinesin motors, for example, the walking distance is estimated to be 5N–1/N micrometers, which implies that seven or eight kinesin molecules are sufficient to attain an average walking distance in the centimeter range. If the cargo particle is pulled against an external load force, this force is shared between the motors, which provides a nontrivial motor–motor coupling and a generic mechanism for nonlinear force–velocity relationships. With increasing load force, the probability distribution of the instantaneous velocity is shifted toward smaller values, becomes broader, and develops several peaks. Our theory is consistent with available experimental data and makes quantitative predictions that are accessible to systematic in vitro experiments. PMID:16287974