title: Regulation of dopamine transporter function and cell surface expression by D3 dopamine receptors.
abstract: D(3) dopamine receptors are expressed by dopamine neurons and are implicated in the modulation of presynaptic dopamine neurotransmission. The mechanisms underlying this modulation remain ill defined. The dopamine transporter, which terminates dopamine transmission via reuptake of released neurotransmitter, is regulated by receptor- and second messenger-linked signaling pathways. Whether D3 receptors regulate dopamine transporter function is unknown. We addressed this issue using a fluorescent imaging technique that permits real time quantification of dopamine transporter function in living single cells. Accumulation of the fluorescent dopamine transporter substrate trans-4-[4-(dimethylamino)styryl]-1-methylpyridinium (ASP(+)) in human embryonic kidney cells expressing human dopamine transporter was saturable and temperature-dependent. In cells co-expressing dopamine transporter and D3 receptors, the D2/D3 agonist quinpirole produced a rapid, concentration-dependent, and pertussis toxin-sensitive increase of ASP(+) uptake. Similar agonist effects were observed in Neuro2A cells and replicated in human embryonic kidney cells using a radioligand uptake assay in which binding to and activation of D3 receptors by [(3)H]dopamine was prevented. D3 receptor stimulation activated phosphoinositide 3-kinase and MAPK. Inhibition of either kinase prevented the quinpirole-induced increase in uptake. D3 receptor activation differentially affected dopamine transporter function and subcellular distribution depending on the duration of agonist exposure. Biotinylation experiments revealed that the rapid increase of uptake was associated with increased cell surface and decreased intracellular expression and increased dopamine transporter exocytosis. In contrast, prolonged agonist exposure reduced uptake and transporter cell surface expression. These results demonstrate that D3 receptors regulate dopamine transporter function and identify a novel mechanism by which D3 receptors regulate extracellular dopamine concentrations.
authors.affiliation: Integrative Neuroscience Section, National Institutes of Health/National Institute on Drug Abuse Intramural Research Program/Department of Health and Human Services, Baltimore, Maryland 21224, USA.
authors.forename: Agustin
authors.initials: A
authors.lastname: Zapata
authors.forename: Bronwyn
authors.initials: B
authors.lastname: Kivell
authors.forename: Yang
authors.initials: Y
authors.lastname: Han
authors.forename: Jonathan A
authors.initials: JA
authors.lastname: Javitch
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Bolan
authors.forename: David
authors.initials: D
authors.lastname: Kuraguntla
authors.forename: Vanaja
authors.initials: V
authors.lastname: Jaligam
authors.forename: Murat
authors.initials: M
authors.lastname: Oz
authors.forename: Lankupalle D
authors.initials: LD
authors.lastname: Jayanthi
authors.forename: Devadoss J
authors.initials: DJ
authors.lastname: Samuvel
authors.forename: Sammanda
authors.initials: S
authors.lastname: Ramamoorthy
authors.forename: Toni S
authors.initials: TS
authors.lastname: Shippenberg
mesh_terms: D002460:Cell Line; D004298:Dopamine / Q000378:metabolism*; D018491:Dopamine Agonists / Q000494:pharmacology; D050483:Dopamine Plasma Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D006801:Humans; D008856:Microscopy, Fluorescence; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D018377:Neurotransmitter Agents / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases; D019257:Quinpirole / Q000494:pharmacology; D050637:Receptors, Dopamine D3 / Q000235:genetics / Q000378:metabolism*; D015290:Second Messenger Systems / Q000187:drug effects / Q000502:physiology*
chemical_list: D018491:Dopamine Agonists; D050483:Dopamine Plasma Membrane Transport Proteins; D018377:Neurotransmitter Agents; D050637:Receptors, Dopamine D3; D019257:Quinpirole; D019869:Phosphatidylinositol 3-Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; D004298:Dopamine