DOPAMINE TRANSPORTER I--STRUCTURE/FUNCTION STUDIES

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The dopamine transporter (DAT) has been identified as the principal brain receptor site best correlated with the rewarding and euphoric properties of cocaine. Euphoric responses to rapid administration of cocaine can be much more prominent than those that follow slower rates of administration. In previous FYs, investigators in this Branch have found that activators of protein kinase C (PKC) modulate dopamine transport in transiently-expressing COS cells (Eur. J. Pharmacol., 268, 115-119). In this FY, we have begun to explore the extent to which direct phosphorylation of the DAT may produce the sorts of acute adaptations that could yield differing responses to rapid and slow rates of transporter occupancy. We have replicated effects of PKC activators in LLC-PK1 cells that stably express DAT, and used specific anti-DAT antisera developed in previous FYs to immunoprecipitate phosphor-DAT from striatal slice and stably transfected cell preparations. Initial evidence suggests that PKC activation does enhance levels of DAT phosphorylation, and thus increases evidence for phosphorylation as a candidate mechanism for rapid adaptations in dopaminergic systems relevant to cocaine-induced euphoria.