The dopamine transporter (DAT) has been identified as a principal brain receptor site previously 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 years, investigators in this Branch have found that activators of protein kinase C (PKC) modulate dopamine transport in transiently-expressing COS cells and identified DAT as a phosphoprotein effected by several PKC, MAP kinase, MEKkinase and IP3 kinase agents. These studies have documented that the DAT regulation can occur as internalization of the transporter or as downregulation of the function of the transporter that remains expressed on cell surfaces, using activation of distince kinase pthways. Work during this year has also provided insights into functions of the human allelic variants in the DAT 5' flanking region. Human variants and haplotypes from this region have beeen identified by genomic resequencing. Large constructions containing a number of variants in each of the two most prominent haplotypes have been expressed in two cell expression systems as luciferase fusion constructions. Interestingly, these results provide evidence for a 30-50% haplotype-dependent difference in levels of DAT expression. In vivo PET and postmortem studies appear to confirm these observations in vivo. This work appears to complement studies of regulation at a protein phospphorylation level to provide a more complete picture of the state- and trait-dependent features of regulation of this important determinant of dopaminergic functions including those important for drug reward.
