? The dopamine (DA) transporter (DAT) is the primary mechanism of removing released dopamine from the synaptic cleft. Improper DAT function or expression has been correlated with many clinical pathologies, including schizophrenia and Parkinson's disease. Of particular interest is regulation due to substance abuse since the DAT is the primary site of action of psychostimulants such as cocaine and amphetamines. Activation of the DA D2 type receptor initiates many signaling cascades, including G-protein mediated stimulation of phospholipase A2 (PLA2) and adenylyl cyclase. PLA2 liberates arachidonic acid (AA) from membrane phospholipids. The AA signaling cascade has been implicated in mediating psychostimulant induced sensitization, thus it is important to examine the role of AA in DAT regulation. In this proposal we focus on the hypothesis that PLA2 generated AA regulates DAT function and cellular localization in rat primary neuronal DA cell cultures.
The specific aims of this proposal are: 1) to characterize the effect of exogenously added A.A on DAT function and localization, 2) examine the effect of PLA2 activation and production of endogenous AA on DAT regulation, and 3) determine the effect of DA receptor activation on PLA2 mediated DAT regulation. The proposed experiments will use radioligand uptake assays to measure AA mediated changes in DAT function. Cellular expression of the DAT will be determined using fluorescent confocal microscopy and live cell imaging will be used to visualize membrane trafficking. These studies will be important for determining the role of the AA signaling cascade in regulating dopaminergic transmission via the DAT. Additional understanding concerning mechanisms of substance abuse will be gained through these experiments. ? ? ? ?
| Keith, Dove J; Wolfrum, Katherine; Eshleman, Amy J et al. (2012) Melittin initiates dopamine transporter internalization and recycling in transfected HEK-293 cells. Eur J Pharmacol 690:13-21 |
| Keith, Dove J; Eshleman, Amy J; Janowsky, Aaron (2011) Melittin stimulates fatty acid release through non-phospholipase-mediated mechanisms and interacts with the dopamine transporter and other membrane-spanning proteins. Eur J Pharmacol 650:501-10 |
