Regulation of the D2-Like Dopamine Autoreceptor by the Dopamine Transporter.

Abstract

Despite its relatively low abundance in the brain, the neurotransmitter dopamine (DA) is vitally important for controlling motor coordination, motivation, reward, and cognition, among other processes. The amount of DA in the extracellular space determines the amount of DA signaling and is primarily controlled by two presynaptic proteins: the dopamine transporter (DAT), which removes DA from the extracellular space, and the D2-like dopamine autoreceptor. D2 autoreceptor decreases extracellular DA by inhibiting DA synthesis, decreasing DA exocytosis, and increasing DA reuptake by DAT. My thesis focuses on understanding the regulation of D2 autoreceptor and I determined that D2 autoreceptor regulation changes depending on its context in the membrane. D2 autoreceptor activation increases surface DAT localization, particularly in time of high neuronal stimulation, such as in response to natural rewards or abused drugs. I investigated the converse, DAT regulation of the D2 autoreceptor and found that co-expression of DAT with D2 in a heterologous cell system transforms the regulation of D2R through a novel D2R-DAT context. Within this context, less D2R was on the surface as compared to expression without DAT, an effect dependent on protein kinase C beta (PKCb) activity. The D2-DAT context was disrupted by removing PKC phosphorylation sites from D2 and DAT, suggesting PKC stabilizes this context. Normally, PKC causes internalization and desensitization of D2R; using PKCb knockout mice and specific PKCb inhibitors, I found that PKCb decreases D2 autoreceptor activity. Furthermore, when co-expressed with DAT, agonist stimulation of D2R increased surface D2R localization, reminiscent of the D2 autoreceptor-mediated increase in surface DAT localization. Interaction with DAT increases D2R signaling through ERK, perhaps through an arrestin-mediated mechanism. Because the D2 autoreceptor stimulated increase of DA uptake only occurs during neuronal burst firing, I propose that the D2 autoreceptor-DAT context is a mechanism to quickly decrease the extracellular DA concentration following burst firing through increased DA reuptake. During tonic DA release, D2 autoreceptor regulates extracellular DA by suppressing DA synthesis and exocytotic release. My results identify a novel, DAT-mediated mechanism for regulation of D2 autoreceptor and further our understanding of D2R regulation.