Amphetamine-induced Dopamine Efflux

Abstract

Amphetamine (AMPH) elicits its behavioral effects by acting on the dopamine (DA) transporter (DAT) to in- duce DA overflow into the synaptic cleft. Facilitated exchange diffusion is the classical model used to de- scribe AMPH-induced DA efflux. This model hypothe- sizes that AMPH-induced DA efflux is mediated by DAT and results from the transport of AMPH into the cell followed by a counter movement of DA out to the extra- cellular compartment. To further characterize the ac- tion ofAMPH, we used the patch clamp technique in the whole-cell configuration combined with amperometry on human embryonic kidney HEK-293 cells stably trans- fected with the human DAT (DAT cells). In DAT cells, AMPH-induced DAT-mediated currents were blocked by cocaine. We demonstrate that DA efflux mediated by DAT is voltage-dependent, electrogenic, and dependent on intracellular Naⴙ concentration in the recording electrode. Intracellular Naⴙ fluorescence, as measured by confocal microscopy using a Naⴙ-sensitive dye, was enhanced by AMPH application. Furthermore, the abil- ity ofAMPH to induce DA efflux was regulated by intra- cellular Naⴙ concentration and correlated with the size of the DAT-mediated, AMPH-induced ion flux across the plasma membrane. In the absence of intracellular Naⴙ but the presence of high intracellular Clⴚ, AMPH-in- duced inward currents elicited DA efflux proportionally to their dimension and duration. Thus, we propose that AMPH-induced DA efflux depends on two correlated transporter processes. First, AMPH binds to the DAT and is transported, thereby causing an inward current. Second, because of this AMPH-induced inward current, Naⴙ becomes more available intracellularly to the DAT, thereby enhancing DAT-mediated reverse transport of DA. The