Effects of cocaine and repeated cocaine followed by withdrawal. Alterations of dopaminergic transporter turnover with no changes in kinetics of substrate recognition.

The turnover of the transporter for dopamine (ca. 1.5 sec-1) and the apparent second order rate constant of association of dopamine with the outward facing form of the transporter protein (ca. 10(6) M-1sec-1) were estimated using kinetic data from rotating disk voltammetric measures of the inward transport of dopamine in striatal suspensions, standard treatments of the kinetics of transport, and values in the literature for density of striatal transporter sites. Under apparent steady-state conditions of transporter functioning, inhibition of the transport of dopamine by cocaine following its addition to the incubation buffer was found to decrease the turnover of the transporter and not affect the kinetics of substrate recognition. The kinetics of binding of dopamine to the transporter were estimated also by apparent pre-steady-state kinetics. These experiments confirmed the second order nature of the binding of dopamine to the transporter and the numerical value of the rate constant estimated under steady-state conditions; they also demonstrated that the binding of dopamine has an absolute dependence on Na+, and that the second order rate constant of association of dopamine with its transporter is not influenced by cocaine. In separate studies, similar experiments were conducted in tissues from animals that had been treated with cocaine for 3 days and withdrawn for 1 day or 2 weeks. Repeated treatments with cocaine followed by either a 24-hr or 2-week period of withdrawal resulted in increases in the Vmax and turnover of the transporter with no apparent changes in the kinetics of association of substrate. No differences between the Ki for cocaine observed in direct inhibitions of the transport of dopamine and the Ki for cocaine observed in tissues obtained from animals treated repeatedly with cocaine were observed. Taken together, these data suggest that cocaine exerts its effects by altering an intramembrane translocation step for the movement of dopamine and not by changing the recognition of dopamine by the externally facing binding site or the apparent Ki for cocaine. Finally, repeated treatments with cocaine followed by a period of withdrawal appear to kinetically activate the transporter for dopamine.