A multisubstrate kinetic mechanism of dopamine transport in the nucleus accumbens and its inhibition by cocaine.

Kinetic studies of dopamine transport into suspensions of nucleus accumbens (NAcc) and effects of Na+ and Cl- as cosubstrates were performed using rotating disk electrode voltammetry. To mimic chemical neurotransmission, dopamine was added as a rapid pulse, and transporter-mediated clearance of dopamine was evaluated kinetically. This paradigm was shown to approximate a zero trans entry transport experiment. Dopamine was taken up with apparent Km and Vmax values of 1.3 microM and 375 pmol/s/g wet weight, respectively. Transport exhibited apparent trans acceleration. Substitution of Na+ with choline or Cl- with isethionate reduced dopamine transport with reaction orders of two and unity, respectively, accompanied by reductions in Vmax with no changes in Km. Apparent K(Na) and K(Cl) values were 70.0 and 92.1 mM, respectively. Dopamine transport in NAcc was found to follow a partially random, sequential mechanism in which dopamine and Na+ bind randomly to the transporter followed by binding of Cl- before transport. Cocaine inhibited dopamine transport and the influences of the other substrates allosterically with an overall Ki of 0.30 microM. Thus, the general kinetic mechanism of the transport of dopamine in the NAcc is identical to that previously reported by this laboratory for dopamine transport in the striatum. However, the dopamine transporter in the NAcc is more tightly regulated by Na+, possesses a higher kinetic turnover rate, is four times more sensitive to cocaine than the striatal transporter, and exhibits cocaine inhibition independent of [substrate]. These findings suggest that cocaine modulates chemical signaling in NAcc differently than in striatum, providing down-regulation of function irrespective of [substrate], thereby enhancing dopaminergic signaling more robustly in the NAcc than in the striatum.