HIV-1 Tat protein-induced rapid and reversible decrease in [3H]dopamine uptake: dissociation of [3H]dopamine uptake and [3H]2beta-carbomethoxy-3-beta-(4-fluorophenyl)tropane (WIN 35,428) binding in rat striatal synaptosomes.

Abstract

Human immunodeficiency virus (HIV)-1 Tat protein plays a key role in the pathogenesis of both HIV-1-associated cognitive-motor disorder and drug abuse. Dopamine (DA) transporter (DAT) function is strikingly altered in patients with HIV-1-associated dementia and a history of chronic drug abuse. This study is the first in vitro evaluation of potential mechanisms underlying the effects of Tat protein on DAT function. Rat striatal synaptosomes were incubated with recombinant Tat(1-86) protein, and [(3)H]DA uptake and the binding of [(3)H]2beta-carbomethoxy-3-beta-(4-fluorophenyl)tropane (WIN 35,428) and [(3)H]1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)-piperazine (GBR 12935) were determined. Tat decreased [(3)H]DA uptake, [(3)H]WIN 35,428 binding, and [(3)H]GBR 12935 binding in a time-dependent manner. The potency of Tat for inhibiting [(3)H]DA uptake (K(i) = 1.2 microM) was the same as that for inhibiting [(3)H]GBR 12935 binding but 3-fold less than that for inhibiting [(3)H]WIN 35,428 binding. Mutant Tat proteins did not alter [(3)H]DA uptake. Kinetic analysis of [(3)H]DA uptake revealed that Tat (1 or 10 microM) decreased the V(max) value and increased the K(m) value in a dose-dependent manner. The V(max) value, decreased by Tat (1 microM), returned to the control level after washout of Tat, indicating that the inhibitory effect of Tat on DA uptake was reversible. Saturation studies revealed that Tat decreased the B(max) value and increased the K(d) value of [(3)H]WIN 35,428 binding, whereas Tat decreased the B(max) value of [(3)H]GBR 12935 binding, without a change in the K(d) value. These findings provide new insight into understanding the pharmacological mechanisms of Tat-induced dysfunction of the DAT in the dopaminergic system in HIV-infected patients.