Dopamine transporter: Solubilization from dog caudate nucleus

Abstract

³H-GBR 12935 was used to label the digitonin-solubilized dopamine transporter from dog caudate nucleus. Specific binding to soluble fractions was observed in dog caudate but was absent in rat cerebellum. Binding to the solubilized transporter sites was saturable and of high affinity (B_max = 2.57 ± 0.60 pmol/mg protein, K_D = 23.42 ± 2.24 nM, n = 4). Heating or addition of trypsin abolished specific binding in the soluble fractions. In competition studies, soluble ³H-GBR 12935 binding was inhibited by mazindol, GBR 12909, nomifensine, dimethocaine, dopamine, (−) cocaine, and (+) cocaine in a manner typical of binding to the dopamine transporter. As expected, tomoxetine and citalopram, inhibitors of norepinephrine and serotonin uptake, respectively, were weak competitors of ³H-GBR 12935 binding.