J. Neurochem. (2012) 123, 288–297.
Amphetamine (AMPH) and methamphetamine (METH) alter dopamine transporter (DAT) function. In vitro heterologous cell line and synaptosome studies demonstrate AMPH-induced DAT internalization, implicating relocalization in reduced DAT uptake following drug exposure. However, few studies have evaluated DAT localization following in vivo AMPH/METH administration. To determine DAT subcellular localization following drug administration, a centrifugation technique was developed to isolate striatal synaptosomal membrane and vesicle fractions. DAT was distributed between the synaptosomal membrane (60%) and endosomal vesicles (40%), and in vitro application of the protein kinase C activator phorbol 12-myristate 13-acetate to striatal synaptosomes caused DAT internalization into the vesicle fractions. In contrast, neither single nor repeated in vivo AMPH and/or METH administrations altered DAT localization 5, 15, 30, or 60 min post-treatment, despite reduced DAT uptake. Importantly, repeated METH injections uniformly decreased total DAT immunoreactivity within all fractions 7 days post-treatment. These findings suggest that factors other than internalization can contribute to the observed acute and persistent DAT dysfunction and dopaminergic deficits following in vivo AMPH or METH administration.