Neurotoxic Dopamine Transporter Substrates Differentially Affect DAT Regulation, Uptake, and Reverse Transport
Authors: Stanislowski Daniel J. , Johnson Brett M. , Adkins Steve D. , Vaughan Roxanne A. , Foster James D. 
Published: June 2021

Synaptic action of the neurotransmitter dopamine (DA) is primarily terminated by reuptake into presynaptic dopaminergic neurons via dopamine transporters (DATs). DATs are integral plasmalemmal proteins whose reuptake function is tightly coordinated by interplay between the post translational modifications phosphorylation and palmitoylation. The psychostimulant amphetamine (AMPH) is a DAT substrate known to empty sequestered DA from vesicles and induce protein kinase C (PKC)‐dependent DAT phosphorylation, resulting in reverse transport of DA (efflux) greatly increased DA signaling. Elevated DA concentrations similar to those produced by AMPH create neurotoxic conditions as DA oxidation produces free radicals and hinders mitochondrial respiration, thus strict spatial and temporal control of cytosolic and synaptic DA is essential for neuronal health. DA induced oxidative stress is an important pathogenic mediator in Parkinson's disease (PD), and animal models of PD are generated by the neurotoxins 1‐methyl‐4‐phenylpyridinium ion (MPP+) and 6‐hydroxydopamine (6OHDA) which selectively target DAT‐expressing neurons, are translocated to the cytosol by DAT, and perturb the oxidative balance within the cell, producing nigrostriatal DA neuron death observed in PD. It is therefore intriguing to investigate DAT‐involved mechanisms in terms of PD etiology as atypical DAT function could contribute to DA oxidative stress and is hypothesized to play a role in other neurological disorders including attention deficit/hyperactivity disorder (ADHD) and addiction. DAT expression occurs around synaptic terminals, along axons, and on dendritic spines, and efflux via DATs located on dendrites in the substantia nigra has been shown to diminish dopaminergic signaling. It is possible, therefore, for aberrant regulation and localization of DA clearance and DAT‐mediated DA efflux to profoundly influence the excitability of neurons and play a significant role in neurodegeneration in the progression of PD. This is even more interesting in the light of recent studies revealing DAT polymorphisms in patients diagnosed with a psychiatric disorder or early onset PD that display an elevated basal DA efflux phenotype. In this study, we are investigating the effect of DAT substrates and PD‐inducing neurotoxins on DAT phosphorylation, palmitoylation, down‐regulation, and reverse transport to better understand aberrant DAT mechanisms which may contribute the onset of PD. Our findings thus far indicate MPP+ and 6OHDA differentially alter DAT function. 6OHDA inhibits basal and attenuates PMA‐induced DAT phosphorylation, does not significantly reduce DAT function, and decreases DA efflux. MPP+ does not change basal or PMA stimulated DAT phosphorylation yet it induces significant DA efflux which may be an additional means by which MPP+ produces neurodegeneration. This MPP+ induced efflux was not blunted by pretreatment with the PKC inhibitor bisindolylmaleimide (BIM). Although AMPH‐induced efflux was blunted after BIM treatment, significant efflux remained suggesting mechanisms independent of PKC mediated DAT phosphorylation for the stimulation of DA efflux.

Support or Funding Information

Supported by the National Institute on Drug Abuse Grant DA 031991 JDF, P20 RR017699 (to U.N.D.) from the COBRE program of the National Center for Research Resources, and P20 RR016741 (to U.N.D.) from the INBRE program of the National Center for Research Resources.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.