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In Vitro Assays for the Functional Characterization of the Dopamine Transporter (DAT)
Journal article   Open access

In Vitro Assays for the Functional Characterization of the Dopamine Transporter (DAT)

Shaili Aggarwal and Ole V Mortensen
Current protocols in pharmacology, v 79(1), pp 12.17.1-12.17.21
20 Dec 2017
PMID: 29261226
url
https://europepmc.org/articles/pmc5741089View
Accepted (AM)Open Access (License Unspecified) Open

Abstract

Animals Biological Assay Cell Line Dopamine - metabolism Dopamine Plasma Membrane Transport Proteins - metabolism Dopamine Uptake Inhibitors - pharmacology Dose-Response Relationship, Drug Humans Kinetics Ligands
Detailed in this unit are protocols for studying the in vitro uptake of dopamine (DA) as a means for defining the functional characteristics of dopamine transporters. All assays are performed using commercially available cell lines that transiently express the transporter under investigation. The three main assays provided are: a kinetic assay to calculate the affinity (K ) and maximal velocity (V ) of radiolabeled DA uptake into cells; concentration-response assays to measure the potencies (IC /K values) of test compounds as transport inhibitors; and an efflux assay to assess the ability and potency (EC ) of a ligand to elicit reverse transport of DA accumulated in the cell. Although the methods are described using DAT and its ligands, the same procedure can be employed for studying serotonin and norepinephrine transporters as well. © 2017 by John Wiley & Sons, Inc.

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