Novel Fluorescent Ligands Enable Single-Molecule Localization Microscopy of the Dopamine Transporter.

ABSTRACT: The dopamine transporter (DAT) is critical for spatiotemporal control of dopaminergic neurotransmission and is the target for therapeutic agents, including ADHD medications, and abused substances, such as cocaine. Here, we develop new fluorescently labeled ligands that bind DAT with high affinity and enable single-molecule detection of the transporter. The cocaine analogue MFZ2-12 (1) was conjugated to novel rhodamine-based Janelia Fluorophores (JF₅₄₉ and JF₆₄₆). High affinity binding of the resulting ligands to DAT was demonstrated by potent inhibition of [³H]dopamine uptake in DAT transfected CAD cells and by competition radioligand binding experiments on rat striatal membranes. Visualization of binding was substantiated by confocal or TIRF microsc

SUBMITTER: Guthrie DA 

PROVIDER: S-EPMC8283659 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature