Design, Synthesis, and Characterization of New δ Opioid Receptor-Selective Fluorescent Probes and Applications in Single-Molecule Microscopy of Wild-Type Receptors

The delta opioid receptor (δOR or DOR) is a G protein-coupled
receptor (GPCR) showing a promising profile as a drug target for nociception
and analgesia. Herein, we design and synthesize new fluorescent antagonist
probes with high δOR selectivity that are ideally suited for
single-molecule microscopy (SMM) applications in unmodified, untagged
receptors. Using our new probes, we investigated wild-type δOR
localization and mobility at low physiological receptor densities
for the first time. Furthermore, we investigate the potential formation
of δOR homodimers, as such a receptor organization might exhibit
distinct pharmacological activity, potentially paving the way for
innovative pharmacological therapies. Our findings indicate that the
majority of δORs labeled with these probes exist as freely diffusing
monomers on the cell surface in a simple cell model. This discovery
advances our understanding of OR behavior and offers potential implications
for future therapeutic research.