The long-term goal of this research is to develop selective, fluorescent probes for delta, kappa, and mu opioid receptor types. The probes will be designed so that functioning opioid receptors can be visualized using the confocal scanning laser microscope. This goal will be pursued by the use of selective pharmacopheres derived from naltrindole (delta), norbinaltorphimine (kappa), and oxymorphone and naltrexone (mu). Three different colored fluorophores will be attached to the pharmacophores through strategically placed hydrophilic spacers so that affinity, selectivity, and potency of the ligands are retained. The different colored fluorophores will be employed in an effort to ultimately image three opioid receptor types simultaneously. These ligand will be evaluated in four in vitro preparations: binding in guinea pig brain membranes, antagonist and agonist potencies in the guinea pig ileum and mouse vas deferens, and confocal microscopic visualization in three cell populations (NGl08-l5, SH-SY5Y, guinea pig cerebellum) that contain defined opioid receptor populations. Promising ligands will then undergo antinociceptive testing and, ultimately, neural tissues from the tested animals will be examined to evaluate the ability of the probe to image opioid receptor populations in vivo. The attainment of this long-range goal will afford an armamentarium of selective opioid receptor probes for scanning laser confocal microscopy.
