Abstract

Fluorophor and biotin derivatives of dopamine agonist and antagonist drugs were synthesized and evaluated for binding affinity and selectivity at D1 and D2 dopamine receptors in membranes prepared from monkey (Macaca fascicularis) caudate putamen. Binding was measured using [3H]SCH 23390 to label D1 receptors and [3H]spiperone to label D2 receptors. The selective D1 antagonist SKF 83566, whether coupled to 7-nitrobenz-2-oxa-1,3-diazole-4-yl (NBD), to fluorescein, or to biotin retained high affinity for D1 dopamine receptors (Ki, 5.3 16 and 3.5 nM, respectively) and high D1/D2 receptor selectivity (130-, 300, and 600-fold, respectively). The selective D2 antagonist derivative N-(p-aminophenethyl)spiperone, (NAPS) coupled either to biotin or to NBD via the N-aminoethylphenyl group, likewise retained high D2 receptor affinity (Ki, 0.58 and 0.66 nM, respectively) and high D2/D1 selectivity (190- and 150-fold, respectively). The affinity of the NBD-coupled derivative of (S)-2-(N-phenylethyl-N-propyl)-amino-5-hydroxytetralin hydrochloride [(S)-PPHT], a selective D2 agonist, was actually higher than that of the parent compound (Ki, 0.30 versus 2.1 nM), whereas the affinity of fluorescein-coupled (S)-PPHT was lower (Ki, 4.8 nM). Sensitivity to GTP, a characteristic of agonist binding at dopamine receptors, was demonstrated for NBD-coupled (S)-PPHT, because D2 receptor affinity was somewhat reduced in the presence of GTP. PPHT-fluorescein fluorescence labeling rimmed cells in monkey and rat anterior pituitary and outlined cells in the striatum. Fluorescent and biotin probes based on selective high affinity ligands for dopamine receptors may expedite studies of receptor localization and mobility at the cellular level.