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Fluorescent probes have been designed and developed for dopamine D-1 and D-2 receptors. Fluorescein and/or NBD (7-nitrobenz-2-oxa-1,3-diazol-4-yl) derivatives of PPHT (D-2 agonist), spiperone (D-2 antagonist), SKF 38393 (D-1 agonist), and SKF 83566 (D-1 antagonist) were synthesized via their amino-functionalized analogues and all ligands were pharmacologically evaluated by measuring their ability to displace [3H]SCH 23390 and [3H]spiperone from D-1 and D-2 receptor sites in caudate putamen of monkeys (Macaca fascicularis). The fluorescein derivatives of PPHT and SKF 83566 and the NBD derivatives of spiperone and SKF 83566 retained the high affinity and selectivity of the parent ligands. The NBD derivatives of PPHT showed higher D-2 receptor affinity and selectivity than their parent ligands. The enantiomers of the fluorescent derivatives of PPHT were also synthesized and were found to exhibit stereoselectivity in binding to the D-2 receptor, with the S enantiomers having a considerably higher affinity than their R analogues. In contrast to these results, the fluorescein derivative of SKF 38393 showed only a low affinity for the D-1 receptor. These fluorescein- and NBD-coupled D-1 and D-2 receptor ligands have considerable significance as potential probes in the study of distribution of the receptors at the cellular/subcellular level and of their mobility in membranes in normal/diseased states by use of fluorescence microscopic and fluorescence photobleaching recovery techniques, respectively. The development of these novel fluorescent probes should also provide new leads for the design and synthesis of additional fluorescent ligands with better fluorescent properties and/or higher affinity/selectivity for the DA receptors.

Receptor-selective fluorescent ligands have come into increasing use as scientific tools for the study of receptor physiology and pathophysiology at the cellular and even the subcellular level. Furthermore, they are being increasingly investigated as tools in drug discovery research. In both cases, techniques employing receptor-selective fluorescent ligands have proved to be complementary to, and in several cases even superior to, the traditional-radioligand based techniques. increasing costs and public concerns associated with radioactive isotope use and dispersal are also making the use of fluoresecent ligands more attractive in research and diagnostic use. With the increasing rate of discovery of new receptors and receptor subtypes and newer more potent and/or selective receptor ligands and drugs, there will be an accompanying need for the design and development of new highly potent and selective fluorescent ligands to aid in the investigation of the physiological and pathophysiological functions of these new receptors and also in the development of drugs acting specifically at these receptors. This review presents a background of development of selected fluorescent ligands, primarily those for small molecule and small peptide based ligands. o 1994 \ViIey-Li

Substituted 1-phenyl-3-benzazepines (e.g., SKF 38393 and fenoldopam) exhibit stereoselectivity in moderately high-affinity binding to and partial agonist activation of D1 dopamine receptors. The 3-allyl (APB) and the 3-allyl-6-chloro (6-Cl-APB) analogues of SKF 38393 are reported to have higher affinity and selectivity for the D1 DA receptor and higher in vivo central neuropharmacologic activity than SKF 38393. We recently reported the corresponding 3-allyl-6-bromo analogue (6-Br-APB) also to be a high-affinity D1 agonist. We now describe the synthesis and characterization of the R-(+) and S-(-) enantiomers of both APB and 6-Br-APB and their comparison with corresponding enantiomers of SKF 38393 with respect to D1 receptor binding affinity and D1 and D2 selectivity. The R-(+) enantiomers of both novel substituted 1-phenyl-3-benzazepines bound to the D1 receptor sites in rat forebrain tissue with much higher affinity and selectivity than their S-(-) antipodes. R-(+)-3-Allyl-6-bromo-7,8-dihydroxy-1-phenyl-2,3,4,5-tetrahydro-1H-3- benzazepine [(R)-(+)-6-Br-APB, 18] exhibits the highest affinity of the reported 1-phenyl-3-benzazepine D1 agonists.

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