Characterization of benzodiazepine receptors with fluorescent ligands.
Fluorescein conjugates of the high-affinity benzodiazepine receptor ligands Ro 15-1788 and Ro 7-1986 were synthesized. The binding of these fluorescent ligands (BD 621 and BD 607) to benzodiazepine receptors was characterized by direct fluorescence measurement. Both the equilibrium dissociation constants (KD) of BD 621 and BD 607 and the maximum number of binding sites (Bmax) estimated by fluorescence monitoring were consistent with values obtained by using radioligand binding techniques. The binding of BD 621 and BD 607 assessed by fluorescence measurement was reversible, abolished by photoaffinity labeling with Ro 15-4513, and unaffected by a variety of substances that do not bind to benzodiazepine receptors. The potencies of chemically diverse benzodiazepine receptor compounds to inhibit fluorescent ligand binding were highly correlated (r = 0.94, P less than 0.001), with potencies obtained from radioligand binding techniques. These findings demonstrate the feasibility of using direct fluorescence measurement techniques to quantitate ligand-receptor interactions.
McCabe
R T
RT
Laboratory of Neuroscience, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
de Costa
B R
BR
Miller
R L
RL
Havunjian
R H
RH
Rice
K C
KC
Skolnick
P
P
D000818:Animals; D001569:Benzodiazepines / Q000138:chemical synthesis* / Q000378:metabolism; D001921:Brain / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000378:metabolism; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D013058:Mass Spectrometry; D051381:Rats; D011919:Rats, Inbred Strains; D011963:Receptors, GABA-A / Q000378:metabolism*; D013329:Structure-Activity Relationship
D005456:Fluorescent Dyes; D008024:Ligands; D011963:Receptors, GABA-A; D001569:Benzodiazepines