Photoaffinity labeling of [3H]flunitrazepam- and [3H]Ro15-4513-bound pellets in rat cerebral cortex and cerebellum.
Irreversible incorporation of [3H]flunitrazepam and [3H]Ro15-4513 into GABA/benzodiazepine receptor subunits was studied by UV irradiation using ligand-bound membrane pellets from rat cerebral cortical and cerebellar synaptic membranes. Specific incorporation for [3H]flunitrazepam was greater in the pellet than in the suspension. The incorporation was identical for [3H]Ro15-4513 in both pellet and suspension. With the ligand-bound pellets, 50% of the available binding sites were photolabeled by both ligands in cortex and cerebellum. SDS polyacrylamide gel electrophoresis and fluorography of [3H]flunitrazepam photo-labeled receptor revealed the same number of major sites in both brain regions. In contrast, [3H]Ro15-4513 appears to label fewer sites in cortex and cerebellum. Photoaffinity labeling with [3H]flunitrazepam in ligand-bound membrane pellet provides a more selective and reliable method for studying the subunit structure of GABA/benzodiazepine receptor complex.
Chiu
T H
TH
Department of Pharmacology, Medical College of Ohio, Toledo 43699.
Yu
O F
OF
Rosenberg
H C
HC
D000345:Affinity Labels; D000818:Animals; D001386:Azides / Q000378:metabolism*; D001569:Benzodiazepines / Q000378:metabolism*; D002531:Cerebellum / Q000378:metabolism*; D002540:Cerebral Cortex / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005445:Flunitrazepam / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011963:Receptors, GABA-A / Q000032:analysis*; D013535:Suspensions; D013570:Synaptic Membranes / Q000378:metabolism; D013997:Time Factors
D000345:Affinity Labels; D001386:Azides; D011963:Receptors, GABA-A; D013535:Suspensions; D001569:Benzodiazepines; D005445:Flunitrazepam; C042957:Ro 15-4513