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Effect of Diethyl Pyrocarbonate Modification of Benzodiazepine Receptors on [ 3 H]Ro 15‐4513 Binding
Author(s) -
UusiOukari Mikko
Publication year - 1992
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1992.tb09406.x
Subject(s) - chemistry , diazepam , flunitrazepam , gabaa receptor , benzodiazepine , binding site , stereochemistry , receptor , pharmacology , medicinal chemistry , biochemistry , medicine
The effects of treatment of brain membranes with diethyl pyrocarbonate (DEP), a histidine‐modifying reagent, on the binding of 3 H‐labeled Ro 15‐4513 (ethyl‐8‐azido‐5,6‐dihydro‐5‐methyl‐6‐oxo‐4 H ‐imidazo[1,5‐ a ]‐[1,4]benzodiazepine‐3‐carboxylate) and [ 3 H]diazepam were compared. DEP pretreatment produced a dose‐dependent decrease in [ 3 H]diazepam binding, whereas low DEP concentrations enhanced the binding of [ 3 H]Ro 15‐4513. These effects were reversed by incubation with hydroxylamine after the treatment. The enhancement of [ 3 H]Ro 15‐4513 binding was due to an increase in the affinity of the binding sites ( K D ), without any effect on binding capacity ( B max . The enhancement was perceived in cerebral cortical, cerebellar, and hippocampal membranes. DEP treatment decreased the displacement of [ 3 H]Ro 15‐4513 binding by diazepam and FG 7142 (N‐niethyl‐β‐carboline‐3‐carboxamide) but not by Ro 15‐4513 and Ro 19‐4603 ( tert ‐butyl‐5,6‐dihydro‐5‐methyl‐6‐oxo‐4 H ‐imidazo[1,5‐ a ]thieno[2,3‐ f ][1,4]diazepine‐3‐carboxylate). Although the stimulating effect of γ‐aminobutyric acid (GABA) on [ 3 H]‐diazepam binding was not affected by DEP treatment, such treatment reduced the inhibitory effect of GABA on [ 3 H]Ro 15‐4513 binding. The enhancement of [ 3 H]Ro 15‐4513 binding was observed in membranes pretreated with DEP in the presence of flunitrazepam, whereas such pretreatment reduced significantly the inhibitory effect of DEP on [ 3 H]‐diazepam binding. The results indicate that a histidine residue in the benzodiazepine binding domain is critical for [ 3 H]diazepam but not [ 3 H]Ro 15‐4513 binding. They also suggest that the cerebellar granule cell‐specific “diazepam‐insensitive” [ 3 H]Ro 15‐4513 binding sites lack this critical histidine.