Binding kinetics of gamma-aminobutyric acidA receptor noncompetitive antagonists: trioxabicyclooctane, dithiane, and cyclodiene insecticide- induced slow transition to blocked chloride channel conformation

JE Hawkinson and JE Casida

Department of Entomological Sciences, University of California, Berkeley 94720.

Binding kinetics and affinities are determined for 25 antagonists interacting with the noncompetitive blocker site of the gamma- aminobutyric acidA receptor complex present in bovine brain membranes. Four radiolabeled noncompetitive antagonists are 4-tert- butylbicyclophosphoro[35S]thionate ([35S]TBPS), 4-tert- butylbicycloortho[3',4'-3H2]benzoate, 4'-cyano-4-sec-[3,4- 3H2]butylbicycloorthobenzoate, and the new 4'-ethynyl-4-n-[2,3- 3H2]propylbicycloorthobenzoate. The other 21 antagonists are unlabeled inhibitors of three chemical classes (other trioxabicyclooctane, dithiane, and cyclodiene insecticides). The radioligands bind to a single noninteracting site in the membranes, based on linear Scatchard plots and monophasic association and dissociation kinetics. The kinetics of unlabeled ligands are estimated by their effect on the [35S]TBPS association curve, using the theoretical model of Motulsky and Mahan [Mol. Pharmacol. 25:1-9 (1984)]. The receptor affinities of trioxabicyclooctanes and dithianes correlate with their association rates, whereas those of cyclodienes correlate with their dissociation rates. The low association rate constants for all ligands (< or = 3 x 10(7) M-1 min-1 at 25 degrees) are consistent with a slow transition to a blocked receptor conformation upon binding of these channel blockers. The association rate-controlled affinity for the trioxabicyclooctanes and dithianes is suggestive of an induced-fit model in which binding of the ligand initiates a conformational change in the receptor complex to the blocked state.

Volume 42, Issue 6, pp. 1069-1076, 12/01/1992
Copyright © 1992 by American Society for Pharmacology and Experimental Therapeutics