Mechanism of Inhibition of the GluA2 AMPA Receptor Channel Opening by 2,3-Benzodiazepine Derivatives

Abstract

2,3-Benzodiazepine (2,3-BDZ) compounds are a group of AMPA receptor inhibitors and are drug candidates for treating neurological diseases involving excessive AMPA receptor activity. We investigated the mechanism by which GluA2Qflip receptor channel opening is inhibited by two 2,3-BDZ derivatives, i.e. (2,3-BDZ-11-2) and its 1-(4-amino-3-chlorophenyl) analogue (2,3-BDZ-11-4). Both compounds have a 7,8-ethylenedioxy moiety instead of the 7,8-methylenedioxy feature in the structure of GYKI 52466, the prototypic 2,3-BDZ compound. Using a laser-pulse photolysis approach with a time resolution of ∼60 μs and a rapid solution flow technique, we characterized the effect of the two compounds on the channel-opening process of the homomeric GluA2Qflip receptor. Both 2,3-BDZ-11-2 and 2,3-BDZ-11-4 inhibit the receptor noncompetitively with specificity for the closed-channel conformation of the GluA2Qflip receptor. 2,3-BDZ-11-4 is ∼10-fold stronger, defined by its inhibition constant for the closed-channel conformation (i.e., KI = 2 μM), than 2,3-BDZ-11-2. From double-inhibitor experiments, we find that both compounds bind to the same site, but this site is different from the two other noncompetitive sites we reported earlier (Ritz et al., 2008, Ritz et al., 2011). In other words, 2,3-BDZ-11-2 and 2,3-BDZ-11-4 do not bind to either the site where GYKI 52466 binds or the site where 2,3-BDZ-2 binds (see their structures below). Our results provide mechanistic clues to better understand AMPA receptor regulation and establish a structure-activity relationship for designing more potent 2,3-BDZ compounds with predictable properties for this new noncompetitive site.