Subunit-dependent interaction of propoxazepam and its metabolite with the -aminobuturic acid type A receptor

Abstract

Benzodiazepines (BDZ) are widely used in clinics in the treatment of psychiatric disorders, and their main action is considered to be determined by more selective binding with α1, α2, α3 or α5 subunits of GABA receptor. The aim of this work was studying of the molecular mechanism of action of new analgesic – propoxazepam and its metabolite (3-hydroxypropoxazepam) on α1, α2, α3, α4 or α5 subunits containing GABAA channels. Materials and methods GABA ha1b3g2, ha2b3g2, ha3b3g2, ha4b3g2 and ha5b3g2 ionotropic GABAARs expressed in HEK293 were used on the automated SP384PE Patch Clamp system. In addition, Propoxazepam, 3-hydroxypropoxazepam, diazepam (positive allosteric modulator) and GABA (positive control) were administered at concentrations 0.001–300 nM to determine the EC50 and Emax for corresponding substances. Results The α subunit plays a significantl role in determining the receptor’s affinity for propoxazepam and 3-hydroxypropoxazepam. The rank order of decreasing EC50 are α1 = α5> α2 > α3 > α4 (propoxazepam) and α1> α2> α5 > α3 > α4 (3-hydroxypropoxazepam), and for Emax α3 > α2 >α5 > α1 > α4 (propoxazepam), α3 > α1 > α2> α5 > α4 (3-hydroxypropoxazepam). The data, transformed to Emax/EC50, show that propoxazepam exhibits tenfold (compared to diazepam) activity (taking into account the magnitude of the maximum effect) to the α3 subunit, which distinguishes it from 3-hydroxypropoxazepam. Conclusion Due to the determined selectivity of propoxazepam for binding with different α subunit-containing GABAA-receptors (mostly α3 and α2 types), it has the potential to provide analgesia with less sedation than non-selective BDZ.