Role of the histidine residue at position 105 in the human α5 containing GABAA receptor on the affinity and efficacy of benzodiazepine site ligands

A histidine residue in the N-terminal extracellular region of α1,2,3,5 subunits of the human GABAA receptor, which is replaced by an arginine in α4 and α6 subunits, is a major determinant for high affinity binding of classical benzodiazepine (BZ)-site ligands. The effect of mutating this histidine at position 105 in the α5 subunit to an arginine (α5H105R) on BZ-site pharmacology has been investigated using radioligand binding on HEK293 and L(tk-) cells and two electrode voltage clamp recording on Xenopus oocytes in which GABAA receptors of subtypes α5, α5H105R, α4 and α6 were co-expressed with β3γ2s. The classical BZs, diazepam and flunitrazepam (full agonists on the α5 receptor) showed negligible affinity and therefore negligible efficacy on α5H105R receptors. The β-carbolines DMCM and βCCE (inverse agonists on the α5 receptor) retained some affinity but did not exhibit inverse agonist efficacy at α5H105R receptors. Therefore, the α5H105R mutation confers an α4/α6-like pharmacology to the classical BZs and β-carbolines. Ro15-4513, flumazenil, bretazenil and FG8094, which share a common imidazobenzodiazepine core structure, retained high affinity and were higher efficacy agonists on α5H105R receptors than would be predicted from an α4/α6 pharmacological profile. This effect was antagonized by DMCM, which competes for the BZ-site and therefore is likely to be mediated via the BZ-site. These data indicate that the conserved histidine residue in the α subunit is not only a key determinant in the affinity of BZ-site ligands on α5 containing GABAA receptors, but also influences ligand efficacy. British Journal of Pharmacology (2002) 135, 248–256; doi:10.1038/sj.bjp.0704459