6,3′-Dinitroflavone is a low efficacy modulator of GABAA receptors

6,3′-Dinitroflavone (6,3′-DNF) is a synthetic flavone derivative that exerts anxiolytic effects in the elevated plus maze. Based on the finding that this effect is blocked by Ro15-1788 (ethyl-8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate) which is a specific antagonist at the benzodiazepine binding site of GABAA receptors we investigated the interaction of 6,3′-DNF with several recombinant GABAA receptor subtypes. Inhibition of [3H]flunitrazepam binding to recombinant GABAA receptors in transiently transfected HEK293 cells indicated that 6,3′-DNF exhibited the highest affinity for GABAA receptors composed of α1β2γ2 subunits and a 2–20 fold lower affinity for homologous receptors containing α2, α3, or α5 subunits. Two-electrode voltage-clamp experiments in Xenopus oocytes indicated that 6,3′-DNF does not induce chloride flux in the absence of GABA, but exerts low efficacy inverse agonistic modulatory effects on GABA-elicited currents in the GABAA receptor subtypes α1β2γ2 and α5β2γ2. In the subtypes α2β2γ2, α3β2γ2, α4β2γ2, α6β2γ2 or α4β2δ and α4β3δ, 6,3′-DNF exerts either none or very low efficacy positive modulatory effects. In contrast, 100 nM Ro15-1788 exhibited weak to moderate partial agonistic effects on each receptor investigated. These data indicate that Ro15-1788 only can antagonize the weak inverse agonist effects of 6,3′-DNF on α1β2γ2 and α5β2γ2 receptors, but will enhance the weak agonistic effects on the other receptor subtypes investigated. The possible mechanism of the Ro15-1788 sensitive anxiolytic effect of 6,3′-DNF is discussed.