The role of transcriptional and translational mechanisms in flumazenil-induced up-regulation of recombinant GABAA receptors

The aim of this study was to further elucidate the mechanisms involved in adaptive changes of GABAA receptors following prolonged exposure to flumazenil, the antagonist of benzodiazepine binding sites on GABAA receptors. The effects of prolonged flumazenil treatment were studied on recombinant α1β2γ2S GABAA receptors stably expressed in human embryonic kidney (HEK 293) cells. Using radioligand binding experiments we found enhancement in the maximum number of [3H]muscimol labeled binding sites in different preparations of HEK 293 cells. The parallel increase of [3H]flunitrazepam binding sites in the membranes was reduced in the presence of actinomycin D and cycloheximide, inhibitors of RNA and protein synthesis, respectively. Chronic flumazenil also raised the steady-state level of mRNA encoding α1 receptor subunit. The results suggest that the up-regulation of GABAA receptors, observed after prolonged flumazenil treatment is at least partly due to increased de novo synthesis of receptor proteins at both transcriptional and translational level.