Formula omitted]t-butylbicycloorthobenzoate binding to recombinant α 1β 2γ 2s GABA A receptor

Abstract

The interaction of several selected compounds with the binding of the cage convulsant t- [ 3 H] butylbicycloorthobenzoate ( [ 3 H] TBOB) to membranes isolated from human embryonic kidney (HEK) 293 cells stably transfected with α 1β 2γ 2s subtype of GABA A receptors was studied. Scatchard analysis of binding data revealed the existence of a single type of binding site for [ 3 H] TBOB with a K d of 47.06±4.06 nM and a B max value of 6.72±0.52 pmol/mg protein. GABA, thiopental, TBOB, picrotoxin and the neurosteroid dehydroepiandrosterone sulfate displaced concentration—dependently the binding of [ 3 H] TBOB to this recombinant receptor. Dehydroepiandrosterone sulfate reversed the 5 μM GABA-induced inhibition of specific [ 3 H] TBOB binding. It is concluded that membranes isolated from HEK 293 cells stably transfected with α 1β 2γ 2s subunits exhibit specific high-affinity [ 3 H] TBOB binding. The potency of drugs to inhibit [ 3 H] TBOB binding mainly corresponded to that observed for the inhibition of the binding of cage convulsants to the native receptors or to transiently transfected HEK 293 cells.