Pericić D, Mirković K, Jazvinsćak M, Besnard F
Ruder Bosković Institute, Laboratory for Molecular Neuropharmacology, Zagreb, Croatia.
Eur J Pharmacol. 1998 Oct 30;360(1):99-104. doi: 10.1016/s0014-2999(98)00661-x.
The interaction of several selected compounds with the binding of the cage convulsant t-[3H]butylbicycloorthobenzoate ([3H]TBOB) to membranes isolated from human embryonic kidney (HEK) 293 cells stably transfected with alpha1beta2gamma2s subtype of GABA(A) receptors was studied. Scatchard analysis of binding data revealed the existence of a single type of binding site for [3H]TBOB with a Kd of 47.06+/-4.06 nM and a Bmax value of 6.72+/-0.52 pmol/mg protein. GABA, thiopental, TBOB, picrotoxin and the neurosteroid dehydroepiandrosterone sulfate displaced concentration-dependently the binding of [3H]TBOB to this recombinant receptor. Dehydroepiandrosterone sulfate reversed the 5 microM GABA-induced inhibition of specific [3H]TBOB binding. It is concluded that membranes isolated from HEK 293 cells stably transfected with alpha1beta2gamma2s subunits exhibit specific high-affinity [3H]TBOB binding. The potency of drugs to inhibit [3H]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.
研究了几种选定化合物与笼形惊厥剂t-[3H]丁基双环邻苯二甲酸酯（[3H]TBOB）与人胚肾（HEK）293细胞稳定转染GABA(A)受体α1β2γ2s亚型后分离的膜结合之间的相互作用。结合数据的Scatchard分析显示存在一种单一类型的[3H]TBOB结合位点，其解离常数（Kd）为47.06±4.06 nM，最大结合量（Bmax）值为6.72±0.52 pmol/mg蛋白质。γ-氨基丁酸（GABA）、硫喷妥钠、TBOB、印防己毒素和硫酸脱氢表雄酮以浓度依赖性方式取代[3H]TBOB与该重组受体的结合。硫酸脱氢表雄酮逆转了5 microM GABA诱导的特异性[3H]TBOB结合抑制。得出的结论是，从稳定转染α1β2γ2s亚基的HEK 293细胞中分离的膜表现出特异性高亲和力的[3H]TBOB结合。药物抑制[3H]TBOB结合的效力主要与观察到的笼形惊厥剂与天然受体或瞬时转染的HEK 293细胞结合抑制情况相对应。
