Hamano H, Nagata K, Fukada T N, Shimotahira H, Ju X L, Ozoe Y
Department of Life Science and Biotechnology, Shimane University, Matsue, Japan.
Bioorg Med Chem. 2000 Mar;8(3):665-74. doi: 10.1016/s0968-0896(00)00009-2.
Acyclic noncompetitive antagonists of ionotropic gamma-aminobutyric acid (GABA) receptors, bearing an ester or ether linkage, were designed, synthesized, and assayed for their inhibition of the specific binding of [3H]4'-ethynyl-4-n-propylbicycloorthobenzoate (EBOB), a radiolabeled noncompetitive antagonist, to rat brain and housefly head membranes. 5-[4-(3,3-Dimethylbutoxycarbonyl)phenyl]-4-pentynoic acid (DBCPP), a butyl benzoate analogue, was found to competitively inhibit the binding of [3H]EBOB in rat brain membranes, with an IC50 of 88 nM. The potency conferred by the p-substituent decreased in the order C(triple bond)C(CH2)2COOH > C(triple bond)C(CH2)2COOCH3 > C(triple bond) CH > Br. Pentyl phenyl ethers were equally potent compared with butyl benzoates, while phenyl pentanoates and benzyl butyl ethers were less pont. These compounds were generally less active in housefly head membranes than in rat brain membranes. The introduction of an isopropyl group into the 1-position of the 3,3-dimethylbutyl group of a butyl benzoate and two benzyl butyl ethers caused an increase in potency in housefly GABA receptors, whereas this modification at the corresponding position of other compounds led to an unchanged or decreased potency. In the case of rat receptors, this modification resulted in a decrease in potency except for a phenyl pentanoate. To confirm that DBCPP interferes with GABA receptor function, we performed whole-cell patch clamp experiments with rat dorsal root ganglion neurons in the primary culture. Repeated co-applications of GABA and DBCPP suppressed GABA-induced whole-cell currents with an IC50 of 0.54 microM and a Hill coefficient of 0.7. These findings indicate that DBCPP and its derivatives inhibit ionotropic GABA receptors by binding to the EBOB site and that there might be structural difference in the noncompetitive antagonist-binding site between rat and housefly GABA receptors.
设计、合成了带有酯键或醚键的离子型γ-氨基丁酸（GABA）受体的无环非竞争性拮抗剂，并检测了它们对放射性标记的非竞争性拮抗剂[3H]4'-乙炔基-4-正丙基双环邻苯二甲酸酯（EBOB）与大鼠脑和家蝇头部膜特异性结合的抑制作用。发现5-[4-(3,3-二甲基丁氧基羰基)phenyl]-4-戊炔酸（DBCPP），一种苯甲酸丁酯类似物，能竞争性抑制[3H]EBOB与大鼠脑膜的结合，IC50为88 nM。对位取代基赋予的效力按C（三键）C(CH2)2COOH > C（三键）C(CH2)2COOCH3 > C（三键）CH > Br的顺序降低。戊基苯基醚与苯甲酸丁酯的效力相当，而苯基戊酸酯和苄基丁基醚的效力较低。这些化合物在家蝇头部膜中的活性通常比在大鼠脑膜中低。在苯甲酸丁酯和两种苄基丁基醚的3,3-二甲基丁基的1-位引入异丙基会导致家蝇GABA受体的效力增加，而在其他化合物的相应位置进行这种修饰会导致效力不变或降低。对于大鼠受体，这种修饰除了苯基戊酸酯外会导致效力降低。为了证实DBCPP干扰GABA受体功能，我们对原代培养的大鼠背根神经节神经元进行了全细胞膜片钳实验。重复共同应用GABA和DBCPP抑制了GABA诱导的全细胞电流，IC50为0.54 microM，希尔系数为0.7。这些发现表明DBCPP及其衍生物通过与EBOB位点结合来抑制离子型GABA受体，并且大鼠和家蝇GABA受体的非竞争性拮抗剂结合位点可能存在结构差异。
