Cole L M, Roush R T, Casida J E
Department of Environmental Science, Policy and Management, University of California, Berkeley 94720-3112.
Life Sci. 1995;56(10):757-65. doi: 10.1016/0024-3205(95)00006-r.
The non-competitive blocker site of the GABA-gated chloride ion channel in normal susceptible strains of Drosophila melanogaster and simulans binds 4-n-[3H]propyl-4'-ethynylbicycloorthobenzoate ([3H]EBOB) at specific sites with KdS of 1.6-1.9 nM and BmaxS of 171-181 fmol/mg protein. This specific binding of [3H]EBOB is strongly inhibited by: a large number and variety of insecticidal channel blockers at 20 nM (lindane, alpha-endosulfan, dieldrin, 12-ketoendrin, fipronil, and a representative bicycloorthobenzoate and dithiane) or 200 nM (picrotoxinin); the insecticidal channel activators avermectin and moxidectin at 20 nM; muscimol at 30 microM and GABA at 300 microM. Cyclodiene resistance in D. melanogaster has been attributed to a mutation resulting in an Ala302-->Ser replacement in the Rdl GABA receptor subunit and in D. simulans to an homologous Ala-->Ser or Gly replacement. These mutations are shown here to greatly reduce [3H]EBOB binding, i.e. lower affinity and apparent number of binding sites. The Ala-->Ser replacement with both melanogaster and simulans almost always reduces the potency in inhibiting [3H]EBOB binding of each of eight channel blockers and of muscimol and GABA. The Ala-->Gly replacement in D. simulans is generally less effective than the Ala-->Ser modification in reducing sensitivity to the channel blockers and to muscimol and GABA. The channel activators avermectin and moxidectin usually retain their inhibitory potency in the Rdl subunit mutants. Thus, it appears that replacement of Ala by Ser generally modifies the non-competitive blocker site and its coupling to the GABA-recognition site with less effect on the channel activator site. In contrast, the Ala-->Gly replacement has less impact in protecting the chloride channel from the action of insecticidal blockers. Each of the resistant strains has the same level of resistance to the lethal action of the five channel blockers examined but none to avermectins and muscimol.
在正常敏感品系的黑腹果蝇和拟果蝇中，GABA门控氯离子通道的非竞争性阻断剂位点在特定位点结合4 - n - [³H]丙基 - 4'-乙炔基双环邻苯二甲酸酯（[³H]EBOB），解离常数（KdS）为1.6 - 1.9 nM，最大结合量（BmaxS）为171 - 181 fmol/mg蛋白质。[³H]EBOB的这种特异性结合受到以下物质的强烈抑制：20 nM的大量且种类繁多的杀虫通道阻断剂（林丹、α - 硫丹、狄氏剂、12 - 酮狄氏剂、氟虫腈以及一种代表性的双环邻苯二甲酸酯和二噻烷）或200 nM的印防己毒素；20 nM的杀虫通道激活剂阿维菌素和莫昔克丁；30 μM的蝇蕈醇和300 μM的GABA。黑腹果蝇中的环二烯抗性归因于一种突变，该突变导致Rdl GABA受体亚基中的Ala302被替换为Ser，而在拟果蝇中则是同源的Ala被替换为Ser或Gly。此处显示这些突变会极大地降低[³H]EBOB的结合，即降低亲和力和结合位点的表观数量。黑腹果蝇和拟果蝇中Ala被替换为Ser几乎总是会降低八种通道阻断剂以及蝇蕈醇和GABA对[³H]EBOB结合的抑制效力。拟果蝇中Ala被替换为Gly在降低对通道阻断剂以及蝇蕈醇和GABA的敏感性方面通常不如Ala被替换为Ser有效。通道激活剂阿维菌素和莫昔克丁在Rdl亚基突变体中通常保留其抑制效力。因此，似乎Ser替换Ala通常会改变非竞争性阻断剂位点及其与GABA识别位点的偶联，而对通道激活剂位点的影响较小。相比之下，Ala被替换为Gly在保护氯离子通道免受杀虫阻断剂作用方面的影响较小。每个抗性品系对所检测的五种通道阻断剂的致死作用具有相同水平的抗性，但对阿维菌素和蝇蕈醇均无抗性。
