Winters C J, Reeves W B, Andreoli T E
Department of Internal Medicine, University of Arkansas College of Medicine, Little Rock 72205.
J Membr Biol. 1993 Aug;135(2):145-52. doi: 10.1007/BF00231440.
A unique property of basolateral membrane Cl- channels from the mTAL is that the Cl- concentration facing the intracellular aspects of these channels is a determinant of channel open time probability (Po). The K1/2 for maximal activation of Po by Cl- facing intracellular domains of these channels is 10 mM Cl-. The present experiments evaluated the nature of these Cl(-)-interactive sites. First, we found that the impermeant anion isethionate, when exposed to intracellular Cl- channel faces, could augment Po with a K1/2 in the range of 10 mM isethionate without affecting conductance (gCl, pS). Second, pretreatment of the solutions facing the intracellular aspects of the channels with either 1 mM phenylglyoxal (PGO), an arginine-specific reagent, or the lysine/terminal amine reagent trinitrobenzene sulfonic acid (TNBS, 1 mM), prevented the activation of Po usually seen when the Cl- concentration of solutions facing intracellular channel domains was raised from 2 to 50 mM. However, when the Cl- channel activity was increased by first raising the Cl- concentration bathing intracellular channel faces from 2 to 50 mM, subsequent addition of either PGO or TNBS to solutions bathing intracellular Cl- channel faces had no effect on Po. We conclude that the intracellular aspects of these Cl- channels contain Cl(-)-interactive loci (termed [Cl]i) which are accessible to impermeant anions in intracellular fluids and which contain arginine- and lysine-rich domains which can be inactivated, at low ambient Cl- or isethionate concentrations, by interactions with PGO or TNBS.
髓袢升支粗段基底外侧膜氯离子通道的一个独特特性是，这些通道细胞内面所面对的氯离子浓度是通道开放时间概率（Po）的一个决定因素。这些通道细胞内结构域面对的氯离子使Po最大激活的半数最大浓度（K1/2）为10 mM氯离子。本实验评估了这些氯离子相互作用位点的性质。首先，我们发现，当不可通透的阴离子羟乙基磺酸暴露于细胞内氯离子通道表面时，可增强Po，其K1/2在10 mM羟乙基磺酸范围内，且不影响电导（gCl，皮安）。其次，用1 mM苯乙二醛（PGO，一种精氨酸特异性试剂）或赖氨酸/末端胺试剂三硝基苯磺酸（TNBS，1 mM）预处理通道细胞内面的溶液，可阻止当通道细胞内结构域面对的溶液氯离子浓度从2 mM提高到50 mM时通常出现的Po激活。然而，当通过首先将细胞内通道表面的灌流氯离子浓度从2 mM提高到50 mM来增加氯离子通道活性时，随后向灌流细胞内氯离子通道表面的溶液中添加PGO或TNBS对Po没有影响。我们得出结论，这些氯离子通道的细胞内面含有氯离子相互作用位点（称为[Cl]i），细胞内液中的不可通透阴离子可接近这些位点，且这些位点含有富含精氨酸和赖氨酸的结构域，在低环境氯离子或羟乙基磺酸浓度下，通过与PGO或TNBS相互作用可使其失活。
