Winters C J, Reeves W B, Andreoli T E
Department of Internal Medicine, University of Arkansas College of Medicine, Little Rock, USA.
Kidney Int. 1999 Apr;55(4):1444-9. doi: 10.1046/j.1523-1755.1999.00401.x.
This article reports studies on the kinetics of chloride (Cl-) conductance in Cl- channels fused into bilayers from basolaterally enriched vesicles from rabbit outer medulla. A considerable body of evidence indicates that these channels represent rbClC-Ka, a 77 kDa kidney-specific protein of the ClC family of Cl- channels. rbClC-Ka, a candidate channel for mediating net Cl- absorption in the medullary thick ascending limb (MTAL), has been cloned from rabbit outer medulla and localized by immunofluorescence to basolateral membranes of the MTAL. Thus, this is the first account, to our knowledge, of the kinetics of ion permeation through a renal Cl- channel mediating net basolateral Cl- absorption in the thick ascending limb of Henle (TALH), and this channel may represent rbClC-Ka.
The electrophysiological properties of these channels were studied by fusing basolaterally enriched MTAL vesicles into planar bilayer membranes.
Cl- conductance through these channels was concentration dependent and saturable. The relationship between gCl (pS) and symmetrical aqueous Cl- concentrations could be expressed in terms of the Michaelis equation with a limiting conductance (GClmax, pS) of 114 pS at infinitely high aqueous Cl- concentrations and a K1/2 of 163 mM Cl-. A log-log plot of the conductance-Cl- concentration relations, in the nonsaturating Cl- concentration range, had a slope of 0.91, that is, virtually unity. The relatively impermeant anion I- produced a voltage-dependent conductance blockade that could be overcome at high electric field strengths.
The experimental data described earlier here fulfill the traditional criteria for a first-order process with a single Cl- ion occupying these channels at a given time. Although the channels may contain multiple ion binding sites, the latter function, in integral kinetic terms, as a single rate-limiting locus.
本文报道了关于氯离子（Cl-）通道电导率动力学的研究，这些通道融合于从兔外髓质基底外侧富集的囊泡形成的双层膜中。大量证据表明，这些通道代表rbClC-Ka，它是Cl-通道ClC家族中一种77 kDa的肾脏特异性蛋白。rbClC-Ka是髓袢升支粗段（MTAL）介导净Cl-吸收的候选通道，已从兔外髓质克隆出来，并通过免疫荧光定位到MTAL的基底外侧膜。因此，据我们所知，这是首次对通过介导亨氏袢升支粗段（TALH）基底外侧净Cl-吸收的肾脏Cl-通道进行离子渗透动力学的描述，且该通道可能代表rbClC-Ka。
通过将基底外侧富集的MTAL囊泡融合到平面双层膜中来研究这些通道的电生理特性。
通过这些通道的Cl-电导率呈浓度依赖性且具有饱和性。gCl（pS）与对称水相Cl-浓度之间的关系可以用米氏方程表示，在无限高的水相Cl-浓度下，极限电导率（GClmax，pS）为114 pS，K1/2为163 mM Cl-。在非饱和Cl-浓度范围内，电导率-Cl-浓度关系的对数-对数图斜率为0.91，即几乎为1。相对不易通透的阴离子I-产生电压依赖性的电导率阻断，在高电场强度下可被克服。
本文前面所述的实验数据符合传统的一级过程标准，即在给定时间单个Cl-离子占据这些通道。尽管通道可能包含多个离子结合位点，但从整体动力学角度来看，后者作为单一的限速位点起作用。
