Sackin H, Palmer L G
Am J Physiol. 1987 Sep;253(3 Pt 2):F476-87. doi: 10.1152/ajprenal.1987.253.3.F476.
Potassium (K+) channels in the basolateral membrane of unperfused Necturus proximal tubules were studied in both cell-attached and excised patches, after removal of the tubule basement membrane by manual dissection without collagenase. Two different K+ channels were identified on the basis of their kinetics: a short open-time K+ channel, with a mean open time less than 1 ms, and a long open-time K+ channel with a mean open time greater than 20 ms. The short open-time channel occurred more frequently than the longer channel, especially in excised patches. For inside-out excised patches with Cl- replaced by gluconate, the current-voltage relation of the short open-time K+ channel was linear over +/- 60 mV, with a K+-Na+ selectivity of 12 +/- 2 (n = 12), as calculated from the reversal potential with oppositely directed Na+ and K+ gradients. With K-Ringer in the patch pipette and Na-Ringer in the bath, the conductance of the short open-time channel was 47 +/- 2 pS (n = 15) for cell-attached patches, 26 +/- 2 pS (n = 15) for patches excised (inside out) into Na-Ringer, and 36 +/- 6 pS (n = 3) for excised patches with K-Ringer on both sides. These different conductances can be partially explained by a dependence of single-channel conductance on the K+ concentration on the interior side of the membrane. In experiments with a constant K+ gradient across excised patches, large changes in Na+ at the interior side of the membrane produced no change in single-channel conductance, arguing against a direct block of the K+ channel by Na+. Finally, the activity of the short open-time channel was voltage gated, where the mean number of open channels decreased as a linear function of basolateral membrane depolarization for potentials between -60 and 0 mV. Depolarization from -60 to -40 mV decreased the mean number of open K+ channels by 28 +/- 8% (n = 6).
在不使用胶原酶、通过手工剥离去除肾小管基底膜后,采用细胞贴附式和膜片钳全细胞式记录方法,对未灌注的美西螈近端肾小管基底外侧膜中的钾(K+)通道进行了研究。根据其动力学特性鉴定出两种不同的K+通道:一种是开放时间短的K+通道,平均开放时间小于1毫秒;另一种是开放时间长的K+通道,平均开放时间大于20毫秒。开放时间短的通道比开放时间长的通道出现得更频繁,尤其是在膜片钳全细胞式记录中。对于用葡萄糖酸盐替代Cl-的外向式膜片钳全细胞式记录,开放时间短的K+通道的电流-电压关系在+/- 60 mV范围内呈线性,根据反向的Na+和K+梯度下的反转电位计算,K+-Na+选择性为12 +/- 2(n = 12)。当膜片钳微管中为K-林格液、浴槽中为Na-林格液时,对于细胞贴附式膜片,开放时间短的通道的电导为47 +/- 2 pS(n = 15);对于外向式(外翻式)记录到Na-林格液中的膜片,电导为26 +/- 2 pS(n = 15);对于两侧均为K-林格液的膜片钳全细胞式记录,电导为36 +/- 6 pS(n = 3)。这些不同的电导可以部分地通过单通道电导对膜内侧K+浓度的依赖性来解释。在对膜片钳全细胞式记录施加恒定K+梯度的实验中,膜内侧Na+的大幅变化并未引起单通道电导的改变,这表明Na+不会直接阻断K+通道。最后,开放时间短的通道的活性受电压门控,在-60至0 mV的电位范围内,开放通道的平均数量随着基底外侧膜去极化呈线性减少。从-60 mV去极化到-40 mV使开放K+通道的平均数量减少了28 +/- 8%(n = 6)。