Assessment of swelling-activated Cl- channels using the halide-sensitive fluorescent indicator 6-methoxy-N-(3-sulfopropyl)quinolinium.
Assessment of swelling-activated Cl- channels using the halide-sensitive fluorescent indicator 6-methoxy-N-(3-sulfopropyl)quinolinium.
复制标题
使用卤化物敏感荧光指示剂 6-甲氧基-N-(3-磺基丙基)喹啉鎓评估膨胀激活的 Cl-通道。
DOI：
10.1016/s0006-3495(98)77499-5
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Hughes,BA
中科院分区：
文献类型：
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作者：
Srinivas,SP;Bonanno,JA;Hughes,BA
本研究使用卤化物敏感荧光染料6-甲氧基- n -（3-磺丙基）喹啉（SPQ），定量分析了通过膨胀激活的Cl -通道增强阴离子渗透性。培养的牛角膜内皮单层灌注NO3 -林格氏氮，在等渗和随后的低渗条件下暴露于I -脉冲。在低渗条件下，由于I -内流引起的SPQ荧光变化明显快于等渗条件。在等渗和低渗条件下，质膜电位（Em）分别为- 58和- 32mV。结合Stern-Volmer方程（用于模拟SPQ荧光被I -猝灭）和Goldman通量方程（用于模拟电扩散单向I -内流），导出了低渗条件下与等渗条件下I -渗透率之比（称为增强比或ER）的表达式。在I−内流期间，SPQ荧光曲线拐点处的荧光值和斜率，以及在等渗和低渗条件下，被用来计算ER。基于这种方法，当低渗休克为110±10 mosM时，内皮细胞表达ER=4.9的肿胀激活Cl -通道。这些结果说明了基于spq的方法在单层中膨胀激活Cl -通道定量表征中的应用。
This study describes a quantitative analysis of the enhancement in anion permeability through swelling-activated Cl−channels, using the halide-sensitive fluorescent dye 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ). Cultured bovine corneal endothelial monolayers perfused with NO3−Ringer's were exposed to I−pulses under isosmotic and, subsequently, hyposmotic conditions. Changes in SPQ fluorescence due to I−influx were significantly faster under hyposmotic than under isosmotic conditions. Plasma membrane potential (Em) was −58 and −32mV under isosmotic and hyposmotic conditions, respectively. An expression for the ratio of I−permeability under hyposmotic condition to that under isosmotic condition (termed enhancement ratio or ER) was derived by combining the Stern-Volmer equation (for modeling SPQ fluorescence quenching by I−) and the Goldman flux equation (for modeling the electrodiffusive unidirectional I−influx). The fluorescence values and slopes at the inflection points of the SPQ fluorescence profile during I−influx, together withEmunder isosmotic and hyposmotic conditions, were used to calculate ER. Based on this approach, endothelial cells were shown to express swelling-activated Cl−channels with ER=4.9 when the hyposmotic shock was 110±10 mosM. These results illustrate the application of the SPQ-based method for quantitative characterization of swelling-activated Cl−channels in monolayers.