Sherstobitov A O, Gusev G P
Laboratory of Comparative Biochemistry of Inorganic Ions, Sechenov Institute, Russian Academy of Sciences, St Petersburg, Russia.
J Exp Biol. 1998 Mar;201(Pt 5):693-700.
The existence of a furosemide-sensitive Cl- transport pathway activated by external Ca2+ and Mg2+ has been demonstrated previously in studies of Cl- influx across the lamprey erythrocyte membrane. The aim of the present study was to characterize further specific Cl- transport pathways, especially those involved in Cl- efflux, in the red blood cell membrane of Lampetra fluviatilis. Cl- efflux was inhibited by 0.05 mmol l-1 dihydroindenyloxyalkanoic acid (DIOA) (81%), 1 mmol l-1 furosemide (76%) and 0.1 mmol l-1 niflumic acid (54%). Bumetanide (100 mumol l-1) and DIDS (100 mumol l-1) had no effect effect on Cl- efflux. Substitution of external Cl- by gluconate, but not by NO3-, led to a gradual decline of Cl- efflux. In addition, the removal of external Ca2+ resulted in a significant reduction in the rate of Cl- efflux. Membrane depolarization caused by increasing external K+ concentration or by inhibiting K+ channels with 1 mmol l-1 Ba2+ did not affect Cl- efflux. The furosemide-sensitive component of Cl- influx was a saturable function of external [Cl-] with an apparent K(m) of approximately 92 mmol l-1 and Vmax of approximately 17.8 mmol l-1 cells-1 h-1. Furosemide did not affect intracellular Cl- concentration (57.6 +/- 5.2 mmol l-1 cell water), measured using an ion-selective Cl- electrode, showing that a furosemide-sensitive pathway is not involved in net Cl- movement. A gradual fall (from 28.1 +/- 1.4 to 15.0 +/- 1.3 mmol l-1 cells-1 h-1) in unidirectional Cl- influx with time was observed within 3 h of cell preincubation in the standard physiological medium. These data provide evidence for the existence for an electroneutral furosemide-sensitive anion-exchange pathway in the lamprey erythrocyte membrane that accepts chloride and nitrate, but not bicarbonate or bromide.
先前在研究氯离子跨七鳃鳗红细胞膜内流时已证明，存在一种由细胞外钙离子和镁离子激活的对呋塞米敏感的氯离子转运途径。本研究的目的是进一步表征河七鳃鳗红细胞膜中特定的氯离子转运途径，尤其是那些参与氯离子外流的途径。0.05 mmol l-1的二氢茚氧基链烷酸（DIOA）（81%）、1 mmol l-1的呋塞米（76%）和0.1 mmol l-1的氟尼辛（54%）可抑制氯离子外流。布美他尼（100 μmol l-1）和二异硫氰酸二苯乙烯酯（DIDS）（100 μmol l-1）对氯离子外流无影响。用葡萄糖酸盐而非硝酸盐替代细胞外氯离子会导致氯离子外流逐渐下降。此外，去除细胞外钙离子会导致氯离子外流速率显著降低。通过增加细胞外钾离子浓度或用1 mmol l-1的钡离子抑制钾离子通道引起的膜去极化不影响氯离子外流。氯离子内流的呋塞米敏感成分是细胞外[Cl-]的饱和函数，表观米氏常数（K(m)）约为92 mmol l-1，最大反应速度（Vmax）约为17.8 mmol l-1细胞-1 h-1。使用离子选择性氯离子电极测量发现，呋塞米不影响细胞内氯离子浓度（57.6 ± 5.2 mmol l-1细胞内液），这表明呋塞米敏感途径不参与氯离子的净移动。在标准生理培养基中对细胞进行预孵育3小时内，观察到单向氯离子内流随时间逐渐下降（从28.1 ± 1.4降至15.0 ± 1.3 mmol l-1细胞-1 h-1）。这些数据为七鳃鳗红细胞膜中存在一种电中性的、对呋塞米敏感的阴离子交换途径提供了证据，该途径可接受氯离子和硝酸盐，但不接受碳酸氢根或溴离子。
