Wu Q, Delpire E, Hebert S C, Strange K
Anesthesiology Research Division, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA.
Am J Physiol. 1998 Dec;275(6):C1565-72. doi: 10.1152/ajpcell.1998.275.6.C1565.
The function of the apical Na+-K+-2Cl- cotransporter in mammalian choroid plexus (CP) is uncertain and controversial. To investigate cotransporter function, we developed a novel dissociated rat CP cell preparation in which single, isolated cells maintain normal polarized morphology. Immunofluorescence demonstrated that in isolated cells the Na+-K+-ATPase, Na+-K+-2Cl- cotransporter, and aquaporin 1 water channel remained localized to the brush border, whereas the Cl-/HCO-3 (anion) exchanger type 2 was confined to the basolateral membrane. We utilized video-enhanced microscopy and cell volume measurement techniques to investigate cotransporter function. Application of 100 microM bumetanide caused CP cells to shrink rapidly. Elevation of extracellular K+ from 3 to 6 or 25 mM caused CP cells to swell 18 and 33%, respectively. Swelling was blocked completely by Na+ removal or by addition of 100 microM bumetanide. Exposure of CP cells to 5 mM BaCl2 induced rapid swelling that was inhibited by 100 microM bumetanide. We conclude that the CP cotransporter is constitutively active and propose that it functions in series with Ba2+-sensitive K+ channels to reabsorb K+ from cerebrospinal fluid to blood.
顶端Na⁺-K⁺-2Cl⁻协同转运蛋白在哺乳动物脉络丛（CP）中的功能尚不确定且存在争议。为了研究协同转运蛋白的功能，我们开发了一种新型的大鼠CP解离细胞制备方法，其中单个分离的细胞保持正常的极化形态。免疫荧光显示，在分离的细胞中，Na⁺-K⁺-ATP酶、Na⁺-K⁺-2Cl⁻协同转运蛋白和水通道蛋白1水通道仍定位于刷状缘，而2型Cl⁻/HCO₃（阴离子）交换体则局限于基底外侧膜。我们利用视频增强显微镜和细胞体积测量技术来研究协同转运蛋白的功能。应用100微摩尔布美他尼可使CP细胞迅速收缩。将细胞外K⁺浓度从3毫摩尔升高到6或25毫摩尔可使CP细胞分别肿胀18%和33%。Na⁺去除或添加100微摩尔布美他尼可完全阻止肿胀。将CP细胞暴露于5毫摩尔BaCl₂可诱导快速肿胀，100微摩尔布美他尼可抑制这种肿胀。我们得出结论，CP协同转运蛋白具有组成性活性，并提出它与Ba²⁺敏感的K⁺通道协同作用，将脑脊液中的K⁺重吸收至血液中。
