Orlov Iu N, Bresler V M, Kazbekov E N, Sukhodolova A T
Tsitologiia. 1987 Mar;29(3):365-8.
Basolateral membrane vesicles were isolated from the rat kidney cortex by a modified method of cation precipitation. Different steps of preparation were analysed using the marker enzymes: Na+,K+-ATPase (for basolateral membrane), alkaline phosphatase (for apical membrane), glucose-6-phosphatase (for membranes of endoplasmic reticulum) and succinate dehydrogenase (for mitochondria). The basolateral membrane was purified by a 8-9-fold treatment with Na+,K+-ATPase, while other membrane contaminations were as low as 2% (as compared to homogenate). The transport of 3H-p-aminohippurate (3H-PAH) by basolateral membrane vesicles was measured under different experimental conditions. The 3H-PAH uptake was found to be Na-gradient dependent. The initial rate of 3H-PAH uptake in the presence of NaCl gradient (500 pM/mg X min) was higher than without the gradient (88 pM/mg X min). It is concluded that the PAH transfer across the basolateral membrane may be energized by the Na+ chemical gradient.
采用改良的阳离子沉淀法从大鼠肾皮质中分离出基底外侧膜囊泡。使用标记酶对制备的不同步骤进行分析:Na⁺,K⁺-ATP酶(用于基底外侧膜)、碱性磷酸酶(用于顶端膜)、葡萄糖-6-磷酸酶(用于内质网膜)和琥珀酸脱氢酶(用于线粒体)。通过用Na⁺,K⁺-ATP酶进行8至9倍处理来纯化基底外侧膜,而其他膜污染低至2%(与匀浆相比)。在不同实验条件下测量基底外侧膜囊泡对³H-对氨基马尿酸(³H-PAH)的转运。发现³H-PAH摄取依赖于Na⁺梯度。在存在NaCl梯度(500 pM/mg×分钟)时³H-PAH摄取的初始速率高于不存在梯度时(88 pM/mg×分钟)。得出的结论是,PAH跨基底外侧膜的转运可能由Na⁺化学梯度提供能量。