Localization of sodium pump sites in frog urinary bladder.
Localization of sodium pump sites in frog urinary bladder.
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青蛙膀胱钠泵部位的定位。
DOI：
10.1016/0005-2736(75)90194-7
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发表时间：
1975
期刊：
影响因子：
--
通讯作者：
Stephan A. Ernst
中科院分区：
文献类型：
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作者：
John W. Mills;Stephan A. Ernst
用短路电流法、闪烁计数法和放射自显影法研究了[3 H]哇巴因在蛙和蟾蜍膀胱中的结合。SCC的数据和分析的组织细胞后seradine暴露于哇巴因表明，哇巴因的结合和抑制Na+转运是完全可逆的蟾蜍膀胱，而在青蛙膀胱，[3 H]哇巴因是紧密结合和Na+转运仍然受到抑制，即使在60分钟的冲洗。粘液暴露的青蛙膀胱[3 H]哇巴因或seradine暴露后预孵育未标记的哇巴因导致显着减少结合。通过调节cecrtain（Na+−K+）-ATP酶配体（K+，ATP）的浓度至已知降低哇巴因结合的水平，进一步评估结合的特异性。高K+浓度和缺氧条件下孵育的内源性ATP耗竭导致[3 H]哇巴因结合显着下降。放射自显影分析表明，颗粒主要定位于基底侧质膜的颗粒细胞，提供直接的形态学证据的位置Na+泵在这些网站。虽然放射自显影没有提供足够的分辨率，以排除明确哇巴因结合到富含线粒体的细胞，形态学证据表明，颗粒密度显着高于相邻的颗粒细胞之间的颗粒细胞-线粒体丰富的细胞界面。
[3H]Ouabain binding in frog and toad urinary bladder was investigated by short-circuit current (SCC), scintillation counting and authoradiographic techniques. SCC data and analysis of tissue digests following serosal exposure to ouabain showed that ouabain binding and inhibition of Na+transport was completely reversible in toad bladder whereas, in frog bladder, [3H] ouabain was tightly bound and Na+transport remained suppressed even after a 60-min washout. Mucosal exposure of frog bladder to [3H]ouabain or serosal exposure after preincubation with unlabeled ouabain led to a marked reduction in binding. Specificity of binding was assessed further by adjusting the concentration of cecrtain (Na+−K+)-ATPase ligands (K+, ATP) to levels known to reduce ouabain binding. High K+concentrations and depletion of endogenous ATP by incubation under anoxic conditions resulted in a significant drop in [3H]ouabain binding. Autoradiographic analysis showed that grains are localized primarily to the basolateral plasma membranes of the granular cells, providing direct morphological evidence for the location of Na+pumps at these sites. Although autoradiographs did not provide sufficient resolution to rule out unequivocally ouabain binding to the mitochondria-rich cell, morphological evidence suggests that grain densities are significatly higher between adjacent granular cells than between granular cell-mitochondria-rich cell interfaces.