An NMR study of cellular phosphates and membrane transport in renal proximal tubules.
An NMR study of cellular phosphates and membrane transport in renal proximal tubules.
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肾近曲小管细胞磷酸盐和膜转运的核磁共振研究。
DOI：
10.1152/ajprenal.1995.268.3.f375
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Brazy,PC
中科院分区：
文献类型：
--
作者：
Chobanian,MC;Anderson,ME;Brazy,PC
细胞磷酸盐测量的技术局限性阻碍了对肾近端小管（PT）细胞中细胞Pi、其转运及其代谢之间相互关系的研究。我们已经开发了一种非侵入性的31 P-核磁共振（NMR）探针灌注系统来测量细胞Pi，并利用该系统来研究犬PT细胞膜运输和细胞内容之间的关系Pi。在细胞外培养基中具有1.2mM Pi的情况下，PT的细胞Pi含量平均为4.94 +/-0.55nmol/mg蛋白质。通过去除细胞外Pi来抑制Pi摄取迅速地将所有细胞磷酸盐化合物降低至对照的55%至85%之间的值。部分替代细胞外Pi（0.4 mM）增加细胞磷酸盐高达84-100%的控制值。通过加入哇巴因抑制Na（+）-K（+）-腺苷三磷酸酶的摄取，未能改变细胞的Pi或有机磷酸盐。降低基底外侧膜电位与氯化钡的加入增加了近30%的细胞Pi含量。细胞Pi和ATP的最大含量在0.4 mM Pi的存在下，在向内定向的Na+梯度和在0.8 mM Pi的情况下，实现。这些数据表明，在犬PT细胞的Pi含量是由Na（+）依赖性和非依赖性的运输机制和跨基底外侧膜的膜电位。最后，发现细胞ATP含量在生理范围内与细胞Pi含量成正比。
Technical limitations in the measurement of cellular phosphates have hindered studies of interrelationships between cellular Pi, its transport, and its metabolism in renal proximal tubule (PT) cells. We have developed a noninvasive 31P-nuclear magnetic resonance (NMR) probe-perifusion system to measure cellular Pi and have utilized this system to investigate relationships in canine PT cells between the membrane transport and the cellular content of Pi. With 1.2 mM Pi in the extracellular medium, the cellular Pi content of PT averaged 4.94 +/- 0.55 nmol/mg protein. Inhibition of Pi uptake by removal of extracellular Pi rapidly decreased all cellular phosphate compounds to values that were between 55 and 85% of control. Partial replacement of extracellular Pi (0.4 mM) increased cellular phosphates up to 84-100% of control values. Inhibition of Na(+)-K(+)-adenosinetriphosphatase uptake by the addition of ouabain failed to change either cellular Pi or organic phosphates. Reducing the basolateral membrane potential with the addition of barium chloride increased cellular Pi content by nearly 30%. Maximal contents of cellular Pi and ATP were achieved at 0.4 mM Pi in the presence of an inwardly directed Na+ gradient and at 0.8 mM Pi in its absence. These data indicate that cellular Pi content in canine PT is regulated by Na(+)-dependent and -independent transport mechanisms and by the membrane potential across the basolateral membrane. Lastly, cellular ATP content was found to be directly proportional to the cellular Pi content over a physiological range.