新生大鼠脑原代星形胶质细胞培养物的阳离子转运和膜电位特性

Cation transport and membrane potential properties of primary astroglial cultures from neonatal rat brains.

作者信息

Kimelberg H K, Bowman C, Biddlecome S, Bourke R S

出版信息

Brain Res. 1979 Nov 30;177(3):533-50. doi: 10.1016/0006-8993(79)90470-0.

Abstract

This paper describes K+ and Na+ content and transport in primary monolayer cultures from dissociated newborn rat brains, considered to consist predominantly of astroglial cells. Net changes in cation content after addition of ouabain, and steady state fluxes using 86Rb+ as a marker for K+ and 22Na+ as a marker for Na+, were measured. The results found indicate that the cells maintained a conventional pattern of cation homeostasis with net efflux of K+ being balanced by its active uptake and net uptake of Na+ balanced by active extrusion mediated by a ouabain sensitive (Na+K) pump. These processes maintained internal measured K+:Na+ ratios of 12--25:1. The cells were normally flat but addition of DBcAMP caused them to round up and form numerous processes, an appearance resembling that of astroglial cells in vivo. DBcAMP treatment also reduced the steady state levels of K+ measured with 86Rb+ by 15--30%, and had no effect on initial rates of 86Rb+ and 22Na+ uptake. The membrane potentials of cells treated with DBcAMP were studied, since only these were easily impaled. The membrane potentials of separate groups of cells gave means ranging from --65 to --75 mV at 35 degrees C, at an external K+ concentration ([K+]o) of 4.5 mM. The dependence of the membrane potentials of individual cells and groups of cells on [K+]o was studied. The slope of the potential per 10-fold change in [K+]o was 55--57 mV, at concentrations of K+ greater than 10--20 mM K+, and diverged from this slope at concentrations below this. This shows that these cells had some permeability to ions other than K+. Assuming that Na+ was the only other ion affecting the membrane potential, it was calculated that the permeability to Na+ was about 30 times less than K+. A similar result was obtained based on estimates of Na+ and K+ permeability from transport experiments on cells also treated with DBcAMP. The results obtained from these cells are compared to those found for other cultured glial cells and glial cells in vivo. We conclude that the membrane potentials of the cultured cells used in the present study show the closest resemblance so far to glia in vivo, since they are large and negative and are determined mainly by K+. However, the cultured cells have different properties from those reported in some studies for glial cells in vivo by showing free permeability to ions other than K+.

摘要

本文描述了来自新生大鼠脑解离后的原代单层培养物中钾离子（K⁺）和钠离子（Na⁺）的含量及转运情况，这些培养物主要由星形胶质细胞组成。测量了加入哇巴因后阳离子含量的净变化，以及使用⁸⁶Rb⁺作为K⁺的标志物和²²Na⁺作为Na⁺的标志物的稳态通量。研究结果表明，细胞维持了阳离子稳态的传统模式，K⁺的净外流通过其主动摄取得到平衡，而Na⁺的净摄取则通过由哇巴因敏感的（Na⁺ - K⁺）泵介导的主动外排得到平衡。这些过程维持了细胞内测得的K⁺:Na⁺比值为12 - 25:1。细胞通常是扁平的，但加入二丁酰环磷腺苷（DBcAMP）会使它们变圆并形成许多突起，这种外观类似于体内的星形胶质细胞。DBcAMP处理还使用⁸⁶Rb⁺测得的K⁺稳态水平降低了15% - 30%，并且对⁸⁶Rb⁺和²²Na⁺的初始摄取速率没有影响。对用DBcAMP处理的细胞的膜电位进行了研究，因为只有这些细胞容易被刺入。在35℃、外部钾离子浓度（[K⁺]o）为4.5 mM时，单独一组细胞的膜电位平均值在 - 65至 - 75 mV之间。研究了单个细胞和细胞组的膜电位对[K⁺]o的依赖性。在K⁺浓度大于10 - 20 mM K⁺时，[K⁺]o每10倍变化时膜电位的斜率为55 - 57 mV，而在低于此浓度时则偏离该斜率。这表明这些细胞对除K⁺以外的离子也有一定的通透性。假设Na⁺是影响膜电位的唯一其他离子，经计算其对Na⁺的通透性比K⁺约低30倍。基于对用DBcAMP处理的细胞进行的转运实验对Na⁺和K⁺通透性的估计，也得到了类似的结果。将这些细胞获得的结果与其他培养的胶质细胞以及体内胶质细胞的结果进行了比较。我们得出结论，本研究中使用的培养细胞的膜电位迄今为止显示出与体内胶质细胞最相似，因为它们数值大且为负，并且主要由K⁺决定。然而，这些培养细胞与一些关于体内胶质细胞的研究报道的细胞具有不同的特性，因为它们对除K⁺以外的离子具有自由通透性。