甲状腺激素和血清对大鼠脑培养物中钠钾 - 三磷酸腺苷酶的发育及相关离子通量的影响

Effect of thyroid hormone and serum on the development of Na+, K+-adenosine triphosphatase and associated ion fluxes in cultures from rat brain.

作者信息

Atterwill C K, Atkinson D J, Bermudez I, Balazs R

出版信息

Neuroscience. 1985 Jan;14(1):361-73. doi: 10.1016/0306-4522(85)90185-x.

Abstract

The effect of culture conditions, serum supplementation or chemically defined medium and the influence of thyroid hormone were studied on the development of the Na+, K+-adenosine triphosphatase (Na+,K+-ATPase) and on the intracellular content of K+ and Na+ ions in cultures which either were greatly enriched in a neuronal cell type, the cerebellar granule cells, or contained a mixed population of cells (brain reaggregates). Foetal rat brain reaggregates displayed lower Na+,K+-ATPase activity when cultured in chemically defined medium than in the presence of serum. Supplementation of the serum-free medium with thyroid hormone resulted in a rise in the Na+,K+-ATPase activity and [3H]ouabain binding to levels similar to those found in the cultures grown in the serum-containing medium. Thyroid hormone had no significant effect on the Mg2+-ATPase activity and on the intracellular content of Na+ and K+ ions. In the granule cell-enriched cerebellar surface cultures the Na+,K+-ATPase activity was lower when the cells were grown in chemically defined medium compared with the serum-containing medium, and the intracellular Na+ to K+ ratio was higher. Thyroid hormone had no effect on the Na+,K+-ATPase activity, [3H]ouabain binding or Mg2+-ATPase activity. The hormone also failed to influence ATPase activities in cerebellar astrocytes maintained in chemically defined medium. Although thyroid hormone had no effect on the Na+,K+-ATPase activity of cultured cerebellar granule cells, treatment with the hormone resulted in a decrease in the ratio of intracellular Na+ to K+ ion content. The effect of the hormone on the Na+,K+-pump activity in live cells was therefore tested by estimating ouabain-sensitive 86Rb uptake. This was regulated as in other cell types, by the rate of Na+ entry: the Na+-ionophore monensin trebled the rate of 86Rb uptake, which was also increased (+30-100%) by 10% foetal calf serum, the maximal response being obtained by about 20 min exposure to serum. The effect was completely blocked by the Na+/H+ exchange inhibitor amiloride. The factor(s) in the serum responsible for the regulation of the Na+,K+-pump were, however, not the thyroid hormones, which failed to affect 86Rb uptake. On the basis of comparing thyroid hormone effects on the different cultures studied it was concluded that not every type of neural cell is target of the hormone action during development.

摘要

研究了培养条件、血清补充或化学成分明确的培养基以及甲状腺激素的影响，这些因素对富含神经元细胞类型（小脑颗粒细胞）的培养物或含有混合细胞群体（脑重聚体）中Na⁺，K⁺ - 腺苷三磷酸酶（Na⁺，K⁺ - ATP酶）的发育以及K⁺和Na⁺离子的细胞内含量的影响。胎鼠脑重聚体在化学成分明确的培养基中培养时，其Na⁺，K⁺ - ATP酶活性低于在有血清存在的情况下。在无血清培养基中添加甲状腺激素会导致Na⁺，K⁺ - ATP酶活性以及[³H]哇巴因结合增加至与在含血清培养基中生长的培养物中发现的水平相似。甲状腺激素对Mg²⁺ - ATP酶活性以及Na⁺和K⁺离子的细胞内含量没有显著影响。在富含颗粒细胞的小脑表面培养物中，与含血清培养基相比，当细胞在化学成分明确的培养基中生长时，Na⁺，K⁺ - ATP酶活性较低，且细胞内Na⁺与K⁺的比率较高。甲状腺激素对Na⁺，K⁺ - ATP酶活性、[³H]哇巴因结合或Mg²⁺ - ATP酶活性没有影响。该激素也未能影响在化学成分明确的培养基中培养的小脑星形胶质细胞的ATP酶活性。尽管甲状腺激素对培养的小脑颗粒细胞的Na⁺，K⁺ - ATP酶活性没有影响，但用该激素处理会导致细胞内Na⁺与K⁺离子含量的比率降低。因此，通过估计哇巴因敏感的⁸⁶Rb摄取来测试该激素对活细胞中Na⁺，K⁺ - 泵活性的影响。其调节方式与其他细胞类型相同，受Na⁺进入速率的影响：Na⁺离子载体莫能菌素使⁸⁶Rb摄取速率增加两倍，10%胎牛血清也使其增加（30% - 100%），暴露于血清约20分钟可获得最大反应。该效应被Na⁺/H⁺交换抑制剂氨氯地平完全阻断。然而，血清中负责调节Na⁺，K⁺ - 泵的因素不是甲状腺激素，甲状腺激素未能影响⁸⁶Rb摄取。基于比较甲状腺激素对所研究的不同培养物的影响，得出结论：并非每种神经细胞类型在发育过程中都是该激素作用的靶点。