Atterwill C K, Cunningham V J, Balázs R
J Neurochem. 1984 Jul;43(1):8-18. doi: 10.1111/j.1471-4159.1984.tb06672.x.
The activities of certain properties of sodium, potassium-activated adenosine triphosphatase (Na+,K+-ATPase; EC 3.6.1.3) were examined in cultures and perikarya fractions enriched in rat cerebellar nerve cells or astrocytes, in comparison with preparations from whole immature and adult rat cerebellum and derived synaptosomal fractions, as well as nonneural tissue such as the kidney. The specific activity of Na+,K+-ATPase was markedly higher in the freshly isolated astrocytes than in the nerve cells (3-15-fold greater depending on neuronal cell type). In contrast, the specific activity of the enzyme was about twice as high in the primary neuronal as in the astrocytic cultures after 14 days in vitro. In membrane preparations from the whole cerebellum, synaptosomal fractions, and total perikarya suspensions the inhibition of enzyme activity by ouabain indicated complex kinetics, which were consistent with the presence of two forms of the Na+,K+-ATPase (apparent Ki values of about 10(-7) M and 10(-4)-10(-5) M, respectively), the high-affinity form accounting for 60-75% of the total activity. The interaction of the enzyme with ouabain was apparently similar in perikarya preparations of granule neurones, Purkinje cells, and astrocytes. Differences were, however, observed in the properties of the Na+,K+-ATPase of cultured neurones and astrocytes. The latter contained predominantly, but not exclusively, an Na+,K+-ATPase with low affinity for ouabain (73% of the total) that is similar to the single enzyme form in the kidney. This form constituted a significantly smaller proportion of the Na+,K+-ATPase in the cultured neuronal preparations (55%). It would appear, therefore, that in membrane fractions from preparations enriched in different separated and cultured neural cell types both the high- and the low-affinity forms of the enzyme, in terms of interaction with ouabain, are expressed. Depending on the class of cells these enzyme forms constituted a different proportion of the total activity, but both forms seemed to be present in every type of cell examined, even after taking into account the contribution in the enriched preparations of the contaminating cell types. In contrast with the results on the Na+,K+-ATPase activity determined under optimal conditions in preparations derived from disrupted cells, differences could not be detected between the cultured cell types when the effect of ouabain on the uptake of 86Rb into "live cells" was estimated as a measure of in situ ion pump activity.(ABSTRACT TRUNCATED AT 400 WORDS)
研究了钠钾激活的三磷酸腺苷酶（Na⁺，K⁺ - ATP酶；EC 3.6.1.3）某些特性的活性，实验对象为富含大鼠小脑神经细胞或星形胶质细胞的培养物和核周体组分，并与来自未成熟和成年大鼠整个小脑的制剂、衍生的突触体组分以及非神经组织（如肾脏）进行比较。新鲜分离的星形胶质细胞中Na⁺，K⁺ - ATP酶的比活性明显高于神经细胞（根据神经元细胞类型，高3 - 15倍）。相反，体外培养14天后，原代神经元培养物中该酶的比活性约为星形胶质细胞培养物中的两倍。在整个小脑、突触体组分和总核周体悬浮液的膜制剂中，哇巴因对酶活性的抑制呈现复杂的动力学，这与存在两种形式的Na⁺，K⁺ - ATP酶一致（表观Ki值分别约为10⁻⁷M和10⁻⁴ - 10⁻⁵M），高亲和力形式占总活性的60 - 75%。颗粒神经元、浦肯野细胞和星形胶质细胞核周体制剂中，该酶与哇巴因的相互作用明显相似。然而，在培养的神经元和星形胶质细胞的Na⁺，K⁺ - ATP酶特性上观察到了差异。后者主要但并非唯一地含有对哇巴因亲和力低的Na⁺，K⁺ - ATP酶（占总量的73%），类似于肾脏中的单一酶形式。在培养的神经元制剂中，这种形式在Na⁺，K⁺ - ATP酶中所占比例明显较小（55%）。因此，在富含不同分离和培养神经细胞类型的制剂的膜组分中，就与哇巴因的相互作用而言，该酶的高亲和力和低亲和力形式均有表达。根据细胞类别，这些酶形式在总活性中所占比例不同，但即使考虑到污染细胞类型在富集制剂中的贡献，似乎在所检查的每种细胞类型中都存在这两种形式。与在来自破碎细胞的制剂中在最佳条件下测定的Na⁺，K⁺ - ATP酶活性结果相反，当将哇巴因对⁸⁶Rb进入“活细胞”的摄取的影响作为原位离子泵活性的指标进行评估时，在培养的细胞类型之间未检测到差异。（摘要截短于400字）
