Analysis of mechanisms regulation ion transport activities of Na pump isoforms in neurons.

神经元Na泵亚型离子转运活性调节机制分析。

• 批准号: 13680847
• 负责人: INOUE Nobuo
• 金额: $ 2.3万
• 依托单位: Yokohama City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680847/
• 关键词: Neuron; Na pump; Na, K-ATPase; Isoform; Ion transport; ES cell; Differentiation; Na, K-ATPase; 細胞外カリウム; リン酸化

Three Na pump isoforms α1, α2 and α3 isoforms, are present in cultured neurons and ion transport activities of the isoforms are differentially regulated. Here we show that the inhibition of α2/α3, a neuron type Na pump, isoform by extracellular K^+ is a key mechanism of the differential regulation. Only the α2/α3 isoform is inhibited by physiological concentrations of K^+ under basal conditions in mature neurons. The inhibition was vanished after glutamate excitation of the neurons and by incubation with a calmodulin antagonist or an inhibitor of CaM kinase II. In contrast to mature neurons the inhibition is small in immature neurons, in which the isoform activities are not regulated differentially. Additionally, when ATP concentration in the neurons becomes low, the inhibition diminishes and the differential regulation becomes increasingly prominent. We also show that astrocyte-derived factors instruct mouse ES cells to differentiate into neurons. Cultured in astrocyte-conditioned med … More ium (ACM) under free-floating conditions, within 4 days, colonies of undifferentiated ES cells give rise to floating spheres with a concentric stratiform structure. Specifically, the spheres have a periphery of neural stem cells, a core of dividing ES cells and an intermediate layer. Culturing the spheres on an adhesive substrate in ACM promotes neurogenesis. The attached spheres give rise to clusters of cells containing neurons at the periphery, and many neurons with neurites migrate from the clusters within 5 days. Gene expression analyses and electrophysiology confirm the neuronal properties of the cells. The neurons express a neuron type Na pump, the α2 and α3 isoforms. Immunofluorescence analyses show that many NF^+ neurons and neurites are also positive for tyrosine hydroxylase. In contrast, neither astrocytes nor oligodendrocytes are formed. This pathway provides a new insight into mechanisms of neurogenesis and also provides a simple procedure for generation of neural stem cells and neurons. Less

三种 Na 泵亚型 α1、α2 和 α3 亚型存在于培养的神经元中，并且亚型的离子转运活性受到不同的调节。在这里，我们表明细胞外 K^+ 对 α2/α3（一种神经元型 Na 泵同工型）的抑制是差异调节的关键机制。在成熟神经元的基础条件下，只有 α2/α3 亚型会被生理浓度的 K^+ 抑制。谷氨酸刺激神经元后，并与钙调蛋白拮抗剂或 CaM 激酶 II 抑制剂一起孵育，抑制作用消失。与成熟神经元相比，未成熟神经元的抑制较小，其中亚型活性没有受到差异性调节。此外，当神经元中的 ATP 浓度变低时，抑制作用减弱，差异调节变得越来越突出。我们还表明星形胶质细胞衍生因子指导小鼠 ES 细胞分化为神经元。在星形胶质细胞条件培养基 (ACM) 中自由漂浮条件下培养，4 天内，未分化的 ES 细胞集落产生具有同心层状结构的漂浮球体。具体来说，球体具有神经干细胞的外围、分裂的 ES 细胞的核心和中间层。在 ACM 中的粘合基质上培养球体可促进神经发生。附着的球体在周围产生含有神经元的细胞簇，并且许多带有神经突的神经元在 5 天内从细胞簇中迁移出来。基因表达分析和电生理学证实了细胞的神经元特性。神经元表达神经元型 Na 泵、α2 和 α3 亚型。免疫荧光分析表明许多 NF^+ 神经元和神经突也呈酪氨酸羟化酶阳性。相反，星形胶质细胞和少突胶质细胞均不形成。该途径提供了对神经发生机制的新见解，并且还提供了生成神经干细胞和神经元的简单程序。较少的

项目成果

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Sasaki Y, Cheng C, Uchida Y, Nakajima O, Ohshima T, Yagi T, Taniguchi M, Nakayama T, Kishida R, Kudo Y, Ohno S, Nakamura F and Goshima Y: "Fyn and cdk5 mediate Semaphorin-3A signaling, which is involved in regulation of dendrite orientation in cerebral co

Sasaki Y、Cheng C、Uchida Y、Nakajima O、Ohshima T、Yagi T、Taniguchi M、Nakayama T、Kishida R、Kudo Y、Ohno S、Nakamura F 和 Goshima Y：“Fyn 和 cdk5 介导 Semaphorin-3A 信号传导，其中