Increased Na, K-ATPase α₂ isoform gene expression by ammonia in astrocytes and in brain in vivo

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• 作者：Zhanxia Xue ; Baoman Li ; Li Gu ; Xiaoling Hu ; Min Li ; Roger F. Butterworth ; Liang Peng
• 关键词：Astrocytes ; Na ; K-ATPase α ; ₂ isoform ; Ammonia ; EGF receptor ; Ouabain-like compounds ; Gene expression
• 刊名：Neurochemistry International
• 出版年：2010
• 出版时间：November 2010
• 年：2010
• 卷：57
• 期：4
• 页码：395-403
• 全文大小：1174 K

文摘

In mouse astrocyte cultures identical to those used in the present study ammonia increases the production of ouabain-like compounds and Na, K-ATPase activity (Kala et al., 2000). Increased activity of Na, K-ATPase could be the result of enhanced production of ouabain-like compounds, since cultured rat astrocytes react to prolonged exposure to a high concentration of ouabain with an upregulation of the Na, K-ATPase α₁ isoform (Hosoi et al., 1997). However, unlike astrocytes in brain in vivo and mouse primary cultures, cultured rat astrocytes do not express the astrocyte-specific α₂ isoform, which shows a higher affinity for ouabain (EC₅₀ 0.1 μM) than the α₁ isoform (EC₅₀  10 μM). In the present study we have investigated (i) effects of ammonia on mRNA and protein expression of α₁ and α₂ isoforms in primary cultures of mouse astrocytes; (ii) effects of hyperammonia obtained by urease injection on mRNA and protein expression of α₁ and α₂ isoforms in the brain in vivo; and (iii) effect on observed upregulation of gene expression of AG1478, an inhibitor of the EGF receptor-tyrosine kinase, PP1, an inhibitor of Src, and GM6001, an inhibitor of Zn²⁺-dependent metalloproteinases in the cultured cells. It was established that α₂ mRNA and protein expression, but not α₁ expression, was upregulated in cultured astrocytes by 1–4 days of exposure to 3 or 5 mM ammonia and that similar upregulation, contrasted by a downregulation of the neuronal α₃ subunit occurred in the hyperammonemic brain. Based on the effects of the inhibitors and literature data it is concluded that ammonia activates formation of an endogenous ouabain-like compound, which binds to the Na, K-ATPase, activating Src, which in turn stimulates the receptor-tyrosine kinase of the EGF receptor, leading to activation of the Ras, Raf, MEK pathway and phosphorylation of ERK_1/2, which eventually causes upregulation of α₂ gene expression.