Astrocyte dysfunction is a primary factor in hepatic encephalopathy (HE) impairing neuronal activity under hyperammonemia. In particular the early events causing ammonia-induced toxicity to astrocytes are not well understood. Using established cellular HE models, we show that mitochondria rapidly undergo fragmentation in a reversible manner upon hyperammonemia. Further, within a timescale of minutes mitochondrial respiration and glycolysis were hampered which occurred in a pH-independent manner. Using metabolomics an accumulation of numerous amino acids, including branched chain amino acids and glucose was observed. Metabolomic tracking of ¹⁵N-labeled ammonia showed rapid incorporation of ¹⁵N into glutamate and glutamate-derived amino acids. Downregulating human GLUD2, encoding mitochondrial glutamate dehydrogenase 2 (GDH2), inhibiting GDH2 activity by SIRT4 overexpression, and supplementing cells with glutamate or glutamine alleviated ammonia-induced inhibition of mitochondrial respiration. Metabolomic tracking of ¹³C-glutamine showed that hyperammonemia can inhibit anaplerosis of TCA-cycle intermediates. Contrary to its classical anaplerotic role, we show that under hyperammonemia GDH2 rather catalyzes the removal of ammonia by reductive amination of α-ketoglutarate which efficiently and rapidly inhibits the TCA-cycle. Overall, we propose a critical GDH2-dependent mechanism in HE models that on the one hand helps to remove ammonia but on the other hand impairs energy metabolism in mitochondria rapidly.

中文翻译：

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氨以快速且谷氨酸脱氢酶 2 依赖性的方式抑制星形胶质细胞的能量代谢。


星形胶质细胞功能障碍是肝性脑病（HE）损害高氨血症下神经元活动的主要因素。特别是引起氨诱导的星形胶质细胞毒性的早期事件尚不清楚。使用已建立的细胞 HE 模型，我们发现线粒体在高氨血症时以可逆的方式快速发生断裂。此外，在几分钟的时间尺度内，线粒体呼吸和糖酵解受到阻碍，这以与 pH 无关的方式发生。使用代谢组学观察到多种氨基酸的积累，包括支链氨基酸和葡萄糖。 ¹⁵ N 标记氨的代谢组学追踪显示¹⁵ N 快速掺入谷氨酸和谷氨酸衍生氨基酸中。下调人GLUD2 、编码线粒体谷氨酸脱氢酶 2 (GDH2)、通过 SIRT4 过表达抑制 GDH2 活性以及向细胞补充谷氨酸或谷氨酰胺可减轻氨诱导的线粒体呼吸抑制。 ¹³ C-谷氨酰胺的代谢组学追踪表明，高氨血症可以抑制 TCA 循环中间体的回补。与其经典的回补作用相反，我们发现在高氨血症下，GDH2 通过 α-酮戊二酸的还原胺化来催化氨的去除，从而有效且快速地抑制 TCA 循环。总的来说，我们在 HE 模型中提出了一个关键的 GDH2 依赖性机制，一方面有助于去除氨，但另一方面会迅速损害线粒体的能量代谢。