Ammonia inhibits energy metabolism in astrocytes in a rapid and glutamate dehydrogenase 2-dependent manner

Abstract

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.

N ‐acetylaspartate release by glutaminolytic ovarian cancer cells sustains protumoral macrophages

“…This evidence is in line with our results on the metabolic rewiring following GLUL ablation in OVCAR3 cells. It is conceivable to hypothesize that ammonia accumulation in GLUL-deficient cells might lower TCA cycle dehydrogenases flux (Ott et al, 2005;Ott & Vilstrup, 2014;Drews et al, 2020), leading to citrate accumulation, as found in our model (Fig 3I). In the mitochondria, oxaloacetate might be rerouted to transamination, facilitated by high Glu levels, leading to Asp, which is promptly acetylated by NAT8L.…”

Sirtuins functions in central nervous system cells under neurological disorders

“…SIRT4 is an ADP-ribosyltransferase whereas SIRT5 is a desuccinylase and demalonylase with weak deacetylase activity ( Kida and Goligorsky, 2016 ). In hepatic encephalopathy, overexpression SIRT4 in astrocyte inhibited GDH2 activity and restored mitochondrial respiration ( Drews et al, 2020 ), which suggests SIRT4 exert a protective effect in astrocyte. In an AD model, the expression of SIRT5 was significantly decreased, along with suppressed autophagy in neuron.…”

Sirtuins-Mediated System-Level Regulation of Mammalian Tissues at the Interface between Metabolism and Cell Cycle: A Systematic Review

“…Progression of fatty liver disease into cirrhosis can result in increased levels of ammonia in the brain, which potentially inhibits mitochondrial respiration. Overexpression of SIRT4 alleviated this inhibition in astrocytes by blocking glutamate metabolism in rat and human astrocytes [ 494 ].…”