Background: Chronic liver failure leads to hyperammonemia, a central component in the pathogenesis of hepatic encephalopathy. Neurotoxic effects of ammonia have been shown to induce numerous metabolic alterations such as increased brain lactate and glutamine levels. Newly developed imaging techniques have demonstrated brain edema is present in patients with chronic liver failure however the pathophysiological mechanisms are not clearly understood. The present study investigates the role of de novo synthesis of brain lactate and glutamine in pathogenesis of brain edema in 2 different rat models of chronic liver failure/hepatic encephalopathy; 1) portacaval anastomosis (PCA), and 2) bile-duct ligation (BDL). Methods: PCA and BDL rats along with their respective sham-operated controls were injected with [1-13C] glucose (500 mg/kg, i.p) and sacrificed 30 minutes later. Glutamine and lactate were assessed in brain tissue by 1H and 13C nuclear magnetic resonance spectroscopy. Ammonia levels were measured in arterial plasma and cerebrospinal fluid using a commercially available kit. Brain water content was measured in frontal cortex using a specific gravimetric technique. Results: Hyperammonemia developed in both PCA (177.3 ± 8.9μM vs sham: 52.1 ± 11.3μM; p< 0.0001) and BDL (155.0 ± 8.9μM vs sham: 41.0 ± 11.2μM; p< 0.0001); with no significant difference between the groups (p> 0.05). Similar results were found in cerebrospinal fluid (PCA: 179.5 ± 10.3μM vs sham: 34.9 ± 6.3μM; p< 0.0001 and BDL: 142.6 ± 28.3μM vs sham: 27.9 ± 4.9μM; p< 0.0001); with no significant difference between the groups (p> 0.05). An increase in brain water content was observed in rats with BDL (81.88 ± 0.12% vs sham: 81.13 ± 0.15%, p< 0.05) whereas no significant change in brain water content was found in PCA vs their respective sham-operated controls. Glutamine levels significantly increased 3 fold in PCA rats and 2 fold in BDL rats vs respective sham-operated controls. A significant increase in de novo synthesis of 13C lactate and 13C glutamine from 13C glucose was found in BDL rats (1.6 fold and 2 fold respectively vs sham), whereas in PCA rats only de novo synthesis of 13C glutamine increased. Conclusion: BDL and PCA rats both presented with similar levels of hyperammonemia and increased brain ammonia, increased de novo synthesis of glutamine from glucose however only BDL rats demonstrated an increase in brain water as well as an increase in de novo synthesis of lactate from glucose. Our findings suggest lactate is an important pathogenic factor implicated in the pathogenesis of brain edema in chronic liver failure.