Caecal ligation and puncture induced sepsis in the rat results in increased brain water content and perimicrovessel oedema.

To investigate brain water content and ultrastructure in a rat caecal ligation and puncture (CLP) model of sepsis, adult male Wistar rats were assigned to one of the following experimental groups: CLP, Un-operated or Sham. CLP was performed under anaesthesia, Sham rats were exposed to anaesthesia, laparotomy and caecal mobilisation and Un-operated rats did not experience anaesthesia or surgery. CLP and Sham rats were sacrificed 18-20 h following recovery from surgery and Un-operated rats were sacrificed at the same time. Frontal cortex samples (CLP n = 9; Un-operated n = 10; Sham n = 8) were taken immediately post mortem and their water content determined using gravimetry. Similar samples were taken from other rats (CLP n = 8; Un-operated n = 8; Sham n = 8), processed for electron microscopy and subjected to morphometric analysis. There was significantly more brain water in CLP than Un-operated (P < 0.01) and Sham (P < 0.05) rats. Electron microscopy revealed significantly more peri-microvessel oedema in CLP than Un-operated (P < 0.001) and Sham rats (P < 0.05). Microvessel endothelial cell lumen cross-sectional area was significantly smaller in CLP than Un-operated (P < 0.001) and Sham (P < 0.05) rats and microvessel endothelial cell cross-sectional area was significantly smaller in CLP than Un-operated (P < 0.05) rats. Significantly more endothelial cell cytoplasmic area was occupied by mitochondria in CLP than Un-operated (P < 0.05) and Sham (P < 0.05) rats. However, experimental group did not affect the number of mitochondria present in endothelial cell profiles, or their cross-sectional area. Therefore, sepsis-induced cerebral oedema involves an increase in and a redistribution of brain water, together with ultrastructural changes to cerebral microvessels and adjacent tissue.

Role of aquaporin-4 in the development of brain oedema in liver failure.

BACKGROUND & AIMS: Liver failure is associated with progressive cytotoxic brain oedema (astrocyte swelling), which underlies hepatic encephalopathy (HE). Ammonia and superimposed inflammation are key synergistic factors in HE, but the mechanism(s) involved remain unknown. We aimed to determine whether aquaporin-4 (AQP4), an astrocyte endfeet bi-directional water channel, is associated with the brain oedema of HE. METHOD: Rats (n=60) received sham-operation (sham), 5 days hyperammonaemia-inducing diet (HD), galactosamine (GALN) induced acute liver failure (ALF), 4 weeks bile duct-ligation (BDL) induced cirrhosis, or caecal ligation and puncture (CLP), a 24h model of bacterial peritonitis. Rats from every group (except CLP) were randomised to receive intraperitoneal injections of lipopolysaccharide (LPS; 1mg/kg) or saline, prior to termination 3h later. Brain water, AQP4 protein expression (western blot) and AQP4 localisation by immunogold electron microscopy were investigated. RESULTS: Significant hyperammonaemia was observed in saline-injected BDL (p<0.05), GALN (p<0.01), and HD (p<0.01), compared to sham rats. LPS injection did not affect arterial ammonia or plasma biochemistry in any of the treatment groups. Increased brain water was observed in saline-injected GALN (p<0.05), HD (p<0.01), and CLP (p<0.001) compared to sham rats. Brain water was numerically increased in BDL rats, but this failed to reach significance (p=0.09). LPS treatment further increased oedema significantly in all treatment groups (p<0.05, respectively). AQP4 expression was significantly increased in saline-injected BDL (p<0.05), but not other treatment groups, compared to sham rats. Membrane polarisation was maintained in BDL rats. CONCLUSION: The results suggest that AQP4 is not directly associated with the development of brain oedema in liver failure, hyperammonaemia, or sepsis. In cirrhosis, there is increased AQP4 protein expression, but membrane polarisation, is maintained, possibly in a compensatory attempt to limit severe brain oedema.

Endotoxemia produces coma and brain swelling in bile duct ligated rats.

UNLABELLED: This study explores the hypothesis that the inflammatory response induced by administration of lipopolysaccharide (LPS) exacerbates brain edema in cirrhotic rats; and if so whether this is associated with altered brain metabolism of ammonia or anatomical disturbance of the blood-brain barrier. Adult Sprague-Dawley rats 4 weeks after bile duct ligation (BDL)/Sham-operation, or naïve rats fed a hyperammonemic diet (HD), were injected with LPS (0.5 mg/kg, intraperitoneally) or saline, and killed 3 hours later. LPS administration increased brain water in HD, BDL, and sham-operated groups significantly (P < 0.05), but this was associated with progression to pre-coma stages only in BDL rats. LPS induced cytotoxic brain swelling and maintained anatomical integrity of the blood-brain barrier. Plasma/brain ammonia levels were higher in HD and BDL rats than in sham-operated controls and did not change with LPS administration. Brain glutamine/myoinositol ratio was increased in the HD group but reduced in the BDL animals. There was a background pro-inflammatory cytokine response in the brains of cirrhotic rats, and plasma/brain tumor necrosis factor alpha (TNF-alpha) and IL-6 significantly increased in LPS-treated animals. Plasma nitrite/nitrate levels increased significantly in LPS groups compared with non-LPS controls; however, frontal cortex nitrotyrosine levels only increased in the BDL + LPS rats (P < 0.005 versus BDL controls). CONCLUSION: Injection of LPS into cirrhotic rats induces pre-coma and exacerbates cytotoxic edema because of the synergistic effect of hyperammonemia and the induced inflammatory response. Although the exact mechanism of how hyperammonemia and LPS facilitate cytotoxic edema and pre-coma in cirrhosis is not clear, our data support an important role for the nitrosation of brain proteins.