Aberrant Gene Expression in Dogs with Portosystemic Shunts

Congenital portosystemic shunts are developmental anomalies of the splanchnic vascular system that cause portal blood to bypass the liver. Large-breed dogs are predisposed for intrahepatic portosystemic shunts (IHPSS) and small-breed dogs for extrahepatic portosystemic shunts (EHPSS). While the phenotype resulting from portal bypass of the liver of the two types of shunt is identical, the genotype and molecular pathways involved are probably different. The aim of this study was to gain insight into the pathways involved in the different types of portosystemic shunting. Microarray analysis of mRNA expression in liver tissue from dogs with EHPSS and IHPSS revealed that the expression of 26 genes was altered in either IHPSS or EHPSS samples compared with that in liver samples from control dogs. Quantitative real-time PCR of these genes in 14 IHPSS, 17 EHPSS, and 8 control liver samples revealed a significant differential expression of ACBP, CCBL1, GPC3, HAMP, PALLD, VCAM1, and WEE1. Immunohistochemistry and Western blotting confirmed an increased expression of VCAM1 in IHPSS but its absence in EHPSS, an increased WEE1 expression in IHPSS but not in EHPSS, and a decreased expression of CCBL1 in both shunt types. Regarding their physiologic functions, these findings may indicate a causative role for VCAM1 in IHPSS and WEE1 for IHPSS. CCBL1 could be an interesting candidate to study not yet elucidated aspects in the pathophysiology of hepatic encephalopathy.     

Hyperammonemia and Systemic Inflammatory Response Syndrome Predicts Presence of Hepatic Encephalopathy in Dogs with Congenital Portosystemic Shunts

Hepatic encephalopathy (HE) is an important cause of morbidity and mortality in patients with liver disease. The pathogenesis of he is incompletely understood although ammonia and inflammatory cytokines have been implicated as key mediators. To facilitate further mechanistic understanding of the pathogenesis of HE, a large number of animal models have been developed which often involve the surgical creation of an anastomosis between the hepatic portal vein and the caudal vena cava. One of the most common congenital abnormalities in dogs is a congenital portosystemic shunt (cpss), which closely mimics these surgical experimental models of HE. Dogs with a cPSS often have clinical signs which mimic clinical signs observed in humans with HE. Our hypothesis is that the pathogenesis of HE in dogs with a cPSS is similar to humans with HE. The aim of the study was to measure a range of clinical, haematological and biochemical parameters, which have been linked to the development of HE in humans, in dogs with a cPSS and a known HE grade. One hundred and twenty dogs with a cPSS were included in the study and multiple regression analysis of clinical, haematological and biochemical variables revealed that plasma ammonia concentrations and systemic inflammatory response syndrome scores predicted the presence of HE. Our findings further support the notion that the pathogenesis of canine and human HE share many similarities and indicate that dogs with cPSS may be an informative spontaneous model of human HE. Further investigations on dogs with cPSS may allow studies on HE to be undertaken without creating surgical models of HE thereby allowing the number of large animals used in animal experimentation to be reduced.     

Distribution of extrahepatic congenital portosystemic shunt morphology in predisposed dog breeds

ABSTRACT: BACKGROUND: An inherited basis for congenital extrahepatic portosystemic shunts (EHPSS) has been demonstrated in several small dog breeds. If in general both portocaval and porto-azygous shunts occur in breeds predisposed to portosystemic shunts then this could indicate a common genetic background. This study was performed to determine the distribution of extrahepatic portocaval and porto-azygous shunts in purebred dog populations. RESULTS: Data of 135 client owned dogs diagnosed with EHPSS at the Faculty of Veterinary Medicine of Utrecht University from 2001 - 2010 were retrospectively analyzed. The correlation between shunt localization, sex, age, dog size and breed were studied. The study group consisted of 54 males and 81 females from 24 breeds. Twenty-five percent of dogs had porto-azygous shunts and 75 % had portocaval shunts. Of the dogs with porto-azygous shunts only 27 % was male (P = 0.006). No significant sex difference was detected in dogs with a portocaval shunt. Both phenotypes were present in almost all breeds represented with more than six cases. Small dogs are mostly diagnosed with portocaval shunts (79 %) whereas both types are detected. The age at diagnosis in dogs with porto-azygous shunts was significantly higher than that of dogs with portocaval shunts (P < 0.001). CONCLUSION: The remarkable similarity of phenotypic variation in many dog breeds may indicate common underlying genes responsible for EHPSS across breeds. The subtype of EHPSS could be determined by a minor genetic component or modulating factors during embryonic development.     

Inherited liver shunts in dogs elucidate pathways regulating embryonic development and clinical disorders of the portal vein

Congenital disorders of the hepatic portal vasculature are rare in man but occur frequently in certain dog breeds. In dogs, there are two main subtypes: intrahepatic portosystemic shunts, which are considered to stem from defective closure of the embryonic ductus venosus, and extrahepatic shunts, which connect the splanchnic vascular system with the vena cava or vena azygos. Both subtypes result in nearly complete bypass of the liver by the portal blood flow. In both subtypes the development of the smaller branches of the portal vein tree in the liver is impaired and terminal branches delivering portal blood to the liver lobules are often lacking. The clinical signs are due to poor liver growth, development, and function. Patency of the ductus venosus seems to be a digenic trait in Irish wolfhounds, whereas Cairn terriers with extrahepatic portosystemic shunts display a more complex inheritance. The genes involved in these disorders cannot be identified with the sporadic human cases, but in dogs, the genome-wide study of the extrahepatic form is at an advanced stage. The canine disease may lead to the identification of novel genes and pathways cooperating in growth and development of the hepatic portal vein tree. The same pathways likely regulate the development of the vascular system of regenerating livers during liver diseases such as hepatitis and cirrhosis. Therefore, the identification of these molecular pathways may provide a basis for future proregenerative intervention.     

Alterations in Cortical [3H]Kainate and α-[3H]Amino-3-Hydroxy-5-Methyl-4-Isoxazolepropionic Acid Binding in a Spontaneous Canine Model of Chronic Hepatic Encephalopathy

Excitatory amino acid receptor binding parameters were investigated in a spontaneous dog model of chronic hepatic encephalopathy. L-[3H]Glutamate, (+)-[3H]-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-im ine maleate ([3H]MK-801), [3H]kainate, and alpha-[3H]-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid ([3H]AMPA) binding experiments were performed using crude cerebrocortical synaptosomal membrane preparations from dogs with congenital portosystemic encephalopathy (PSE) and control dogs. There was no change in the affinity or density of L-[3H]-glutamate or [3H]MK-801 binding sites in dogs with congenital PSE compared with control dogs. However, in the PSE dogs there was a significant reduction in the density of [3H]kainate binding sites compared with control dogs and abolition of the low-affinity [3H]AMPA binding site. The relative binding capacity of PSE synaptosomal membranes for [3H]kainate and [3H]AMPA was expressed as the ratio Bmax/KD. There was a significant inverse correlation between the Bmax/KD ratio for [3H]AMPA binding and the worst grade of encephalopathy experienced by each dog. These results suggest that there is a significant perturbation of cerebrocortical non-N-methyl-D-aspartate receptor binding in dogs with congenital PSE which may have relevance to the pathogenesis of hepatic encephalopathy.     

Cerebellar [H]kainate and [H]ampa binding in dogs with congenital portosystemic encephalopathy

Previous studies have indicated that kainate and AMPA receptors are altered in cerebral cortex of dogs with chronic hepatic encephalopathy (HE). To ascertain whether receptors in dog cerebellum are similarly altered in HE [³H]kainate and [³H]-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) binding assays were performed on crude synaptosomal membranes prepared from cerebellar tissue from dogs with congenital portosystemic encephalopathy (PSE) and control dogs. There was no pathophysiologically relevant difference in the affinity or density of kainate or AMPA binding sites in PSE cerebellar tissue compared with control dogs. The failure to demonstrate alterations in these binding parameters in cerebellar tissue was expected as clinical signs of HE reflect cortical rather than cerebellar dysfunction.     

Low Vitamin D Status Is Associated with Systemic and Gastrointestinal Inflammation in Dogs with a Chronic Enteropathy

Introduction

Vitamin D deficiency, as assessed by serum concentrations of 25 hydroxyvitamin D (25(OH)D), has been linked to the development of over-zealous and inappropriate inflammation in humans. However, the relationship between vitamin D status and inflammation in dogs is ill-defined. Chronic enteropathies (CE) are frequently diagnosed in client owned dogs, have a wide range of serum 25(OH)D concentrations, and represent a spontaneous model in which to probe the relationship between vitamin D and inflammation. The hypothesis of this study was that vitamin D status would be negatively associated with systemic and gastrointestinal inflammation in dogs with a CE. The aim of this study was to examine the relationship between serum 25(OH)D concentrations and markers of systemic and gastrointestinal inflammation in a cohort of dogs with CE.

Methods and Materials

Serum 25(OH)D concentrations, together with neutrophil, monocyte, eosinophil and lymphocyte counts, duodenal histopathology scores, serum IL-2, IL-6, IL-8 and TNFα concentrations and were measured in 39 dogs with histologically confirmed CE. A linear regression model examined the relationship between serum 25(OH)D status and measures of inflammation.

Results

Serum 25(OH)D concentrations were negatively associated with neutrophil and monocyte counts, duodenal histopathology scores and serum IL-2 and IL-8 concentrations. Dogs with low serum 25(OH)D concentrations typically had an inflammatory signature characterised by high monocyte and neutrophil numbers together with low lymphocyte numbers. There is a need to establish whether low vitamin D status is a cause or consequence of inflammation.     

Endogenous Benzodiazepine Receptor Ligands in Human and Animal Hepatic Encephalopathy

The role of endogenous benzodiazepine receptor ligands in the pathogenesis of hepatic encephalopathy was studied in humans and in rat models of hepatic encephalopathy. Endogenous benzodiazepine ligands were extracted from rat brain and human CSF by acid treatment and purification by HPLC. Detection and partial characterization of these endogenous benzodiazepine ligands were carried out using both radioreceptor binding assays and radioimmunoassays with anti-benzodiazepine antibodies. Four different benzodiazepine receptor ligands were identified in human and rat tissue, two of which may be diazepam and desmethyldiazepam, based on elution profiles and anti-benzo-diazepine antibody reactivity. Human CSF and serum from patients with hepatic encephalopathy contained approximately 10 times more endogenous benzodiazepine receptor ligand than CSF from controls or nonencephalopathic patients with liver disease. The levels of brain benzodiazepine receptor ligand compounds were also increased approximately 10-fold in rats suffering from fulminant hepatic failure, but not in rats with portacaval shunts, a model of chronic hepatic disease. The increased concentrations of these substances could be behaviorally significant and may contribute to the pathogenesis of hepatic encephalopathy.     

Arterial blood ketone body ratio as an indicator of the no-return point in hepatic inflow occlusion without venous shunt in dogs

The effects of hepatic inflow occlusion without venous shunt on the viability of the liver were investigated with respect to liver energy metabolism in dogs, subjected to portal triad cross-clamping (Pringle's maneuver) for 10, 30 and 60 min. The concentrations of ketone bodies and the arterial blood ketone body ratio (KBR) were decreased markedly by hepatic inflow occlusion, but recovered upon recirculation. The initial velocity of KBR recovery was 0.150/min after 10-min clamping, 0.140/min after 30-min clamping and 0.032/min after 60-min clamping. KBR recovery was delayed when hepatic inflow occlusion exceeded 30 min, which indicates that hepatic inflow occlusion for 60 min causes severe inhibition of energy generation in liver mitochondria. These findings indicate that the safety period for hepatic inflow occlusion without venous shunt is between 30 and 60 min in dogs, and that mortality can be predicted by measuring the initial velocity of KBR recovery upon recirculation.     

A model for experimental bacterial cystitis in the dog

To induce an experimental model of bacterial cystitis, ten preconditioned dogs underwent bladder irritation with a 0.1% alcoholic solution of salicylic acid followed in 24 hours by an intravesicular infusion of Proteus mirabilis. The dogs were observed for the following 14 days (five dogs) and 17 days (five dogs) and then euthanatized and necropsied. Tenesmus, dysuria, hematuria, and pollakiuria occurred in all dogs, but the severity of these signs diminished with time. The total white cell, neutrophil, and monocyte counts in the peripheral blood increased and urinalysis results were consistent with infection and severe inflammation. The infection persisted for the duration of the study, although the average quantitative bacterial count in urine progressively declined. No changes occurred in the measured clinical chemistry values. Severe inflammation was present on gross examination of the bladder and microscopic examination of the bladder, prostate, and renal pelvis. Less severe inflammation was present on microscopic examination of the urethra and ureter.     

Elective End-to-side Portacaval Shunt: Results in 64 Cases

In a series of 64 cases of elective end-to-side portacaval shunts performed for liver disease the success rate—in that the patient survived with a patent shunt, free of subsequent haemorrhage and severe encephalopathy—was 48%.The early postoperative death rate was 12·5% and the five-year survival 65%. Bleeding from oesophagogastric varices after blockage of the shunt was responsible for at least half of the early postoperative deaths, and most late deaths were due to liver failure. A decreased chance of late survival was associated with age over 40 years, active chronic hepatitis, and with a preoperative history of hepatocellular jaundice.Shunt blockage occurred in 16% of patients, and all bled again from oesophagogastric varices. Shunt block is more likely if the portal vein is calcified or thrombosed, and may be more likely if the portal vein diameter, as shown by splenic venography, is 1·5 cm or less.In survivors with a patent shunt the most serious late complication was chronic, severe portal-systemic encephalopathy, which occurred in 38%. Severe encephalopathy was associated with age over 40 years, a preoperative history of any degree of encephalopathy, diabetes mellitus, and with continued drinking in the alcoholic. Most patients who had portal-systemic encephalopathy in the first year postoperatively developed chronic disabling encephalopathy.A preoperative history of transient mild or moderate ascites did not seem adversely to influence the outcome.     

ANIT toxicity toward mouse hepatocytes in vivo is mediated primarily by neutrophils via CD18

Alpha-naphthylisothiocyanate (ANIT) is a hepatotoxicant that causes acute cholestatic hepatitis with infiltration of neutrophils around bile ducts and necrotic hepatocytes. The objective of this study was to determine whether the beta2-integrin CD18, which plays an important role in leukocyte invasion and cytotoxicity, contributes to ANIT-induced hepatic inflammation and liver injury. Mice with varying levels of leukocyte CD18 expression were treated with ANIT and monitored for hepatic neutrophil influx and liver injury over 48 h. Mice that were partially deficient in CD18 (30% of normal levels) developed periportal inflammation and widespread hepatic necrosis after ANIT treatment in a pattern identical to that in wild-type (WT) mice. In contrast, mice that completely lack CD18 (CD18 null) were resistant to ANIT toxicity. Forty-eight hours after ANIT, CD18-null mice displayed 60% lower serum alanine aminotransferase (ALT) levels and 75% less hepatic necrosis, as shown by morphometry, than WT mice. This was true despite evidence that ANIT still provoked hepatic neutrophil influx in CD18-null mice. WT mice could also be protected from ANIT-induced hepatocellular necrosis, by depleting the animals of neutrophils. Notably, neither CD18-null mice nor neutrophil-depleted WT mice exhibited any attenuation of bile duct injury or cholestasis due to ANIT. We conclude from these experiments that neutrophils invade ANIT-treated livers in a CD18-independent fashion but utilize CD18 to induce hepatocellular cytotoxicity. The results emphasize that neutrophil-mediated amplification of ANIT-induced liver injury is directed toward hepatocytes rather than cholangiocytes. In fact, the data indicate that the majority of ANIT toxicity toward hepatocytes in vivo is neutrophil driven.     

The effect of pancreatic and intestinal venous blood on hepatic atrophy and compensatory hyperplasia in the rat

Hepatotrophic effect of pancreatic and intestinal venous blood was studied in rats with mesocaval or distal splenocaval shunt following ligation of a branch of the portal vein supplying 70% of liver mass. Because 2/3 of liver mass was deprived of portal flow the nonligated liver lobes were not hypoperfused due to shunt procedure. During the first three postoperative days the DNA synthesis, mitotic index, and changes in relative weights were measured in both ligated (atrophied) and nonligated (compensatory hyperplasia) parts of the liver. It was found, that the restorative capacity of the liver existed in rats with selective portasystemic shunts. The stimulus to growth was greater in lobes supplied by intestinal venous blood compared to those perfused by pancreatic effluent. The increase in DNA synthesis occurred in lobes undergoing atrophy and the intensity of this response was also dependent on type of shunt since recirculation of intestinal blood by way of the hepatic artery inhibited atrophy to a greater extent than pancreatic venous effluent. Although the patency of arterial branches was confirmed the ligated lobes showed necrotic lesions. Systemic recirculation of intestinal venous blood far more inhibited necrosis than pancreatic venous blood.     

The Relationship Between Plasma and Brain Quinolinic Acid Levels and the Severity of Hepatic Encephalopathy in Animal Models of Fulminant Hepatic Failure

Abstract: Quinolinic acid is an excitatory, neurotoxic tryptophan metabolite proposed to play a role in the pathogenesis of hepatic encephalopathy. This involvement was investigated in rat and rabbit models of fulminant hepatic failure at different stages of hepatic encephalopathy. Although plasma and brain tryptophan levels were significantly increased in all stages of hepatic encephalopathy, quinolinic acid levels increased three- to sevenfold only in the plasma, CSF, and brain regions of animals in stage IV hepatic encephalopathy. Plasma-CSF and plasma-brain quinolinic acid levels in rats and rabbits with fulminant hepatic failure were strongly correlated, with CSF and brain concentrations ∼10% those of plasma levels. Moreover, there was no significant regional difference in brain quinolinic acid concentrations in either model. Extrahepatic indoleamine-2,3-dioxygenase activity was not altered in rats in stage IV hepatic encephalopathy, but hepatic l -tryptophan-2,3-dioxygenase activity was increased. These results suggest that quinolinic acid synthesized in the liver enters the plasma and then accumulates in the CNS after crossing a permeabilized blood-brain barrier in the end stages of liver failure. Furthermore, the observation of low brain concentrations of quinolinic acid only in stage IV encephalopathy suggests that the contribution of quinolinic acid to the pathogenesis of hepatic encephalopathy in these animal models is minor.     

Reduced somatostatin-like immunoreactivity in the brain of dogs with an Eck fistula

Somatostatin-like immunoreactivity (SLI) in the brains of Eck fistula dogs, prepared as an experimental model of hepatic encephalopathy, was measured to investigate the pathogenesis of hepatic encephalopathy. The values were studied in comparison with the concentrations of amino acids where the imbalance was suggested to cause hepatic encephalopathy. SLI levels in the parietal and temporal cortex of Eck fistula dogs were 76.0 +/- 12.0 (mean +/- S.E.M., fmol/mg wet wt.) and 113.4 +/- 23.7, and those of controls were 144.0 +/- 11.8 and 186.9 +/- 19.2, respectively, the differences being statistically significant (P less than 0.005, P less than 0.05). No significant difference in gel filtration profiles of SLI in extracts from parietal and temporal cortex was observed between Eck fistula dogs and controls. Tyrosine and phenylalanine, which are suggested to be precursors of false neurotransmitters, were significantly increased in the parietal cortex of the Eck fistula dogs, and phenylalanine was significantly increased in the temporal cortex of these dogs. There was a significant negative correlation between SLI and phenylalanine concentrations in the parietal and temporal cortex (r = -0.7171, P less than 0.01). These results suggest that the reduced SLI may be one of the factors which cause the neuropsychiatric disturbances in hepatic encephalopathy.     

Metabolomics as a diagnostic tool for hepatology: validation in a naturally occurring canine model

Human hepatopathies are a diagnostic challenge, with many distinct diseases having similar clinical signs and laboratory findings. Naturally occurring canine hepatic disease provides an excellent model for human diseases and similar diagnostic dilemmas exist; differentiating canine congenital portosystemic vascular anomalies (PVA) from acquired hepatopathies is difficult and traditionally requires invasive diagnostic procedures. The emerging post-genomic science of metabolomics is concerned with detecting global changes of populations of endogenous low molecular weight metabolites in biological samples and offers the possibility of identifying surrogate profiles of disease. Metabolomics couples sensitive metabolite analysis with sophisticated pattern recognition techniques. In this study, a metabolomic strategy has been employed to assess metabolite changes in the plasma of dogs with congenital PVA and acquired hepatic disease. Plasma samples were collected from 25 dogs, comprising 9 dogs with congenital PVA, 6 with acquired hepatopathy and 10 with non-hepatic disorders. Low molecular weight metabolites were analyzed by liquid chromatography-mass spectrometry (LC-MS). Following identification of metabolites, multivariate data analysis was used to compare profiles amongst groups. The analysis demonstrated significant disturbances in the plasma bile acid and phospholipid profiles of dogs with portovascular anomalies. In contrast to traditional laboratory parameters, the metabolomic strategy was able to produce a clear segregation between all three study groups. In conclusion, this study demonstrates the potential of metabolomics as a diagnostic tool for naturally occurring hepatic disease. With further validation, this approach will improve diagnostic capabilities, provide an insight into pathogenetic mechanisms, and ultimately inform therapeutic decision making in clinical hepatology. This work was supported in part by grants from The Royal Society, Petplan Charitable Trust and The Waltham Foundation.     

Evidence for an Astrocytic Glutamate Transporter Deficit in Hepatic Encephalopathy

There is increasing evidence to suggest that hepatic encephalopathy in acute liver failure is the result of altered glutamatergic function. In particular, the high affinity uptake of glutamate is decreased in brain slices and synaptosomes from rats with acute liver failure as well as by exposure of cultured astrocytes to concentrations of ammonia equivalent to those reported in brain in acute liver failure. Both protein and gene expression of the recently cloned and sequenced astrocytic glutamate transporter GLT-1 are significantly reduced in the brains of rats with acute liver failure. Decreased expression of GLT-1 in brain in acute liver failure results in increased extracellular brain glutamate concentrations which correlates with arterial ammonia concentrations and with the appearance of severe encephalopathy and brain edema in these animals. Ammonia-induced reductions in expression of GLT-1 resulting in increased extracellular glutamate concentrations could explain some of the symptoms (hyperexcitability, cerebral edema) characteristic of hepatic encephalopathy in acute liver failure.     

TIPS and splenorenal shunt for complications of portal hypertension in chronic hepatosplenic schistosomiasis–A case series and review of the literature

BackgroundTransjugular intrahepatic portosystemic shunt (TIPS) and shunt surgery are established treatment options for portal hypertension, but have not been systematically evaluated in patients with portal hypertension due to hepatosplenic schistosomiasis (HSS), one of the neglected tropical diseases with major impact on morbidity and mortality in endemic areas.MethodsIn this retrospective case study, patients with chronic portal hypertension due to schistosomiasis treated with those therapeutic approaches in four tertiary referral hospitals in Germany and Italy between 2012 and 2020 were included. We have summarized pre-interventional clinical data, indication, technical aspects of the interventions and clinical outcome.FindingsOverall, 13 patients with confirmed HSS were included. 11 patients received TIPS for primary or secondary prophylaxis of variceal bleeding due to advanced portal hypertension and failure of conservative management. In two patients with contraindications for TIPS or technically unsuccessful TIPS procedure, proximal splenorenal shunt surgery in combination with splenectomy was conducted. During follow-up (mean follow-up 23 months, cumulative follow-up time 31 patient years) no bleeding events were documented. In five patients, moderate and transient episodes of overt hepatic encephalopathy were observed. In one patient each, liver failure, portal vein thrombosis and catheter associated sepsis occurred after TIPS insertion. All complications were well manageable and had favorable outcomes.ConclusionsTIPS implantation and shunt surgery are safe and effective treatment options for patients with advanced HSS and sequelae of portal hypertension in experienced centers, but require careful patient selection.     

Amino Acid Release from Cerebral Cortex in Experimental Acute Liver Failure, Studied by In Vivo Cerebral Cortex Microdialysis

Both increased gamma-aminobutyric acid (GABA)-ergic and decreased glutamatergic neurotransmission have been suggested relative to the pathophysiology of hepatic encephalopathy. This proposed disturbance in neurotransmitter balance, however, is based mainly on brain tissue analysis. Because the approach of whole tissue analysis is of limited value with regard to in vivo neurotransmission, we have studied the extracellular concentrations in the cerebral cortex of several neuroactive amino acids by application of the in vivo microdialysis technique. During acute hepatic encephalopathy induced in rats by complete liver ischemia, increased extracellular concentrations of the neuroactive amino acids glutamate, taurine, and glycine were observed, whereas extracellular concentrations of aspartate and GABA were unaltered and glutamine decreased. It is therefore suggested that hepatic encephalopathy is associated with glycine potentiated glutamate neurotoxicity rather than with a shortage of the neurotransmitter glutamate. In addition, increased extracellular concentration of taurine might contribute to the disturbed neurotransmitter balance. The observation of decreasing glutamine concentrations, after an initial increase, points to a possible astrocytic dysfunction involved in the pathophysiology of hepatic encephalopathy.