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Scope: This study addresses the effects of branched-chain amino acids (BCAA) on global gene expression in liver and skeletal muscle and the molecular mechanisms underlying the improvement in liver cirrhosis using DNA microarray analysis combined with RNase protection assay., Methods and Results: Male Wistar rats administered carbon tetrachloride (CCl(4) ) repeatedly for 19 weeks as a decompensated cirrhosis model were thereafter given BCAA-enriched diet (AL) or normal diet (LC) for 5 weeks. The control-diet rats without CCl(4) administration were used as a normal control group. Gene expression in AL was reversed by twofold greater than in LC in the microarray were selected to elucidate the improvements in nutritional and metabolic disorders. Downregulation of fatty acid translocase (FAT)/Cd36, glutamine synthetase, and pyruvate dehydrogenase kinase isoenzyme 4 is believed to promote lower uptake of fatty acids, lower ammonia incorporation, and higher uptake of glucose, and thus to provide an energy source without using BCAA. Ultimately, the catabolism of BCAA and skeletal muscle protein would be slowed, maintaining BCAA concentrations in blood., Conclusion: We established, for the first time, the regulatory gene pathways of processes involved in hepatic fibrosis and energy metabolism (hypoalbuminemia, hyperammonemia, and carbohydrate catabolism, and their relationships) under BCAA supplementation., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Soporific acute hyperammonemic hepatic encephalopathy (aHE) can induce considerable changes in cerebral white and gray matter. This report describes a patient in the subacute phase of aHE grade I without disturbed consciousness and with reversible fine laminar cortical involvement on magnetic resonance imaging (MRI). The 59-year-old patient had esophageal varices bleeding due to primary biliary cirrhosis (ammonium blood level: 140 mmoL/L) and presented with sensory Jacksonian seizures, dysarthria, and increased drowsiness and fatigue. MRI revealed patchy hyperintense (T2-weighted, T2w) white-matter lesions and bilateral signal intensities in the striatum (T1w). During a rise of ammonium blood level to 220 mmoL/L, the patient had increased drowsiness, persistent dysarthria and mild temporary hemiparesis without loss of consciousness. Two weeks later, the patient was asymptomatic and blood ammonium level had reverted to normal value. MRI at that time revealed bihemispheric fine laminar subcortical hyperintensities on T2w and fluid-attenuated inversion recovery (FLAIR) imaging, and partially on T1w sequences, with no swelling or restricted diffusion; the hyperintensities were fully reversible a month later. Such a distinct cortical signal increase not only on T2w images, but also on T1w, in a patient after a mild form of aHE is a new MR finding., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

Hyperammonemia and sarcopenia (loss of skeletal muscle) are consistent abnormalities in cirrhosis and portosystemic shunting. We have shown that muscle ubiquitin-proteasome components are not increased with hyperammonemia despite sarcopenia. This suggests that an alternative mechanism of proteolysis contributes to sarcopenia in cirrhosis. We hypothesized that autophagy could be this alternative pathway since we observed increases in classic autophagy markers, increased LC3 lipidation, beclin-1 expression, and p62 degradation in immunoblots of skeletal muscle protein in cirrhotic patients. We observed similar changes in these autophagy markers in the portacaval anastamosis (PCA) rat model. To determine the mechanistic relationship between hyperammonemia and autophagy, we exposed murine C(2)C(12) myotubes to ammonium acetate. Significant increases in LC3 lipidation, beclin-1 expression, and p62 degradation occurred by 1 h, whereas autophagy gene expression (LC3, Atg5, Atg7, beclin-1) increased at 24 h. C(2)C(12) cells stably expressing GFP-LC3 or GFP-mCherry-LC3 constructs showed increased formation of mature autophagosomes supported by electron microscopic studies. Hyperammonemia also increased autophagic flux in mice, as quantified by an in vivo autophagometer. Because hyperammonemia induces nitration of proteins in astrocytes, we quantified global muscle protein nitration in cirrhotic patients, in the PCA rat, and in C(2)C(12) cells treated with ammonium acetate. Increased protein nitration was observed in all of these systems. Furthermore, colocalization of nitrated proteins with GFP-LC3-positive puncta in hyperammonemic C(2)C(12) cells suggested that autophagy is involved in degradation of nitrated proteins. These observations show that increased skeletal muscle autophagy in cirrhosis is mediated by hyperammonemia and may contribute to sarcopenia of cirrhosis.

The effect of artificial acidification of the intestinal content on neurological manifestations of acute severe cyclophosphamide intoxication was studied in rats. The animals were gavaged with 20 ml/kg sulfuric (0.05 M), hydrochloric, boric, or lactic acids (0.1 M) 3 h before intraperitoneal injections of the cytostatic in doses of 0, 200, 600, or 1000 mg/kg. The decrease in pH (by.0) and ammonia-producing activity of the cecal chyme developed within 3 h after administration of acids. Cyclophosphamide caused hyperammonemia; glutamine/ammonia and urea/ammonia ratios in the blood decreased. These changes augmented after administration of acids (boric acid produced maximum and lactic acid minimum effects). Acid treatment resulted in greatest elevation of ammonia level in the portal venous blood and a lesser elevation in the vena cava posterior blood. Acid treatment promoted manifestation of cyclophosphamide neurotoxic effect and animal death. Hence, acidification of the chyme inhibited the formation of ammonia in it, while ammonia release from the gastrointestinal tract into the blood increased; the treatment augmented hyperammonemia and aggravated the neurological manifestations of cyclophosphamide intoxication.

Hepatic encephalopathy (HE) is a potentially fatal complication of acute liver failure, associated with severe neurological dysfunction and coma. The brain's innate immune cells, microglia, have recently been implicated in the pathophysiology of HE. To date, however, only ex vivo studies have been used to characterize microglial involvement. Our study uses in vivo two-photon imaging of awake-behaving mice expressing enhanced green fluorescent protein (eGFP) under the Cx3cr1 promoter to examine microglial involvement in two different models of encephalopathy - a slower, fatal model of azoxymethane-induced HE and a rapid, reversible acute hyperammonemic encephalopathy (AHE) induced by an ammonia load. To investigate the potential contribution of microglia to the neurological deterioration seen in these two models, we developed a software to analyze microglial activation and motility in vivo. In HE, we found that microglia do not become activated prior to the onset of neurological dysfunction, but undergo activation with mildly impaired motility during the terminal stage IV. We demonstrate that this microglial activation coincides with blood-brain barrier (BBB) opening and brain edema. Conversely, both microglial activation and motility are unchanged during AHE, despite the mice developing pathologically increased plasma ammonia and severe neurological dysfunction. Our study indicates that microglial activation does not contribute to the early neurological deterioration observed in either HE or AHE. The late microglial activation in HE may therefore be associated with terminal BBB opening and brain edema, thus exacerbating the progression to coma and increasing mortality., (Copyright © 2012 IBRO. Published by Elsevier Ltd. All rights reserved.)

A 15 year old boy with SMA type II underwent spinal fusion and suffered a mitochondrial Reye-like catabolic crisis 4 days postop with hypoketotic hypoglycemia, lactic acidaemia, hyperammonemia and liver failure, with 90% coagulative necrosis and diffuse macro- and microvesicular steatosis, requiring orthotopic liver transplantation. This crisis responded in part to mitochondrial therapy and anabolic rescue. He made a dramatic sustained neurological recovery, though his post-transplant liver biopsies revealed micro- and macrosteatosis. We hypothesize that a combination of surgical stress-catecholamine induced lipolysis, prolonged general anaesthesia with propofol and sevoflurane, and perioperative fasting on a background of decreased β-oxidation were potential risk factors for the mitochondrial decompensation., (Copyright © 2012 European Paediatric Neurology Society. All rights reserved.)

A male child suffering from generalized tonic clonic epilepsy, on treatment with valproate, developed fulminant hepatic failure, hyperammonemia and encephalopathy due to drug toxicity. The most extraordinary feature was his MRI (FLAIR image) of brain which showed unilateral hyperintensities in right putamen and caudate nucleus. The patient recovered on withdrawal of valproate with mild residual left sided athetotic movements during remission. Repeat investigation confirmed an improved MRI imaging and normalised blood ammonia levels. The case report is unique because of unilateral involvement of basal ganglia due to valproate-induced encephalopathy.

Brain glutamine synthetase (GS) is an integral part of the glutamate-glutamine cycle and occurs in the glial compartment. In vivo Magnetic Resonance Spectroscopy (MRS) allows noninvasive measurements of the concentrations and synthesis rates of metabolites. (15)N MRS is an alternative approach to (13)C MRS. Incorporation of labeled (15)N from ammonia in cerebral glutamine allows to measure several metabolic reactions related to nitrogen metabolism, including the glutamate-glutamine cycle. To measure (15)N incorporation into the position 5N of glutamine and position 2N of glutamate and glutamine, we developed a novel (15)N pulse sequence to simultaneously detect, for the first time, [5-(15)N]Gln and [2-(15)N]Gln+Glu in vivo in the rat brain. In addition, we also measured for the first time in the same experiment localized (1)H spectra for a direct measurement of the net glutamine accumulation. Mathematical modeling of (1)H and (15)N MRS data allowed to reduce the number of assumptions and provided reliable determination of GS (0.30±0.050 μmol/g per minute), apparent neurotransmission (0.26±0.030 μmol/g per minute), glutamate dehydrogenase (0.029±0.002 μmol/g per minute), and net glutamine accumulation (0.033±0.001 μmol/g per minute). These results showed an increase of GS and net glutamine accumulation under hyperammonemia, supporting the concept of their implication in cerebral ammonia detoxification.

Chronic liver failure leads to hyperammonemia, a central component in the pathogenesis of hepatic encephalopathy (HE); however, a correlation between blood ammonia levels and HE severity remains controversial. It is believed oxidative stress plays a role in modulating the effects of hyperammonemia. This study aimed to determine the relationship between chronic hyperammonemia, oxidative stress, and brain edema (BE) in two rat models of HE: portacaval anastomosis (PCA) and bile-duct ligation (BDL). Ammonia and reactive oxygen species (ROS) levels, BE, oxidant and antioxidant enzyme activities, as well as lipid peroxidation were assessed both systemically and centrally in these two different animal models. Then, the effects of allopurinol (xanthine oxidase inhibitor, 100mg/kg for 10days) on ROS and BE and the temporal resolution of ammonia, ROS, and BE were evaluated only in BDL rats. Similar arterial and cerebrospinal fluid ammonia levels were found in PCA and BDL rats, both significantly higher compared to their respective sham-operated controls (p<0.05). BE was detected in BDL rats (p < 0.05) but not in PCA rats. Evidence of oxidative stress was found systemically but not centrally in BDL rats: increased levels of ROS, increased activity of xanthine oxidase (oxidant enzyme), enhanced oxidative modifications on lipids, as well as decreased antioxidant defense. In PCA rats, a preserved oxidant/antioxidant balance was demonstrated. Treatment with allopurinol in BDL rats attenuated both ROS and BE, suggesting systemic oxidative stress is implicated in the pathogenesis of BE. Analysis of ROS and ammonia temporal resolution in the plasma of BDL rats suggests systemic oxidative stress might be an important "first hit", which, followed by increases in ammonia, leads to BE in chronic liver failure. In conclusion, chronic hyperammonemia and oxidative stress in combination lead to the onset of BE in rats with chronic liver failure., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Objectives: Acute liver failure (ALF) produces cerebral dysfunction and edema, mediated in part by elevated ammonia concentrations, often leading to coma and death. The pathophysiology of cerebral edema in ALF is incompletely understood. In vitro models of the cerebral effects of ALF have predominately consisted of dissociated astrocyte cultures or acute brain slices. We describe a stable long-term culture model incorporating both neural and glial elements in a three-dimensional tissue structure offering significant advantages to the study of astrocytic-neuronal interactions in the pathophysiology of cerebral edema and dysfunction in ALF., Methods: We utilized chronic organotypic slice cultures from mouse forebrain, applying ammonium acetate in iso-osmolar fashion for 72 hours. Imaging of slice thickness to assess for tissue swelling was accomplished in living slices with optical coherence tomography, and confocal microscopy of fluorescence immunochemical and histochemical staining served to assess astrocyte and neuronal numbers, morphology, and volume in the fixed brain slices., Results: Ammonia exposure at 1-10 mM produced swelling of immunochemically identified astrocytes, and at 10 mM resulted in macroscopic tissue swelling, with slice thickness increasing by about 30%. Astrocytes were unchanged in number. In contrast, 10 mM ammonia treatment severely disrupted neuronal morphology and reduced neuronal survival at 72 hours by one-half., Discussion: Elevated ammonia produces astrocytic swelling, tissue swelling, and neuronal toxicity in cerebral tissues. Ammonia-treated organotypic brain slice cultures provide an In vitro model of cerebral effects of conditions relevant to ALF, applicable to pathophysiological investigations.

Glutamine deficiency with hyperammonemia due to an inherited defect of glutamine synthetase (GS) was found in a 2 year old patient. He presented neonatal seizures and developed chronic encephalopathy. Thus, GS deficiency leads to severe neurological disease but is not always early lethal., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Reasons for Performing the Study: Intestinal hyperammonaemia (HA) has been infrequently reported in individual horses; however, there have been no studies describing clinical and laboratory data as well as short- and long-term outcome in a larger number of cases., Objectives: To describe clinical and laboratory data and short- and long-term outcome in a large group of horses with intestinal HA., Methods: Multi-centred, retrospective study; case records of horses with HA were reviewed and any horse with a clinical or post mortem diagnosis of intestinal HA was included. Hyperammonaemia was defined as a blood ammonium (NH(4) (+)) concentration ≥60 µmol/l and horses with a diagnosis of primary hepatic disease were excluded. Relevant data were recorded and, if appropriate, data from survivors were compared to nonsurvivors to identify potential prognostic indicators., Results: Thirty-six cases, 26 mature horses and 10 foals with intestinal HA were identified. Case histories included diarrhoea, colic and neurological signs and the most common clinical diagnosis was colitis and/or enteritis. The most common clinical and laboratory abnormalities included tachycardia, increased packed cell volume, hyperlactataemia and hyperglycaemia. Fourteen horses (39%) survived to discharge; NH(4) (+) concentration on admission was the only parameter significantly associated with survival. All surviving horses and foals for which follow-up information was available recovered completely and returned to their intended use without further complications., Conclusions and Potential Relevance: Intestinal HA occurs in mature horses and foals and can be associated with severe clinical and laboratory abnormalities; further studies are required to investigate predisposing factors and delineate possible differences in aetiologies., (© 2010 EVJ Ltd.)

Background and Purpose: Acute hyperammonemic encephalopathy has significant morbidity and mortality unless promptly treated. We describe the MR imaging findings of acute hyperammonemic encephalopathy, which are not well-recognized in adult patients., Materials and Methods: We retrospectively reviewed the clinical and imaging data and outcome of consecutive patients with documented hyperammonemic encephalopathy seen at our institution. All patients underwent cranial MR imaging at 1.5T., Results: Four patients (2 women; mean age, 42 ± 13 years; range, 24-55 years) were included. Causes included acute fulminant hepatic failure, and sepsis with a background of chronic hepatic failure and post-heart-lung transplantation with various systemic complications. Plasma ammonia levels ranged from 55 to 168 μmol/L. Bilateral symmetric signal-intensity abnormalities, often with associated restricted diffusion involving the insular cortex and cingulate gyrus, were seen in all cases, with additional cortical involvement commonly seen elsewhere but much more variable and asymmetric. Involvement of the subcortical white matter was seen in 1 patient only. Another patient showed involvement of the basal ganglia, thalami, and midbrain. Two patients died (1 with fulminant cerebral edema), and 2 patients survived (1 neurologically intact and the other with significant intellectual impairment)., Conclusions: The striking common imaging finding was symmetric involvement of the cingulate gyrus and insular cortex in all patients, with more variable and asymmetric additional cortical involvement. These specific imaging features should alert the radiologist to the possibility of acute hyperammonemic encephalopathy.

Neuropathology and neuroimaging of long-term survival cases of arginase deficiency are rarely reported. The magnetic resonance imaging (MRI) of our case showed severe multicystic white matter lesions with cortical atrophy, which were more severe compared with previous reports. In this patient, low-protein diet successfully reduced hyperammonemia, but hyperargininemia persisted. These severe neurological and MRI findings may be explained by a compound heterozygote, inheriting both of severe mutant alleles from her parents., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Many theories have been advanced to explain the encephalopathy associated with chronic liver disease and with the less common acute form. A major factor contributing to hepatic encephalopathy is hyperammonemia resulting from portacaval shunting and/or liver damage. However, an increasing number of causes of hyperammonemic encephalopathy have been discovered that present with the same clinical and laboratory features found in acute liver failure, but without liver failure. Here, we critically review the physiology, pathology, and biochemistry of ammonia (i.e., NH3 plus NH4+) and show how these elements interact to constitute a syndrome that clinicians refer to as hyperammonemic encephalopathy (i.e., acute liver failure, fulminant hepatic failure, chronic liver disease). Included will be a brief history of the status of ammonia and the centrality of the astrocyte in brain nitrogen metabolism. Ammonia is normally detoxified in the liver and extrahepatic tissues by conversion to urea and glutamine, respectively. In the brain, glutamine synthesis is largely confined to astrocytes, and it is generally accepted that in hyperammonemia excess glutamine compromises astrocyte morphology and function. Mechanisms postulated to account for this toxicity will be examined with emphasis on the osmotic effects of excess glutamine (the osmotic gliopathy theory). Because hyperammonemia causes osmotic stress and encephalopathy in patients with normal or abnormal liver function alike, the term "hyperammonemic encephalopathy" can be broadly applied to encephalopathy resulting from liver disease and from various other diseases that produce hyperammonemia. Finally, the possibility that a brain glutamine synthetase inhibitor may be of therapeutic benefit, especially in the acute form of liver disease, is discussed., (Copyright © 2010 The American Society for Experimental NeuroTherapeutics, Inc. Published by Elsevier Inc. All rights reserved.)

Background and Purpose: In acute hepatic encephalopathy, MR imaging abnormalities have been described in the PVWM, thalami, and corticospinal tracts. We sought to determine characteristic regions of involvement on FLAIR and DWI, to evaluate their reversibility, and to correlate MR imaging extent with clinical severity., Materials and Methods: Twenty patients who presented clinically with acute hepatic encephalopathy and MR imaging <21 days after symptom onset were reviewed retrospectively. Two neuroradiologists recorded involved regions on FLAIR and DWI in each, measured ADC values in affected regions and NAWM, and scored the MR imaging severity/extent. The initial severity (West Haven grade), follow-up clinical severity (degree of improvement), and maximal PAL within ±8 days of MR imaging were recorded and correlated with the MR imaging severity., Results: On FLAIR and DWI respectively, there were abnormalities in the thalami (85%, 70%), PLIC (75%, 80%), PVWM (80%, 85%), and DBS (70%, 35%) and diffuse cortical involvement (30%, 25%). There were relatively strong significant (P < .005) correlations of FLAIR (r = 0.680, P = .001) and DWI severity (r = 0.690, P = .001) with PAL, and of PAL with the clinical outcome (r = 0.691, P = .001). Both FLAIR (r = 0.592, P = .006) and DWI (r = 0.487, P = .029) severity correlated moderately with the clinical outcome but were not significant at the P < .005 level after Bonferroni correction., Conclusions: Patients with acute hepatic encephalopathy may exhibit characteristic regions of involvement on FLAIR with DWI findings that can be reversible. The MR imaging extent on FLAIR and DWI strongly correlates with the maximal PAL, and PAL correlates well with the clinical outcome. Diffuse cortical involvement has a higher potential for neurologic sequelae but can be reversible.

Background & Aims: Hyperammonemia and inflammation cooperate to induce neurological alterations in hepatic encephalopathy. Recent studies in animal models suggest that chronic hyperammonemia and neuroinflammation impair learning ability by the same mechanism. Chronic hyperammonemia might induce inflammatory factors in the brain that impair cognitive function. We sought to determine whether hyperammonemia itself induces neuroinflammation, whether ammonia-induced neuroinflammation mediates cognitive impairment, and whether neuroinflammation also occurs in rats with bile duct ligation (BDL rats)-a model of chronic liver injury that results in hyperammonemia and hepatic encephalopathy., Methods: Chronic moderate hyperammonemia was induced by feeding male Wistar rats an ammonium-containing diet or performing BDL. Rats that received a standard diet or a sham operation were used as controls. Neuroinflammation was assessed by measuring activation of microglia and inflammatory factors. Brain samples were collected from hyperammonemic and BDL rats; microglial activation was determined by immunohistochemistry and quantification of inflammatory markers (ie, inducible nitric oxide synthase, interleukin-1beta, and prostaglandin E2). Learning ability and motor activity were assessed in hyperammonemic and BDL rats given ibuprofen as an anti-inflammatory agent., Results: Chronic moderate hyperammonemia or BDL activated the microglia, especially in cerebellum; increased inducible nitric oxide synthase, interleukin-1beta, and prostaglandin E2 levels; and impaired cognitive and motor function, compared with controls. Ibuprofen reduced microglial activation and restored cognitive and motor functions in the hyperammonemic and BDL rats., Conclusions: Chronic hyperammonemia is sufficient to induce microglial activation and neuroinflammation; these contribute to the cognitive and motor alterations that occur during hepatic encephalopathy., (Copyright (c) 2010 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Urea cycle disorders are congenital metabolic disorders that often cause episodic hyperammonemia. Neuroimaging in episodic hyperammonemia demonstrates several patterns of brain injuries, including focal lesions in the lentiform nucleus, insula, cingulate gyrus, and perirolandic fissure, as well as diffuse cerebral edema. In cases with neonatal onset of hyperammonemia, similar lesions have also been reported. We herein report a boy with severe neonatal hyperammonemia caused by ornithine transcarbamylase deficiency. He presented with parieto-occipital encephalomalacia, which resembles severe neonatal hypoglycemia on magnetic resonance imaging. This radiological finding may indicate parieto-occipital vulnerability not only to hypoglycemia but also to hyperammonemia., (Copyright 2009 Elsevier B.V. All rights reserved.)

Background and Purpose: MR imaging and (1)H-MR spectroscopy changes are well reported in cirrhotic patients, whereas they are inadequately reported in EHPVO. The aim of this study was to investigate age-related changes in brain MR imaging and metabolite profile in EHPVO with and without MHE and to explore any correlation of imaging and (1)H-MR spectroscopy parameters with blood ammonia., Materials and Methods: Sixty-three patients with EHPVO (children, 7-12 years [n = 22], adolescents, 13-18 years [n = 15] and adults, 19-41 years [n = 26]) and 47 healthy age/sex-matched volunteers were studied. Neuropsychological tests, MR imaging, (1)H-MR spectroscopy, and blood ammonia estimation were performed in all subjects., Results: Of 63 EHPVO patients, 25 (40%) who had MHE showed significantly increased MD, Glx, and blood ammonia in all 3 age groups; however, myo-inositol was significantly lower only in adults when compared with controls. MD positively correlated with blood ammonia and Glx in all age groups. Brain choline levels were normal in all patients with different age groups., Conclusions: Increases in brain MD, Glx, and blood ammonia were associated with MHE in all age groups. Normal brain choline in EHPVO signifies healthy liver and may serve as a diagnostic marker for its differentiation from cirrhosis-induced encephalopathy. Significant decrease of myo-inositol in adults is probably due to cellular osmoregulation secondary to long-standing hyperammonemia.

Aims: The overexpression of alpha-synuclein has been associated with neurodegenerative diseases, especially when the protein aggregates to form insoluble structures. The present study examined the effect of chronic hyperammonaemia on alpha-synuclein expression in the rat cerebellum following portacaval anastomosis (PCA)., Methods: Immunohistochemical and western blot determinations were performed 1 month and 6 months after the PCA procedure., Results: A time-dependent increase in alpha-synuclein expression was seen in the cerebellar grey matter compared with the controls. At 1 month post PCA, alpha-synuclein-immunopositive material was observed in the molecular layer, while the Purkinje cells showed weak alpha-synuclein expression, and alpha-synuclein aggregates were observed throughout the granular layer. At 6 months post PCA, alpha-synuclein expression was significantly increased compared with the controls. alpha-synuclein-immunostained astroglial cells were also found; the Bergmann glial cells showed alpha-synuclein-positive processes in the molecular layer of PCA-exposed rats, and in the granular layer, perivascular astrocytes showed intense alpha-synuclein immunoreactivity, as indicated by colocalization of alpha-synuclein with glial fibrillary acidic protein (GFAP). In addition, ubiquitin-immunoreactive inclusions were present in PCA-exposed rats, although they did not colocalize with alpha-synuclein. Western blotting performed at 6 months post PCA showed a reduction in the level of soluble alpha-synuclein compared with 1 month post PCA and the controls; this reduction was concomitant with an increase in the insoluble form of alpha-synuclein., Conclusions: Although the precise mechanism by which alpha-synuclein aggregates in PCA-treated rats remains unknown, the present data suggest an important role for this protein in the onset and progression of hepatic encephalopathy, probably via its expression in astroglial cells.

Introduction: Animal models used to study hyperammonaemic disorders related to chronic liver disease are unsatisfactory. These animals only develop hyperammonaemia and brain oedema when fed with diets supplemented with amonium acetate., Aim: To develop a novel experimental model of hyperammonaemia and brain oedema in CCl(4)-induced cirrhosis in rats., Methods: Four groups were studied: rats with sham intervention (S), rats with total portal vein ligation (TPVL), cirrhotic rats (LC), and cirrhotic rats with TPVL (LC+TPVL). When ascites was diagnosed, oral glutamine challenge (OGC) test was performed. Blood, liver, lungs and brain samples were collected to quantify liver function parameters, plasmatic and cerebral ammonia, endotoxaemia, liver and brain histology, brain oedema and portosystemic shunting degree., Results: LC+TPVL rats showed a significant increase in portosystemic shunting when compared with LC group and a significant derangement in liver function when compared with TPVL group. These alterations resulted in a significant increase in plasmatic and brain ammonia concentrations and a higher plasmatic endotoxaemia as compared with others. Similarly, the area under OGC curve was significantly increased in LC+TPVL group as compared with the others, and correlates with portal shunting. Low-grade brain oedema was only observed in LC+TPVL group. All cirrhotic groups showed liver regeneration nodules and type-II Alzheimer astrocytes, Conclusion: LC+TPVL reproduce the main alterations - portosystemic shunting, plasmatic and cerebral hyperammonaemia and low-grade brain oedema - observed in cirrhotic patients with hepatic encephalopathy.

There is a lack of awareness of acutely presenting inborn errors of metabolism in adults, of which the X-linked urea cycle defect ornithine transcarbamylase (OTC) deficiency is an example, many comparatively mild mutations having been identified. In male hemizygotes clinical manifestations and age at presentation vary and depend on the mutation. In female heterozygotes the clinical spectrum depends on the extent to which the abnormal gene is expressed. Milder versions of the defect may not cause clear clinical symptoms and may remain unrecognized until the person is subjected to an unusually high nitrogen load when they develop severe hyperammonaemia. During acute episodes liver enzymes may be normal or only slightly elevated and occasionally accompanied by coagulopathy, but the key finding is hyperammonaemia. Boys with these milder forms may exhibit abnormal behaviour and be diagnosed with attention deficit hyperactivity disorder. This case illustrates how late presentation of OTC deficiency in a non-specialist centre can be difficult to differentiate from drug abuse, psychiatric illness or encephalopathy. Failure to measure blood ammonia in adults with unexplained key symptoms - particularly prolonged vomiting without diarrhoea and altered mental state/hallucinations, or to recognize the significance of elevated blood ammonia without evidence of liver decompensation can lead to delayed or missed diagnosis.

Reduced function of the glutamate--nitric oxide (NO)--cGMP pathway is responsible for some cognitive alterations in rats with hyperammonemia and hepatic encephalopathy. Hyperammonemia impairs the pathway in cerebellum by increasing neuronal nitric oxide synthase (nNOS) phosphorylation in Ser847 by calcium-calmodulin-dependent protein kinase II (CaMKII), reducing nNOS activity, and by reducing nNOS amount in synaptic membranes, which reduces its activation following NMDA receptors activation. The reason for increased CaMKII activity in hyperammonemia remains unknown. We hypothesized that it would be as a result of increased tonic activation of NMDA receptors. The aims of this work were to assess: (i) whether tonic NMDA activation receptors is increased in cerebellum in chronic hyperammonemia in vivo; and (ii) whether this tonic activation is responsible for increased CaMKII activity and reduced activity of nNOS and of the glutamate--NO--cGMP pathway. Blocking NMDA receptors with MK-801 increases cGMP and NO metabolites in cerebellum in vivo and in slices from hyperammonemic rats. This is because of reduced phosphorylation and activity of CaMKII, leading to normalization of nNOS phosphorylation and activity. MK-801 also increases nNOS in synaptic membranes and reduces it in cytosol. This indicates that hyperammonemia increases tonic activation of NMDA receptors leading to reduced activity of nNOS and of the glutamate--NO--cGMP pathway.

Cranial magnetic resonance imaging was performed in three cases of acute hyperammonemic encephalopathy with three diverse etiologies: infantile citrullinemia, acute hepatic encephalopathy, and proximal urea cycle disorder. All three patients exhibited diffuse extensive cortical signal changes and swelling. Neurologic outcome was poor in all three cases. Knowledge of the magnetic resonance imaging findings of hyperammonemic encephalopathy may help in early diagnosis and treatment and could influence the neurologic outcome.

Plant flavonoids are emerging as potent therapeutic drugs effective against a wide range of free radical-mediated diseases. Morin (3,5,7,2',4'-pentahydroxyflavone), a member of flavonols, is an important bioactive compound by interacting with nucleic acids, enzymes and protein. In this study, we found that morin (30 mg/kg body weight) by oral administration offers protection against hyperammonemia by means of reducing blood ammonia, oxidative stress and enhancing antioxidant status in ammonium chloride-induced (100 mg/kg body weight; i.p) hyperammonemic rats. Enhanced blood ammonia, plasma urea, lipid peroxidation in circulation and tissues (liver and brain) of ammonium chloride-treated rats was accompanied by a significant decrease in the tissues levels of superoxide dismutase (SOD), catalase, reduced glutathione (GSH) and glutathione peroxidase (GPx). Morin administered rats showed a significant reduction in ammonia, urea, lipid peroxidation with a simultaneous elevation in antioxidant levels. Cotreatment with morin prevented the elevation of liver marker enzymes induced by ammonium chloride. The body weight of the animals decreased significantly on ammonium chloride administration when compared with control group. However, cotreatment with morin significantly prevented the decrease of the body weight caused by ammonium chloride. Hyperammonemic rats show liver fibrosis, steatosis, sinusoidal dilatation, etc., along with necrosis, microcystic degeneration in brain. All these changes were reduced in hyperammonemic rats treated with Morin, which too correlated with the biochemical observations. In conclusion, these findings indicate that morin exert antioxidant potential and offer protection against ammonium chloride-induced hyperammonemia. But the exact underlying mechanism needs to be elucidated.

Acute liver failure (ALF) may lead to rapid death unless the patients receive a liver for transplantation. However, the number of livers available is not enough and a number of patients die before a suitable liver is available for transplantation. The liver has a high capacity for regeneration which may allow complete recovery even in patients with severe liver failure. It would be therefore very useful to have procedures to prevent or delay the mechanisms by which ALF leads to death. These mechanisms are no well understood. Progression of ALF leads to multi-organ failure, systemic inflammatory response, hepatic encephalopathy, cerebral oedema and increased intracranial pressure, which seem the most important immediate causes of mortality in patients with ALF. A main contributor to these events is hyperammonemia, due to impaired ammonia detoxification in the liver. Acute hyperammonemia per se leads to death, which is mediated by activation of the NMDA type of glutamate receptors in brain and may be prevented by antagonists blocking these receptors. Acute liver failure also leads to hyperammonemia and excessive activation of NMDA receptors in brain which contributes to ALF-induced death. Sustained blocking of NMDA receptors by continuous administration of the antagonists MK-801 or memantine increases about twice the survival time of rats with severe ALF due to injection of 2.5g/kg of galactosamine. In rats with milder ALF due to injection of 1.5g/kg of galactosamine, blocking NMDA receptors increases the percentage of surviving rats from 23% to 62% and increases about twice the survival time of the rats which die. These data strongly support that blocking NMDA receptors would improve survival of patients with ALF, either by allowing more time for liver regeneration or to get a liver suitable for transplantation.

Even though ammonia is considered to underlie nervous system symptoms of dysfunction during hyperammonemia, lactate, which increases as a metabolic consequence of high ammonia levels, might also be a contributing factor. The data presented here show that NH4Cl (5 mM) mediates astroglial cell swelling, and that treatment with NH4Cl or lactate (25 mM) causes rearrangements of actin filaments and reduces astroglial glutamate uptake capacity. Co-application with BaCl2, which blocks astroglial uptake of NH4+, prevents NH4Cl-mediated cell swelling and rearrangement of actin filaments, but does not reduce NH4Cl-induced glutamate uptake capacity inhibition. Neither NH4Cl nor lactate affected glutamate uptake or protein expression in microglial cultures, indicating that astroglial cells are more susceptible to the neurotoxic affects of ammonia. Our results suggest that ammonium underlies brain edema, but that lactate can contribute to some of the cellular dysfunctions associated with elevated cerebral levels of ammonia.

Methylmalonic acidemia caused by an l-methylmalonyl-CoA mutase deficiency. The mut(0) type is associated with significant mortality and morbidity, but tandem mass spectrometry has made early detection possible. Five patients were identified through newborn screening for elevated propionylcarnitine (C3-carnitine) levels. These patients received a positive screening result at a median age of 10 days (range, 5-18 days). When treated at a median age of 11 days (range, 3-50 days), 2 patients were asymptomatic, and only one was significantly acidotic (pH <7.2), but all had various degrees of hyperammonemia (range, 127-1,244 mumol/L). Magnetic resonance imaging of the brain was performed in 4 patients shortly after diagnosis, and the results were all abnormal. Four patients were followed. There was no further metabolic decompensation after the initial episodes, but their mean developmental quotient was only 50. These results suggest that early hyperammonemia can lead to significant brain damage in methylmalonic acidemia. Therefore, treatment of this disease in newborns must be more aggressive.

The term hepatic encephalopathy (HE) includes a spectrum of neuropsychiatric abnormalities occurring in patients with liver dysfunction. Most cases are associated with cirrhosis and portal hypertension or portal-systemic shunts, but the condition can also be seen in patients with acute liver failure and, rarely, with portal-systemic bypass and no associated intrinsic hepatocellular disease. Although HE is a clinical condition, several neuroimaging techniques, particularly MR imaging, may eventually be useful for the diagnosis because they can identify and measure the consequences of central nervous system (CNS) increase in substances that under normal circumstances, are efficiently metabolized by the liver. Classic MR imaging abnormalities include high signal intensity in the globus pallidum on T1-weighted images, likely a reflection of increased tissue concentrations of manganese, and an elevated glutamine/glutamate peak coupled with decreased myo-inositol and choline signals on proton MR spectroscopy, representing disturbances in cell-volume homeostasis secondary to brain hyperammonemia. Recent data have shown that white matter abnormalities, also related to increased CNS ammonia concentration, can also be detected with several MR imaging techniques such as magnetization transfer ratio measurements, fast fluid-attenuated inversion recovery sequences, and diffusion-weighted images. All these MR imaging abnormalities, which return to normal with restoration of liver function, probably reflect the presence of mild diffuse brain edema, which seems to play an essential role in the pathogenesis of HE. It is likely that MR imaging will be increasingly used to evaluate the mechanisms involved in the pathogenesis of HE and to assess the effects of therapeutic measures focused on correcting brain edema in these patients.

Acute hyperammonaemia (HA) causes cerebral oedema and severe brain damage in patients with urea cycle disorders (UCDs) or acute liver failure (ALF). Chronic HA is associated with developmental delay and intellectual disability in patients with UCDs and with neuropsychiatric symptoms in patients with chronic liver failure. Treatment often cannot prevent severe brain injury and neurological sequelae. The causes of the brain oedema in hyperammonaemic encephalopathy (HAE) have been subject of intense controversy among physicians and scientists working in this field. Currently favoured hypotheses are astrocyte swelling due to increased intracellular glutamine content and neuronal cell death due to excitotoxicity caused by elevated extracellular glutamate levels. While many researchers focus on these mechanisms of cytotoxicity, others emphasize vascular causes of brain oedema. New data gleaned from expression profiling of astrocytes acutely isolated from hyperammonaemic mouse brains point to disturbed water and potassium homeostasis as regulated by astrocytes at the brain microvasculature and in the perisynaptic space as a potential mechanism of brain oedema development in hyperammonaemia.

Acute hyperammonemia (HA) causes cerebral edema and brain damage in children with urea cycle disorders (UCDs) and in patients in acute liver failure. Chronic HA is associated with developmental delay and mental retardation in children with UCDs, and with neuropsychiatric symptoms in patients with chronic liver failure. Astrocytes are a major cellular target of hyperammonemic encephalopathy, and changes occurring in these cells are thought to be causally related to the brain edema of acute HA. To study the effect of HA on astrocytes in vivo, we crossed the Otc(spf) mouse, a mouse with the X-linked UCD ornithine transcarbamylase (OTC) deficiency, with the hGFAP-EGFP mouse, a mouse selectively expressing green fluorescent protein in astrocytes. We used FACS to purify astrocytes from the brains of hyperammonemic and healthy Otcspf/GFAP-EGFP mice. RNA isolated from these astrocytes was used in microarray expression analyses and qRT-PCR. When compared with healthy littermates, we observed a significant downregulation of the gap-junction channel connexin 43 (Cx43) the water channel aquaporin 4 (Aqp4) genes, and the astrocytic inward-rectifying potassium channel (Kir) genes Kir4.1 and Kir5.1 in hyperammonemic mice. Aqp4, Cx43, and Kir4.1/Kir5.1 are co-localized to astrocytic end-feet at the brain vasculature, where they regulate potassium and water transport. Since, NH4+ ions can permeate water and K+-channels, downregulation of these three channels may be a direct effect of elevated blood ammonia levels. Our results suggest that alterations in astrocyte-mediated water and potassium homeostasis in brain may be key to the development of the brain edema., ((Copyright) 2008 Wiley-Liss, Inc.)

Objectives: To review signs and symptoms of valproate-induced hyperammonaemic encephalopathy without hepatotoxicity in the psychiatric setting, explore its mechanisms, and give recommendations for prevention and treatment., Methods: Medline search with keywords valproate, ammonia, hyperammonaemia, encephalopathy, and then cross-references to articles obtained through this search. Only cases with indication of valproate for psychiatric condition were included., Results: Fourteen cases published in the psychiatric setting are reviewed. Valproate-induced hyperammonaemic encephalopathy is a rare adverse event, occurring almost equally in men and women, with a large age range, and reported in two patients with mental retardation. Symptoms appeared either a few days after initiation of valproate therapy, or after several months or years. The main symptoms were fluctuations in consciousness and disorientation. Clinical severity was not related to blood ammonia levels. All patients recovered after valproate-induced hyperammonaemic encephalopathy diagnosis and treatment, usually involving discontinuation of valproate., Conclusions: Valproate-induced hyperammonaemic encephalopathy is rare and usually reversible in patients without urea cycle disorders when valproate is discontinued. Therapy with carnitine is recommended. Special caution should be used in patients with mental retardation. Psychiatrists should suspect valproate-induced hyperammonaemic encephalopathy when consciousness deteriorates.

Hepatic myelopathy is a rare neurological complication in patients with chronic liver failure and most patients who suffered from this disorder were demonstrated to have portal-systemic shunt. A 31-year-old man who was diagnosed as having adult-onset type II citrullinemia (CTLN2) and had a six-year history of recurrent hepatic encephalopathy showed progressive spastic paraparesis with no involvement of sensation and sphincter function. Examinations of cerebrospinal fluid and spinal MRI were normal. He suddenly died of acute exacerbation of hepatic encephalopathy with severe brain edema. The pathology of the spinal cord disclosed a localized degeneration of both lateral columns, the lesion being more remarkable in the lower segments of the cord. These clinical and pathological findings of hepatic myelopathy have not been noted in the many patients with CTLN2 previously reported, and our patient is unique in developing hepatic myelopathy without porto-caval shunting. Thus, repeated attacks of encephalopathy with hyperammonemia might secondarily have induced the myelopathy in this patient.

Acid-sensitive ion channels (ASICs) are proton-gated and belong to the family of degenerin channels. In the mammalian nervous system, ASICs are most well known in sensory neurons, where they are involved in nociception, occurring when injury or inflammation causes acidification. ASICs also are widely expressed in the CNS, and some synaptic roles have been revealed. Because neuronal activity can produce pH changes, ASICs may respond to local acidic transients and alter the excitability of neuronal circuits more widely than is presently appreciated. Furthermore, ASICs have been found to underlie calcium transients that contribute to neuronal death. Degeneration of midbrain dopamine neurons is characteristic of advanced idiopathic Parkinson's disease. Therefore, we tested for functional ASICs in midbrain dopamine neurons of the ventral tegmental area and substantia nigra compacta. Patch-clamp electrophysiology applied to murine midbrain slices revealed abundant acid-sensitive channels. The ASICs were gated and desensitized by extracellular application of millimolar concentrations of NH(4)Cl. Although the NH(4)Cl solution contains micromolar concentrations of NH(3) at pH 7.4, our evidence indicates that NH(4)(+) gates the ASICs. The proton-gated and the ammonium-gated currents were inhibited by tarantula venom (psalmotoxin), which is specific for the ASIC1a subtype. The results show that acid-sensitive channels are expressed in midbrain dopamine neurons and suggest that ammonium sensitivity is a widely distributed ASIC characteristic in the CNS, including the hippocampus. The ammonium sensitivity suggests a role for ASIC1s in hepatic encephalopathy, cirrhosis, and other neuronal disorders that are associated with hyperammonemia.

The availability of an animal model is crucial in studying the pathophysiological mechanisms of disease and to test possible therapies. Now, there are several models for the study of liver diseases, but there still remains a lack of a satisfactory animal model of chronic liver disease with hepatic encephalopathy (HE) and abnormalities in nitrogen metabolism, as seen in humans. In rats, two models of chronic HE are widely used: rats after portacaval anastomosis (PCA) and rats with chronic hyperammonemia. The first one mimics the situation induced in cirrhosis by collateral circulation, and has the problem of the absence of hepatocellular injury. The model of hyperammonemia is useful to study the effect of ammonia as a brain toxic substance, but also lacks liver failure. Bile-duct ligation has been used to induce cirrhosis and could also be a model of HE, probably with the addition of a precipitant factor. An ideal model of HE in chronic liver disease must have liver cirrhosis and a precipitant factor of HE; it must also show neuropathological characteristic findings of HE, neurochemical alterations in the main pathways impaired in these complications of cirrhosis, and low-grade brain edema.

From 1995-2002, 14 patients with predominantly prehepatic, noncirrhotic portal hypertension were evaluated. At presentation, the eight females and six males had a mean age of 9 years (range 2-18). Seven were admitted with gastrointestinal, mostly esophageal bleeding, three with splenomegaly, three with hepato-pulmonary syndrome, and one with hyperammonemia. Imaging studies showed portal vein obstruction in six patients and non-obstructed but frequently anomalous vascular patterns, including hypoplasia of the portal vein, in the remaining eight patients. At the onset, liver function was marginally abnormal in all patients, but thrombocytopenia of approximately 100 x 10(9)/L was consistently observed, probably reflecting chronic mild consumption coagulopathy and hypersplenism. The most striking and frequent histopathologic finding in 25 liver samples, was the presence of hypoplastic portal triads with collapsed portal vein radicles. In contrast, other triads showed markedly distended and misshapen portal vein radicles and likely lymphatics. These two patterns of collapse and distention presumably reflect areas of impaired versus overloaded intrahepatic portal venous flow. Some of the biopsies showed variable portal/sinusoidal fibrosis. Four patients (two with intestinal bleeding, two with hepatopulmonary syndrome) required liver transplants and are doing well. Eight patients are doing well clinically after surgical or spontaneous vascular shunting. Two patients with intestinal bleeding and hepato-pulmonary syndrome, respectively) who had congenital dyskeratosis, underwent bone marrow transplantation and died of nonhepatic-related complications. It is possible to suggest prehepatic causes of portal hypertension even in needle biopsies when collapsed portal vein radicles are present in portal triads, but more than one biopsy sample with larger bore bioptomes may be required to see the changes. Conversely, identifying these changes may suggest to the clinicians the need to work-up a patient for portal hypertension.

Bone marrow transplantation (BMT) has been shown to reverse or stabilize some manifestations of mucopolysaccharidosis I (Hurler syndrome). Idiopathic hyperammonemia (IHA) is a rare complication of solid organ and BMT that is characterized by elevated serum ammonia, normal liver enzymes, and abrupt onset of neurologic deterioration. We present the case of a 14-month-old male patient with Hurler syndrome who developed fatal IHA (ammonia = 2297 micromol/L) 31 days after a cord blood transplant. A complete autopsy was performed, with examination of both frozen and formalin-fixed paraffin-embedded (FFPE) tissues using a variety of special stains and electron microscopy. Hyperammonemia was documented by analysis of antemortem serum and postmortem cerebrospinal and vitreous fluid. Other causes of hyperammonemia, including Reye syndrome, were excluded. Histologic changes included centrilobular microvesicular steatosis of the liver and storage product present in multiple organs. The highly water-soluble mucopolysaccharide (MPS) storage product was best identified by colloidal iron staining of FFPE and unfixed air-dried fresh frozen liver sections. Alcian blue stains failed to convincingly demonstrate MPS in any of the liver sections. This is the first published report, to our knowledge, of IHA in a posttransplant patient younger than 18 years old or following transplantation for Hurler syndrome. Demonstration of the hepatic centrilobular microvesicular steatosis characteristic of IHA was complicated by the diffuse storage of MPS within the liver. MPS storage can be best detected in the liver using colloidal iron staining. Oil-red-O staining may be useful to document microvesicular steatosis in cases with a clinical history of hyperammonemia following solid organ or BMT. Determining if certain subsets of children are at increased risk for IHA requires further study.

Glutamine synthetase (GS) in brain is located mainly in astrocytes. One of the primary roles of astrocytes is to protect neurons against excitotoxicity by taking up excess ammonia and glutamate and converting it into glutamine via the enzyme GS. Changes in GS expression may reflect changes in astroglial function, which can affect neuronal functions. Hyperammonemia is an important factor responsible of hepatic encephalopathy (HE) and causes astroglial swelling. Hyperammonemia can be experimentally induced and an adaptive astroglial response to high levels of ammonia and glutamate seems to occur in long-term studies. In hyperammonemic states, astroglial cells can experience morphological changes that may alter different astrocyte functions, such as protein synthesis or neurotransmitters uptake. One of the observed changes is the increase in the GS expression in astrocytes located in glutamatergic areas. The induction of GS expression in these specific areas would balance the increased ammonia and glutamate uptake and protect against neuronal degeneration, whereas, decrease of GS expression in non-glutamatergic areas could disrupt the neuron-glial metabolic interactions as a consequence of hyperammonemia. Induction of GS has been described in astrocytes in response to the action of glutamate on active glutamate receptors. The over-stimulation of glutamate receptors may also favour nitric oxide (NO) formation by activation of NO synthase (NOS), and NO has been implicated in the pathogenesis of several CNS diseases. Hyperammonemia could induce the formation of inducible NOS in astroglial cells, with the consequent NO formation, deactivation of GS and dawn-regulation of glutamate uptake. However, in glutamatergic areas, the distribution of both glial glutamate receptors and glial glutamate transporters parallels the GS location, suggesting a functional coupling between glutamate uptake and degradation by glutamate transporters and GS to attenuate brain injury in these areas. In hyperammonemia, the astroglial cells located in proximity to blood-vessels in glutamatergic areas show increased GS protein content in their perivascular processes. Since ammonia freely crosses the blood-brain barrier (BBB) and astrocytes are responsible for maintaining the BBB, the presence of GS in the perivascular processes could produce a rapid glutamine synthesis to be released into blood. It could, therefore, prevent the entry of high amounts of ammonia from circulation to attenuate neurotoxicity. The changes in the distribution of this critical enzyme suggests that the glutamate-glutamine cycle may be differentially impaired in hyperammonemic states.

Alzheimer's disease (AD) is the most common age-related neurodegenerative disorder. Behavioural, cognitive and memory dysfunctions are characteristic symptoms of AD. The formation of amyloid plaques is currently considered as the key event of AD. Other histological hallmarks of the disease are the formation of fibrillary tangles, astrocytosis, and loss of certain neuronal systems in cortical areas of the brain. A great number of possible aetiologic and pathogenetic factors of AD have been published in the course of the last two decades. Among the toxic factors, which have been considered to contribute to the symptoms and progression of AD, ammonia deserves special interest for the following reasons: (a) Ammonia is formed in nearly all tissues and organs of the vertebrate organism; it is the most common endogenous neurotoxic compounds. Its effects on glutamatergic and GABAergic neuronal systems, the two prevailing neuronal systems of the cortical structures, are known for many years. (b) The impairment of ammonia detoxification invariably leads to severe pathology. Several symptoms and histologic aberrations of hepatic encephalopathy (HE), of which ammonia has been recognised as a pathogenetic factor, resemble those of AD. (c) The excessive formation of ammonia in the brains of AD patients has been demonstrated, and it has been shown that some AD patients exhibit elevated blood ammonia concentrations. (d) There is evidence for the involvement of aberrant lysosomal processing of beta-amyloid precursor protein (beta-APP) in the formation of amyloid deposits. Ammonia is the most important natural modulator of lysosomal protein processing. (e) Inflammatory processes and activation of microglia are widely believed to be implicated in the pathology of AD. Ammonia is able to affect the characteristic functions of microglia, such as endocytosis, and cytokine production. Based on these facts, an ammonia hypothesis of AD has first been suggested in 1993. In the present review old and new observations are discussed, which are in support of the notion that ammonia is a factor able to produce symptoms of AD and to affect the progression of the disease.

Hyperammonemia resulting from inherited urea cycle enzyme deficiencies or liver failure results in severe central nervous system dysfunction including brain edema, convulsions and coma. Neuropathologic evaluation in these disorders reveals characteristic alterations of astrocyte morphology ranging from cell swelling (acute hyperammonemia) to Alzheimer Type II astrocytosis (chronic hyperammonemia). Having no effective urea cycle, brain relies on glutamine synthesis for the removal of excess ammonia and the enzyme responsible, glutamine synthetase, has a predominantly astrocytic localization. Accumulation of ammonia in brain results in a redistribution of cerebral blood flow and metabolism from cortical to sub-cortical structures. In addition to changes in astrocyte morphology, increased brain ammonia concentrations result in alterations in expression of key astrocyte proteins including glial fibrillary acidic protein, glutamate and glycine transporters and "peripheral-type" (mitochondrial) benzodiazepine receptors. Such changes result in alterations of astrocytic volume and increased extracellular concentrations of excitatory and inhibitory substances. In addition, the ammonium ion has direct effects on excitatory-inhibitory transmission via distinct mechanisms involving cellular chloride extrusion and postsynaptic receptor function. Acute ammonia exposure leads to activation of NMDA receptors and their signal transduction pathways. Chronic hyperammonemia also results in increased concentrations of neuroactive L-tryptophan metabolites including serotonin and quinolinic acid. Therapy in hyperammonemic syndromes continues to rely on ammonia-lowering strategies via peripheral mechanisms (reduction of ammonia production in the gastrointestinal tract, increased ammonia removal by muscle)., (Copyright 2002 Elsevier Science Ltd)

Acute hyperammonemia resulting from congenital urea cycle disorders, Reye syndrome or acute liver failure results in severe neuronal dysfunction, seizures and death. Increasing evidence suggests that acute hyperammonemia results in alterations of mitochondrial and cellular energy function resulting from ammonia-induced inhibition of the tricarboxylic acid cycle enzyme alpha-ketoglutarate dehydrogenase and by activation of the NMDA receptor. Antagonists of this receptor and NOS inhibitors prevent acute ammonia-induced seizures and mortality and prevent acute ammonia-induced changes in mitochondrial calcium homeostasis and cellular energy metabolism. Acute hyperammonemia also results in decreased activities of free radical scavenging enzymes and again, free radical formation due to ammonia exposure is prevented by either NMDA receptor antagonists or NOS inhibitors. Acute hyperammonemia also results in activation of "peripheral-type" benzodiazepine receptors and monoamine oxidase-B, enzymes which are localized on the mitochondrial membranes of astrocytes in the CNS. Activation of these receptors results in mitochondrial swelling and in increased degradation of monoamines, respectively. Alterations of mitochondrial function could contribute to the neuronal dysfunction characteristic of acute hyperammonemic syndromes.

Studies of animal models suggest that the activation of soluble guanylate cyclase by nitric oxide is altered in liver disease. We studied 77 patients with liver disease and 17 controls, to investigate whether the activation of soluble guanylate cyclase is altered in lymphocytes from patients with liver disease. The basal content of guanosine 3',5'-cyclic monophosphate (cGMP) in lymphocytes was decreased both in patients with liver cirrhosis (by 52%) and in patients with chronic hepatitis (by 62%). Activation of soluble guanylate cyclase by nitric oxide was higher in lymphocytes from patients with cirrhosis (3100+/-1000% of basal) or with hepatitis (5200+/-2500% of basal) than in lymphocytes from controls (1200+/-500% of basal). cGMP in plasma was increased in patients with liver disease. Successful (but not unsuccessful) treatment with interferon of patients with hepatitis due to virus C reversed all the above alterations. Altered modulation of soluble guanylate cyclase by nitric oxide in liver disease may play a role in the hemodynamic alterations found in these patients.