Elevated levels of ammonia in blood and brain result in derangement of cerebral function. Recently, lipid peroxidation and oxidative stress have been implicated in ammonia neurotoxicity. Because ammonia is primarily detoxified in astrocytes, we postulated that pathophysiological concentrations of ammonia might induce free radical formation in these cells. To test this hypothesis, we examined the extent of free radical production in primary cultures of astrocytes that had been preloaded with the fluorescent dye 5- (and 6-)carboxy-2',7'-dichlorodihydrofluorescein diacetate (DCFDA). DCFDA fluorescence was found to be increased in a dose-dependent manner when astrocytes were exposed to 1, 5, and 10 mM NH(4)Cl. This phenomenon was transitory; it peaked at 2.5 min after exposure and declined subsequently. By 2 hr after treatment, DCFDA fluorescence was below control level. Addition of catalase or superoxide dismutase to 5 mM NH(4)Cl-treated astrocytes reduced free radical formation. Pretreatment with 3 mM methionine sulfoximine, an inhibitor of glutamine synthetase, also suppressed free radical formation by 5 mM NH(4)Cl. The results of this study suggest that elevated concentrations of ammonia induce the formation of free radicals in astrocytes and that this process is associated with the synthesis of glutamine. We propose that astrocyte-derived free radicals may be responsible for some of the pathophysiological changes associated with hyperammonemic conditions.

Astrocyte swelling represents the major factor responsible for the brain edema associated with fulminant hepatic failure (FHF). The edema may be of such magnitude as to increase intracranial pressure leading to brain herniation and death. Of the various agents implicated in the generation of astrocyte swelling, ammonia has had the greatest amount of experimental support. This article reviews mechanisms of ammonia neurotoxicity that contribute to astrocyte swelling. These include oxidative stress and the mitochondrial permeability transition (MPT). The involvement of glutamine in the production of cell swelling will be highlighted. Evidence will be provided that glutamine induces oxidative stress as well as the MPT, and that these events are critical in the development of astrocyte swelling in hyperammonemia.

It is generally accepted that astrocyte swelling forms the major anatomic substrate of the edema associated with acute liver failure (ALF) and that ammonia represents a major etiological factor in its causation. The mechanisms leading to such swelling, however, remain elusive. Recent studies have invoked the role of oxidative stress in the mechanism of hepatic encephalopathy (HE), as well as in the brain edema related to ALF. This article summarizes the evidence for oxidative stress as a major pathogenetic factor in HE/ALF and discusses mechanisms that are triggered by oxidative stress, including the induction of the mitochondrial permeability transition (MPT) and activation of signaling kinases. We propose that a cascade of events initiated by ammonia-induced oxidative stress results in cell volume dysregulation leading to cell swelling/brain edema. Blockade of this cascade may provide novel therapies for the brain edema associated with ALF.

Mitochondria, being the principal source of cellular energy, are vital for cell life. Yet, ironically, they are also major mediators of cell death, either by necrosis or apoptosis. One means by which these adverse effects occur is through the mitochondrial permeability transition (mPT) whereby the inner mitochondrial membrane suddenly becomes excessively permeable to ions and other solutes, resulting in a collapse of the inner membrane potential, ultimately leading to energy failure and cell necrosis. The mPT may also bring about the release of various factors known to cause apoptotic cell death. The principal factors leading to the mPT are elevated levels of intracellular Ca 2+ and oxidative stress. Characteristically, the mPT is inhibited by cyclosporin A. This article will briefly discuss the concept of the mPT, its molecular composition, its inducers and regulators, agents that influence its activity and describe the consequences of its induction. Lastly, we will review its potential contribution to acute neurological disorders, including ischemia, trauma, and toxic-metabolic conditions, as well as its role in chronic neurodegenerative conditions such as Alzheimer's disease, Parkinson's disease, Huntington's disease and amyotrophic lateral sclerosis.

Ammonia is a toxin that has been strongly implicated in the pathogenesis of hepatic encephalopathy (HE), and the astrocyte appears to be the principal target of ammonia toxicity. The specific neurochemical mechanisms underlying HE, however, remain elusive. One of the suggested mechanisms for ammonia toxicity is impaired cellular bioenergetics. Because there is evidence that the mitochondrial permeability transition (MPT) is associated with mitochondrial dysfunction, we determined whether the MPT might be involved in the bioenergetic alterations related to ammonia toxicity. Accordingly, we examined the mitochondrial membrane potential (Deltapsi(m)) in cultured astrocytes and neurons using laser-scanning confocal microscopy after loading the cells with the voltage-sensitive dye JC-1. We found that ammonia induced a dissipation of the Deltapsi(m) in a time- and concentration-dependent manner. These findings were supported by flow cytometry using the voltage-sensitive dye tetramethylrhodamine ethyl ester (TMRE). Cyclosporin A, a specific inhibitor of the MPT, completely blocked the ammonia-induced dissipation of the Deltapsi(m). We also found an increase in the mitochondrial permeability to 2-deoxyglucose in astrocytes that had been exposed to 5 mM NH(4)Cl, further supporting the concept that ammonia induces the MPT in these cells. Pretreatment with methionine sulfoximine, an inhibitor of glutamine synthetase, blocked the ammonia-induced collapse of Deltapsi(m), suggesting a role of glutamine in this process. Over a 24-hr period, ammonia had no effect on the Deltapsi(m) in cultured neurons. Collectively, our data indicate that ammonia induces the MPT in cultured astrocytes, which may be a factor in the mitochondrial dysfunction associated with HE and other hyperammonemic states.