Ammonia induces the mitochondrial permeability transition in primary cultures of rat astrocytes

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 underlyi ng HE, however, remain elusive. One of the suggested mechanisms for ammonia toxicity is impaired cellular bioenergetics. Because there is evidence tha t the mitochondrial permeability transition (MPI is associated with mitocho ndrial dysfunction, we determined whether the MPT might be involved in the bioenergetic alterations related to ammonia toxicity. Accordingly, we exami ned the mitochondrial membrane potential (Atom) in cultured astrocytes and neurons using laser-scanning confocal microscopy after loading the cells wi th the voltage-sensitive dye JC-1. We found that ammonia induced a dissipat ion of the Delta psi (m) in a time- and concentration dependent manner. The se findings were supported by flow cytometry using the voltage-sensitive dy e tetramethylrhodamine ethyl ester (TMRE). Cyclosporin A, a specific inhibi tor of the MPT, completely blocked the ammonia-induced dissipation of the A om. We also found an increase in the mitochondrial permeability to 2-deoxyg lucose in astrocytes that had been exposed to 5 mM NH4Cl, further supportin g the concept that ammonia induces the MPT in these cells. Pretreatment wit h methionine sulfoximine, an inhibitor of glutamine synthetase, blocked the ammonia-induced collapse of Delta psi (m), suggesting a role of glutamine in this process. Over a 24-hr period, ammonia had no effect on the Delta ps i (m) in cultured neurons. Collectively, our data indicate that ammonia ind uces the MPT in cultured astrocytes, which may be a factor in the mitochond rial dysfunction associated with HE and other hyperammonemic states. (C) 20 01 Wiley-Liss, Inc.