No abstract

Redox-induced regulation of the Na-K-ATPase was studied in dispersed rat cerebellar granule cells. Intracellular thiol redox state was modulated using glutathione (GSH)-conjugating agents and membrane-permeable ethyl ester of GSH (et-GSH) and Na-K-ATPase transport and hydrolytic activity monitored as a function of intracellular reduced thiol concentration. Depletion of cytosolic and mitochondrial GSH pools caused an increase in free radical production in mitochondria and rapid ATP deprivation with a subsequent decrease in transport but not hydrolytic activity of the Na-K-ATPase. Selective conjugation of cytosolic GSH did not affect free radical production and Na-K-ATPase function. Unexpectedly, overloading of cerebellar granule cells with GSH triggered global free radical burst originating most probably from GSH autooxidation. The latter was not followed by ATP depletion but resulted in suppression of active K(+) influx and a modest increase in mortality. Suppression of transport activity of the Na-K-ATPase was observed in granule cells exposed to both permeable et-GSH and impermeable GSH, with inhibitory effects of external and cytosolic GSH being additive. The obtained data indicate that redox state is a potent regulator of the Na-K-ATPase function. Shifts from an "optimal redox potential range" to higher or lower levels cause suppression of the Na-K pump activity.

Thiol containing NO.-derivatives were found to inhibit the activity of brain and kidney Na/K-ATPase. S-Nitrosogluthatione demonstrated only minor inhibiting activity, while dinitrosyl iron complexes (DNIC) with cysteine or glutathione were much more effective. Brain Na/K-ATPase is more vulnerable to inhibiting action than kidney Na/K-ATPase. Inhibition of the activity is accompanied by a decrease in amount of protein thiol groups and a change in the substrate dependence curve of the enzyme. Restoration of Na/K-ATPase activity by SH-reagent dithiothreitol or cysteine is accompanied by restoration of SH-groups of the enzyme. This suggests that blockade of SH-groups of Na/K-ATPase is responsible for the inhibition. The possibility that this blockade results in disordering of interprotomer interactions within the oligomeric complexes of Na/K-ATPase is suggested. Possible regulatory meaning of the effect of NO. derivatives is discussed.

Multiple lines of evidence demonstrate that reactive oxygen species (ROS) are involved in regulation of normal cell metabolism as second messengers. Under extreme conditions, these molecules induce oxidative stress, which may stimulate (or accompany) a number of neurodegenerative processes. In the glutamatergic system, ROS levels are under control of ionotropic and metabotropic glutamate receptors, which modulate ion fluxes through the neuronal membrane. The Na+/K(+)-pump is also one of the important participants affecting stationary ROS levels through several distinct mechanisms. This review describes the involvement of the Na+/K(+)-pump in intracellular signaling mechanisms via cross-talk between the pump and glutamate receptors in cerebellum granule cells. Selective dysfunction of mGlu II receptors may also lead to abnormal protein phosphorylation (i.e., tau phosphorylation), culminating in neurodegenerative disorders (i.e., Alzheimer disease). Also, unregulated production of intracellular ROS resulting from an imbalance of ionotropic and metabotropic receptors may activate one or more protein kinases. In summary, Glu receptor dysfunction, leading to a deficit in glutamate-mediated signal transduction may represent one of the earliest stages of neurodegenerative disorders. The Na+/K(+)-pump is able to prevent over-production of intracellular ROS, thus increasing oxidative stability of neuronal cells.

We found that methionine added to the ration of pregnant rats (1 g/kg body weight) induced sustained hyperhomocysteinemia and led to the formation of sustained oxidative stress in the brain of their progeny. Newborn animals were characterized by lower body weight, SOD deficiency in the brain, increased neuronal death, and desensitization of NMDA receptors. These factors are associated with impaired cognitive capacity in the Morris test.