Introduction. An important component of the antioxidant defense system is glutathione, which neutralizes lipid peroxides and maintains the reduced state of SH-group proteins, providing their functional activity. Recent studies were connected with the role of glutathione in gene expression, intracellular signaling, regulation of enzyme activity, apoptosis, and other processes.
Purpose. The aim of the study was to examine the performance of the glutathione system under conditions of nitrosative stress and their effect on the mechanisms that trigger neuronal death.
Methods. For in vitro studies, neurons were isolated from the cerebral cortex of 4-week old albino rats. For modeling nitrosative stress in the incubation medium with toxic concentration (250 mcM) it was added dinitrozole iron complex (DNIC). In order to determine the type of neuronal cell death using a morphological technique differential colouring of apoptotic cells with fluorescent dye — Hoechst 33342. Intensity of nitrosative stress in suspension was evaluated by neuronal accumulation of nitrotyrosine. Condition of glutathione level was assessed by its oxidized and reduced form, as well as the activity of key enzymes of its metabolism — glutathione reductase and glutathione-S-transferase (G-S-T).
Results. Introduction to the incubation medium resulted in an increase DNIC aldehyde phenylhydrasones 2.02 times, ketonphenylhydrasones 2.12 times and nitrotyrosine 3.06 times. Also, a shift of thiol-disulfide ratio toward the oxidized thiols, as evidenced by the decrease in glutathione levels restored to 80.5%, which took place against the background of an increase in its oxidized form 3.05 times.
Conclusions. 1. Adding DNIC neurons to the incubation medium resulted in deprivation of glutathione thiol-disulfide-level system, expressed in short supply of reduced glutathione and inhibition of enzyme activity of its metabolism.
2. Adding DNIC caused by the uncontrolled production of reactive oxygen and nitrogen oxide, the development of oxidative and nitrosative stress.
3. The above described pathobiochemical changes caused increased cell death of neurons in the suspension, as evidenced by a statistically significant increase in the number of apoptous-modified neurons colored by Hoechst 33342.