The effect of modulation of Na+/К+-АТPase activity on viability of cerebellar granule cells exposed to oxidative stress in vitro

Introduction. Oxidative stress is an important pathogenic factor in cerebral ischemia, which occupies one of the leading places among various forms of cerebral pathology in mortality and disability of the working-age population and is recognized as an actual problem of experimental and clinical neurology. Naturally, modeling of neurodestructive processes and their correction under the action of oxidative stress in vitro contributes to the study of protective mechanisms that counteract ischemic damage of neurons.

Objective. To reveal the influence of chemical preconditioning induced by transient inhibition of Na⁺/K⁺-ATPase activity on tolerance of cultured cerebellar granule neurons to oxidative stress at different stages of their differentiation in vitro.

Materials and methods. The activity of Na⁺/K⁺-ATPase was inhibited with ouabain, which was added at 3–4 and 7–8 days in vitro to cerebellar cell cultures of 7-day rats at a concentration of 0.1 mM for 24 hours before induction of oxidative stress by hydrogen peroxide (0.05 and 0.075 mM, 4 hours) or paraquat (0.15 and 0.2 mM, 24 hours).

Results. Oxidative stress induced by paraquat causes the most pronounced death of cultured granular neurons in immature (3–4 days) cultures, in which survival was 44±2,5% of neurons, compared to mature (7–8 days) cultures, in which survival was 61±5,4%. Pretreatment of cultures with ouabain has a protective effect, the most significant in mature cultures. The exposure of mature cultures with hydrogen peroxide kills more than 90% of neurons, whereas pretreatment with ouabain increases the survival rate by 44%. At the same time in the immature cultures the damaging effects of H₂O₂ and the protective effect of ouabain is less pronounced.

Conclusion. The increased tolerance of cultured cerebellar granule cells to oxidative stress after transient inhibition of Na⁺/K⁺-ATPase activity by ouabain is shown. The direct dependence of the efficiency of the ouabain protection on the degree of neuronal morphochemical differentiation in vitro is revealed.