Resumo

Objectives: Hyperprolinemia type II is a metabolic disease caused by deficiency of Δ1-pyrroline-5-carboxylic acid dehydrogenase activity characterized by tissue accumulation of proline. Affected patients usually present neurological manifestations including seizures and mental retardation. Na+, K+-ATPase is a membrane-bound enzyme that plays an essential role in controlling neuronal excitability by maintaining electrochemical gradients, through active transport of Na+ and K+ ions across the cell membrane, at the expense of ATP. Considering that chronic hyperprolinemia decreases the activity of this enzyme in brain synaptic plasma membranes, the objectives of the present study were to investigate the effect of chronic proline administration on thiobarbituric acid-reactive substances (TBARS), as well as the influence of antioxidant vitamins E plus C on the effects mediated by proline on Na+, K+-ATPase activity in cerebral cortex of rats. The expression of Na+, K+-ATPase catalytic subunits was also evaluated. Materials and Methods: Wistar rats were chronically treated with proline, administered subcutaneously twice a day with 8 h-interval, from the 6th to the 28th day of age. Concomitantly, vitamin E (40 mg/kg) plus C (100 mg/kg) were injected intraperitoneally once a day. Rats were sacrificed 12 h after the last injection and the cerebral cortex was dissected. Synaptic plasma membrane was prepared and Na+, K+-ATPase activity assay were performed. TBARS levels, an index of lipid peroxidation, were also measured. The analysis of Na+, K+-ATPase catalytic subunits expression were carried out by a semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR) assay. Results: Results showed that proline increased TBARS, suggesting an increase of lipid peroxidation (Sal: 1.37±0.11; Pro: 1.62±0.22; n=7). Besides, concomitant administration of vitamins E plus C significantly prevented the inhibition of Na+, K+-ATPase activity caused by proline, suggesting the involvement of oxidative stress in such effect (Sal: 1661±117; Pro: 1134±126; Vit: 1756±142; Pro + Vit: 1393±87; n=4-5). We did not observe any change in levels of Na+, K+-ATPase mRNA transcripts after chronic exposure to proline and vitamins E plus C (Sal: 0.97±0.03; Pro: 0.94±0.04; Vit: 0.98±0.04; Pro + Vit: 0.95±0.02; n=4-5). Conclusion: Our results suggest that inhibition of Na+, K+-ATPase activity promoted by proline, at least in part, is mediated by oxidative stress, since concomitant administration of vitamins E plus C significantly prevented this effect. However, the inhibition of enzyme activity did not involve alterations in Na+, K+-ATPase mRNA transcripts. These findings provide insights into the mechanisms by which proline exerts its effects on brain function and suggest the possibility that treatment with antioxidants may be beneficial in neurological dysfunctions observed in hyperprolinemic patients.