Protective Effect of MK-801 and Diphenyl Diselenide against Methylmercury-Induced Free Radical Generation in Brain Slices of Rats
Puntel, G. O. 1; Roos, D. H. 1; Santos, M. M. 1; Puntel, R. L. 1; Zeni, G. 1; Aschner, M. 2; Rocha, J. B. T. 1;
1Depto. Química- CCNE, Laboratório de Bioquímica Toxicológica, UFSM, SANTA MARIA, RS.
2Vanderbilt Univ. Sch. Med. Dept. Pediat. B-3307 Med. Ctr. N., Nashville, TN 37232, USA.
A number of mechanisms and molecular targets have been proposed to be involved in methylmercury (MeHg)-induced neurotoxicity. Furthermore, several studies have implicated formation of reactive oxygen species (ROS) and disruption of mitochondrial function as two key mechanisms in MeHg-induced neuronal damage (Neurochem. Intern. 2000: 37, 199-206). MK-801 is an NMDA receptor antagonist which can protect neurons from excitotoxicity resulting from overstimulation of N-methyl-D-aspartate (NMDA) receptors. However, at high concentrations MK-801 is known to be extremely neurotoxic (PNAS 2000: 97, 12885–12890). Diphenyl diselenide has been suggested to possess antioxidant properties because it exhibits glutathione (GSH) peroxidase-like activity (Chem Rev. 2004: 104, 6255-6285). The aim of this study was to investigate whether diphenyl diselenide and/or MK-801 could protect brain slices from MeHg-induced free radical generation.
Slices from cerebral cortex of rats were incubated with MeHg (50-200 mM), buthionine sulfoximine (BSO: 25-100 mM), an inhibitor of g–glutamylcysteine synthetase, and/or MK-801 (25-500 mM) and free radical production was determined spectrofluorometrically, using 2’, 7’-dichlorodihydrofluorescin diacetate (DCFA).
At lower concentration (50 mM), MeHg did not modify DCFA oxidation (p>0.05). However, at 100 and 200 mM MeHg it caused an increase in DCFA oxidation when compared to control (p<0.05). BSO did not increase DCFA oxidation compared to controls (p>0.05). MK-801 at 50 mM caused a significant decrease (p<0.05), whereas at 300-500 mM caused a significant increase in DCFA oxidation (p<0.05). The effect of MeHg (200 mM) was completely abolished by MK-801 (50 mM) or diphenyl diselenide (5 mM).
Taken together these results support the potential antioxidant properties of diphenyl diselenide and its ability to effectively attenuate MeHg-induced free radical generation, as well as the involvement of NMDA receptors in MeHg-induced neurotoxicity.
Supported by FAPERGS, CNPq, CAPES
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