NEUROTOXICITY OF ETHANOL IN RAT CORTICAL NEURONS: AN ULTRASTRUCTURAL STUDY

GUADAGNOLI, T; ARONNE, MP; LOPEZ, LM; GIRONACCI, M; BRUSCO, A

Florencia

Congreso; IBRO2011; 2011

IBRO

Previously, we have demonstrated alterations in the neuronal cytoskeleton in cerebral cortex of ratsprenatally exposed to ethanol. We also showed in rat primary cortical neurons that ethanol inducedneuronal death and a decrease in the elongation and branching of dendrites.The aim of the present work was to evaluate ultrastructural changes in rat cortical neurons exposed To 50mM of ethanol during 24 hours.Primary cortical neuronal cultures were obtained from one-day-old Wistar rats. Cells were plated at 3.5 x 106 cells/35 mm dish and grown in Neurobasal supplemented with B-27 and 2 mM L-glutamine until complete differentiation. 12 h before treatment, cells were B-27 deprived, and exposed to 50 mMethanol during 24 h. After that, cells were fixed with 2 % glutaraldehyde + 1% formaldehyde in 0,1Mcacodylate buffer for 60 min. After washing in this buffer, cells were postfixed in 2 % OsO4 containing 1 % potassium ferrocyanide for 60 min., treated with 0.1 % tannic acid for 1 min, washed and stained in block with 2 % aqueous uranyl acetate for 120 min , and embedded in Eppon 812 Ultrathin sections. Images were taken in a Zeiss electron microscopy at 7000X, 30000X and 50000X.Cortical neurons exposed to ethanol showed indented nucleus, increased number of lipid droplets,mitochondria with disrupted cristae and degenerating bodies (multilamellar vesicles), dilatedendoplasmic reticulum lumen and a disorganized Golgi apparatus with a ring-shape appearance.Heterogeneous distribution with a decreased density of microtubules (MT) in neurites was alsoobserved in cortical neurons exposed to ethanol (control: 139.2 ± 32.52 vs. treated: 105.5 ± 31.56 MT/μm2). In addition, neuronal cultures showed immature synapses which were not observed in ethanol-treated neurons.We conclude that ultrastructural changes observed in cortical neurons exposed to ethanol mayexplain some of the neuronal dysfunctions observed ?in vivo?. Cytoskeleton disorganization mayresult in a dendritic processes decrease.