Ethanol-induced changes on ?in vitro? cortical neurons

GUADAGNOLI, TAMARA; ARONNE, MARIA PAULA; LOPEZ, LIDIA M; GIRONACCI, MARIELA; BRUSCO, ALICIA

Uspallata, Mendoza

Congreso; V Neurotoxicity Society Meeting; 2011

Neurotoxicity Society

Deletereous effects of ethanol (EtOH) on cortical neurons has been demonstrated using ?invivo? animal models of fetal EtOH-exposure, but there are not a fully ?in vitro? study. Ourprevious "in vivo" results showed neuronal damage in EtOH-exposed rats. In this work, westudy the ?in vitro? response of primary culture of cortical neurons to EtOH-exposure.Cortical neurons of Wistar rats postnatal day 1 were used for obtain primary cultures. Cells wereplated at 3.5 x 106 cells/well in 35mm Nunclon?A dishes containing Neurobasal supplementedwith B-27 and 2 mM L-glutamine at 37°C (95% air and 5% CO2) during 6 days. 12 h beforetreatment the cell medium was replaced by Neurobasal, and exposed to 25mM, 50mM or100mM EtOH during 24 h. Control cells were incubated with Neurobasal only. Viability wasstudied by MTT assay. Apoptosis and necrosis were analized using a vital fluorescence test,TUNEL staining and ultrastructural analysis. MAP-2 immunostaining was used as a specificneuronal marker.EtOH-exposure produces a decrease in cells viability, increased the apoptotic neuronspercentage and decrease of the MAP-2+ cells. In addition, total dendrite length per cell, primarydendrite length, secondary dendrite length and number of primary and secondary dendrites perneuron were analized. EtOH-exposure produce a reduction in the total dendrite length as inbranching. These results suggested that 24 hs of EtOH-exposure promotes neuronal death aswell as morphological changes in dendrites and axon corroborate by electron microscopy.Grants UBACYT M-004, PIP 0774