EFFECTS OF EHTANOL IN A NEURON-LIKE CELL LINE

GUADAGNOLI T,; ARONNE MP,; REYNALDO M.; EVRARD SG; BRUSCO HA.

Buzios, Brasil

Congreso; I Congress IBRO/LARC of Neurosciences of Latin America, Caribbean y Iberian Peninsula. XXIII Congress of the Sociedad Argentina de Investigación en Neurociencias (SAN).; 2008

IBRO/LARC of Neurosciences of Latin America, Caribbean y Iberian Peninsula.

Ethanol (EtOH) is a diffusible molecule with access to the CNS tissue. It directly acts on neurons. Our previous "in vivo" results showed neuronal changes in EtOH-exposed rats. In this work, we have studied the “in vitro” response of a neuron-like cell type to EtOH exposure. Neuro-2a (an immortalized murine neuroblastoma) is a suitable model to study the effects of EtOH in the morphology and survival of these cells in a short-term bioassay. Cells were plated at 2 x 104 cells/well in 35mm Nunclon™Ä dishes containing 80ìl DMEM/High Glucose supplemented with 2 mM L-glutamine, 1 mM sodium pyruvate, penicillin G and streptomycin 1% and fetal bovine serum to a final concentration of 10% at 37°C in an atmosphere of 95% air and 5% CO2. 24 h after plating the medium was replaced by OPTIMEM, a reduced serum medium, and exposed to 50mM or 75mM EtOH for 72 h. Control were cells incubated in OPTIMEM only. Caspase-3 immunoreactivity (-ir) showed that a 72-hour treatment with 50 mM or 75 mM EtOH caused an increase in the apoptotic cells percentage (enolase neuron specific-ir) (control: 1.044 ± 0.173 vs 50 mM EtOH: 3.48 ± 0.216, P<0.0001 vs 75 mM EtOH: 2.188 ± 0.2703, P=0.0242). 5HT1Air cells were increased (control: 27.09 ± 1.957 vs 50 mM EtOH: 41.88 ± 2.053, P<0.0001 vs 75 mM EtOH: 41.15 ± 2.166, P<0.0001). Binucleated cells increased in EtOH- treated cells (control: 9.983 ± 1.016 vs 50 mM EtOH: 13.75 ± 0.9739, P=0.016 vs 75 mM EtOH: 15.36 ± 1.502, P=0.0091). In vivo previous works showed that EtOH-altered neuronal development could be partially reversed by a 5HT1A agonists treatment. The present results suggest that EtOH strongly affects neuronal survival and that the 5HT1A receptor is overexpressed in EtOH-treated Neuro-2a cells. Further studies are needed to elucidate the mechanisms involved in the functional relationship between EtOH and 5HT1A receptor.