Ethanol induces Fas/Apo [apoptosis]-1 mRNA and cell suicide in the developing cerebral cortex

Introduction: Animal studies modeling fetal alcohol syndrome have demonstra ted that developmental exposure to alcohol is associated with decreased bra in weight and significant neuronal loss in multiple regions of the developi ng brain. Our previous data suggest that the Fas/Apo [apoptosis]-1 receptor is transiently expressed in the developing cerebral cortex during the peak period of naturally occurring apoptotic cell death and maximum sensitivity to alcohol. Therefore, we hypothesized that ethanol increases the expressi on of suicide receptors such as Fas/Apo-1 in the developing fetal cerebral cortex and leads to an upregulation or extension of the normal period of ap optosis and consequent disorganization of the neural circuitry. Methods: Ethanol was administered in one of four doses (120, 320, 630, and 950 mg/dl) to organotypic explant cultures of the developing cerebral corte x established from postnatal day 2 rats and maintained for 6 days in vitro. The number of cells expressing Fas/Apo-1 receptor mRNA was counted. Apopto sis was measured by the use of two independent assays; a cell death enzyme- linked immunosorbent assay for PNA fragmentation and flow cytometric analys is of Annexin-V binding to phosphatidylserine externalized to the outer lea flet of the plasma membrane. Necrosis was also estimated by two independent measures, the amount of lactate dehydrogenase released into culture medium and now cytometric analysis of cells that were positive for both Annexin-V and propidium iodide. Results: A significantly larger number of developing cortical cells express ed Fas/Apo-1 mRNA at the lower doses (120 and 320 mg/dl) than at the higher doses (630 and 950 mg/dl). Furthermore, ethanol induced apoptosis in a dos e-related manner, with peak apoptosis observed at a dose of 630 mg/dl in th e case of DNA fragmentation and at 630 and 950 mg/dl in the case of phospha tidylserine translocation to the outer leaflet of the plasma membrane. Etha nol did not induce necrosis at any of the administered doses of ethanol. Conclusions: Our data suggest that ethanol induces a susceptibility to apop totic signals at low doses by upregulating the expression of mRNAs for cyto toxic receptors such as Fas/Apo-1 in the developing cerebral cortex. Howeve r, ethanol itself specifically induces apoptosis in the developing cerebral cortex only at higher doses.