Ethanol intake is associated with increase in lipid peroxidation and formation of reactive oxygen species in different cerebral areas, in neurons as well in astrocytes. In this study new born rat hippocampal astrocytes in primary culture were acutely exposed to ethanol. Cells were loaded with fura-2/AM and CM-H2DCFDA for the determination of changes in [Ca²⁺]c and ROS production, respectively. Glutamate secretion was determined by an enzyme-linked system and spectrofluorimetry, and GFAP content was determined by immunocytochemistry and confocal scanning microscopy. Our results show that ethanol (1 mM-200 mM) led to a dose-dependent increase in glutamate secretion. 50 mM ethanol induced release of Ca²⁺ from cytosolic stores and increased ROS production. A reduction in glutamate secretion and ROS production was observed when extracellular Ca²⁺ was omitted and following preincubation of cells in the presence of the intracellular Ca²⁺ chelator BAPTA/AM (10 mM), indicating a Ca²⁺ dependency for these processes. Preincubation of astrocytes in the presence of 10 mM antimycin plus 10 mM oligomycin, to inhibit mitochondria, completely blocked ethanol-evoked ROS production. Finally, incubation of astrocytes in the presence of ethanol increased the content of GFAP, a sign of gliosis. In conclusion, our results suggest that ethanol induces an immediate response through Ca²⁺ mobilization, ROS generation and glutamate secretion and later changes involving GFAP expression, in rat hippocampal astrocytes. These effects may underline various signalling pathways which are important for cell proliferation, differentiation and function, and might represent a situation potentially leading to neurotoxicity in the hippocampus.

Supported by Junta Extremadura-FEDER 2PR04A009.