Regulation of CCAAT/enhancer-binding protein family members by stimulation of glutamate receptors in cultured rat cortical astrocytes

Regulation of mRNA levels, DNA binding activities, and phosphorylation of CCAAT/enhancer-binding protein (C/EBP) family members by stimulation of glutamate receptors were studied in cultured rat cortical astrocytes. Indirect immunofluorescence and immunoblot analyses with specific antibodies to C/EBP family members revealed that both C/EBP beta and C/EBP delta but not C/EBP alpha are expressed in the nuclei of astrocytes. After exposure to glutamate, C/EBP beta mRNA levels increased within 10 min, reached the maximal level at about 1 h, and returned to the basal level within 6 h, In contrast, C/EBP delta mRNA levels decreased by 6 h and were recovered within 12 h, These changes in mRNA levels were accompanied by an increase and a decrease in proteins for C/EBP beta and C/EBP delta, respectively. Elevation of C/EBP beta mRNA levels by glutamate treatment required an increase in intracellular Ca2+ concentration and depended on activations of protein kinase C and calmodulin-dependent protein kinases. Gel mobility shift analysis using nuclear extracts from the glutamate-treated cells showed increases in C/EBP site binding activities 2 h after the exposure to glutamate, Moreover, glutamate stimulated phosphorylation of C/EBP beta in P-32-labeled astrocytes in a Ca2+-dependent manner. These results suggest that glutamate regulates functions of C/EBP family members in brain astrocytes through changes in mRNA levels of C/EBP beta and C/EBP delta as well as through phosphorylation of C/EBP beta.