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/EBPβ and C/EBPδ but not C/EBPα are expressed in the nuclei of astrocytes. After exposure to glutamate, C/EBPβ 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/EBPδ 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/EBPβ and C/EBPδ, respectively. Elevation of C/EBPβ 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/EBPβ in 32p-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/EEPβ and C/EBPδ as well as through phosphorylation of C/EBPβ.
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