Isolation and characterization of a new immortal rat astrocyte with a high expression of NGF mRNA

• Published in: Neuroscience research Vol. 39; no. 2; pp. 205 - 212
• Main Authors: Morikawa, Masayuki, Asai, Kiyofumi, Kokubo, Minoru, Fujita, Kaori, Yoneda, Kazuhiro, Yamamoto, Naoki, Inoue, Yuichiro, Iida, Junzo, Kishimoto, Toshifumi, Kato, Taiji
• Format: Journal Article
• Language: English
• Published: Ireland Elsevier Ireland Ltd 01-02-2001
• Subjects: Animals; Astrocyte; Astrocytes - cytology; Astrocytes - metabolism; Brain - cytology; Brain - metabolism; Brain-Derived Neurotrophic Factor - genetics; Cell Culture Techniques - methods; Cells, Cultured; Embryo, Mammalian; Glial fibrillary acidic protein; Immortalization; Karyotyping; Nerve growth factor; Nerve Growth Factor - genetics; Neurotrophin 3 - genetics; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; SV40 T antigen; Transcription, Genetic; SV40 T antigen; Nerve growth factor; Astrocyte; Immortalization; Glial fibrillary acidic protein
• ISSN: 0168-0102; 1872-8111
• DOI: 10.1016/S0168-0102(00)00217-0

We have established a new line of immortalized rat astrocytes through transfection of plasmid pSV3-neo encoding the large T antigen of simian virus 40 into normal astrocytes. One of these immortalized astrocytes (ACT-57) with a flat and polygonal cell shape, exhibited stable growth in a chemically defined medium (modified N-2 medium) as well as in medium containing ordinary serum. ACT-57, retained a detectable level of expression of glial fibrillary acidic protein (GFAP) and its mRNA, and exhibited a stronger expression of nerve growth factor (NGF) mRNA than that of normal rat astrocytes or C6 glioma cells. NGF mRNA was significantly up-regulated by phorbol ester (12-
O-tetradecanoylphorbol 13-acetate, TPA) and γ-amino-
n-butyric acid (GABA) but not by hydrocortisone. None of stimulants (TPA, dibutyryl cyclic AMP (db-cAMP), hydrocortisone,
l-glutamate, carbacol, GABA, dopamine, or isoproterenol) changed the expression level of either brain-derived neurotrophic factor (BDNF) or neurotrophin-3 (NT-3). There was a discrete difference between ACT-57 and normal astrocytes in the response to GABA and isoproterenol. These findings imply that normal cortical astrocytes possess a functional heterogeneity whereas the clonal astrocyte, ACT-57, does not, indicating that ACT-57 cells may be useful for in vitro studies of neuron-astrocyte interactions involving the induction of neurotrophic factors such as NGF.