Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue

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Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue

KD McCarthy and J de Vellis

A novel method has been developed for the preparation of nearly pure separate cultures of astrocytes and oligodendrocytes. The method is based on (a) the absence of viable neurons in cultures prepared from postnatal rat cerebra, (b) the stratification of astrocytes and oligodendrocytes in culture, and (c) the selective detachment of the overlying oligodendrocytes when exposed to sheer forces generated by shaking the cultures on an orbital shaker for 15--18 h at 37 degrees C. Preparations appear greater than 98% pure and contain approximately 1-2 x 10(7) viable cells (20--40 mg of cell protein). Three methods were used to characterize these two culture t ypes. First, electron microscopic examination was used to identify the cells in each preparation (mixed and separated cultures of astrocytes and oligodendrocytes) and to assess the purity of each preparation. Second, two oligodendroglial cell markers, 2',3'-cyclic nucleotide 3'- phosphohydrolase (EC 3.1.4.37) and glycerol phosphate dehydrogenase (EC 1.1.1.8) were monitored. Third, the regulation of cyclic AMP accumulation in each culture type was examined. In addition to these studies, we examined the influence of brain extract and dibutyryl cAMP on the gross morphology and ultrastructure of each preparation. These agents induced astroglial process formation without any apparent morphological effect on oligodendrocytes. Collectively, the results indicate that essentially pure cultures of astrocytes and of oligodendrocytes can be prepared and maintained. These preparations should significantly aid in efforts to examine the biochemistry, physiology, and pharmacology of these two major classes of central nervous system cells.
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Boos, L., Campbell, I. L., Ames, R., Wetsel, R. A., Barnum, S. R. (2004). Deletion of the Complement Anaphylatoxin C3a Receptor Attenuates, Whereas Ectopic Expression of C3a in the Brain Exacerbates, Experimental Autoimmune Encephalomyelitis. J. Immunol. 173: 4708-4714 [Abstract] [Full Text]
Zhou, D., Xue, J., Gavrialov, O., Haddad, G. G. (2004). Na+/H+ exchanger 1 deficiency alters gene expression in mouse brain. Physiol. Genomics 18: 331-339 [Abstract] [Full Text]
Chaban, V. V., Lakhter, A. J., Micevych, P. (2004). A Membrane Estrogen Receptor Mediates Intracellular Calcium Release in Astrocytes. Endocrinology 145: 3788-3795 [Abstract] [Full Text]
Srinivasan, D., Yen, J.-H., Joseph, D. J., Friedman, W. (2004). Cell Type-Specific Interleukin-1{beta} Signaling in the CNS. J. Neurosci. 24: 6482-6488 [Abstract] [Full Text]
Connors, N. C., Adams, M. E., Froehner, S. C., Kofuji, P. (2004). The Potassium Channel Kir4.1 Associates with the Dystrophin-Glycoprotein Complex via {alpha}-Syntrophin in Glia. J. Biol. Chem. 279: 28387-28392 [Abstract] [Full Text]
Liuzzi, G. M., Latronico, T., Fasano, A., Carlone, G., Riccio, P. (2004). Interferon-beta inhibits the expression of metalloproteinases in rat glial cell cultures: implications for multiple sclerosis pathogenesis and treatment. Mult Scler 10: 290-297 [Abstract]
Bachoo, R. M., Kim, R. S., Ligon, K. L., Maher, E. A., Brennan, C., Billings, N., Chan, S., Li, C., Rowitch, D. H., Wong, W. H., DePinho, R. A. (2004). Molecular diversity of astrocytes with implications for neurological disorders. Proc. Natl. Acad. Sci. USA 101: 8384-8389 [Abstract] [Full Text]
Decker, L., ffrench-Constant, C. (2004). Lipid Rafts and Integrin Activation Regulate Oligodendrocyte Survival. J. Neurosci. 24: 3816-3825 [Abstract] [Full Text]
Denkins, Y., Reiland, J., Roy, M., Sinnappah-Kang, N. D., Galjour, J., Murry, B. P., Blust, J., Aucoin, R., Marchetti, D. (2004). Brain metastases in melanoma: Roles of neurotrophins. Neuro Oncol Duke 6: 154-165 [Abstract]
De Pietri Tonelli, D., Mihailovich, M., Di Cesare, A., Codazzi, F., Grohovaz, F., Zacchetti, D. (2004). Translational regulation of BACE-1 expression in neuronal and non-neuronal cells. Nucleic Acids Res 32: 1808-1817 [Abstract] [Full Text]
Montana, V., Ni, Y., Sunjara, V., Hua, X., Parpura, V. (2004). Vesicular Glutamate Transporter-Dependent Glutamate Release from Astrocytes. J. Neurosci. 24: 2633-2642 [Abstract] [Full Text]
Perez-Ortiz, J. M., Tranque, P., Vaquero, C. F., Domingo, B., Molina, F., Calvo, S., Jordan, J., Cena, V., Llopis, J. (2004). Glitazones Differentially Regulate Primary Astrocyte and Glioma Cell Survival: INVOLVEMENT OF REACTIVE OXYGEN SPECIES AND PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR-{gamma}. J. Biol. Chem. 279: 8976-8985 [Abstract] [Full Text]
Baud, O., Greene, A. E., Li, J., Wang, H., Volpe, J. J., Rosenberg, P. A. (2004). Glutathione Peroxidase-Catalase Cooperativity Is Required for Resistance to Hydrogen Peroxide by Mature Rat Oligodendrocytes. J. Neurosci. 24: 1531-1540 [Abstract] [Full Text]
Romera, C., Hurtado, O., Botella, S. H., Lizasoain, I., Cardenas, A., Fernandez-Tome, P., Leza, J. C., Lorenzo, P., Moro, M. A. (2004). In Vitro Ischemic Tolerance Involves Upregulation of Glutamate Transport Partly Mediated by the TACE/ADAM17-Tumor Necrosis Factor-{alpha} Pathway. J. Neurosci. 24: 1350-1357 [Abstract] [Full Text]