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. 1985 Feb 1;100(2):384–396. doi: 10.1083/jcb.100.2.384

Neuronal regulation of astroglial morphology and proliferation in vitro

PMCID: PMC2113456  PMID: 3881455

Abstract

To analyze the interdependence of neurons and astroglia during central nervous system development, a rapid method for purifying early postnatal cerebellar neurons and astroglia, and recombining them in vitro, has been developed. The influence of neurons on astroglial shape and proliferation has been evaluated with an in vitro model system previously used to describe the role of cerebellar astroglia in neuronal migration and positioning (Hatten, M. E., and R. K. H. Liem, 1981, J. Cell Biol., 90:622-630; and Hatten, M. E., R. K. H. Liem, and C. A. Mason, 1984, J. Cell Biol., 98:193-204. Cerebellar tissue harvested from C57Bl/6J mouse cerebellum on the third or fourth day postnatal was dissociated into a single cell suspension with trypsin, and enriched glial and neuronal fractions were separated with a step gradient of Percoll. Highly purified astroglial and neuronal fractions resulted from subsequently preplanting the cells on a polylysine-coated culture surface. In the absence of neurons, astroglia, identified by staining with antisera raised against purified glial filament protein, assumed a flattened shape and proliferated rapidly. In the absence of astroglia, cerebellar neurons, identified by staining with antisera raised against the nerve growth factor-inducible large external (NILE) glycoprotein and by electron microscopy, formed cellular reaggregates, had markedly impaired neurite outgrowth, and survived poorly. When purified neurons and isolated astroglia were recombined, astroglial proliferation slowed markedly and the flattened shape expressed in the absence of neurons transformed into highly elongated profiles that resembled embryonic forms of cerebellar astroglia. After longer periods (48-72 h) in the presence of neurons, astroglia had "Bergmann-like" or "astrocyte-like" shapes and neurons commonly associated with them. These results suggest that neurons influence the differentiation of astroglia.

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Selected References

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  1. Barkley D. S., Rakic L. L., Chaffee J. K., Wong D. L. Cell separation by velocity sedimentation of postnatal mouse cerebellum. J Cell Physiol. 1973 Apr;81(2):271–279. doi: 10.1002/jcp.1040810215. [DOI] [PubMed] [Google Scholar]
  2. Bignami A., Dahl D. Astrocyte-specific protein and neuroglial differentiation. An immunofluorescence study with antibodies to the glial fibrillary acidic protein. J Comp Neurol. 1974 Jan 1;153(1):27–38. doi: 10.1002/cne.901530104. [DOI] [PubMed] [Google Scholar]
  3. Booher J., Sensenbrenner M. Growth and cultivation of dissociated neurons and glial cells from embryonic chick, rat and human brain in flask cultures. Neurobiology. 1972;2(3):97–105. [PubMed] [Google Scholar]
  4. Bovolenta P., Liem R. K., Mason C. A. Development of cerebellar astroglia: transitions in form and cytoskeletal content. Dev Biol. 1984 Mar;102(1):248–259. doi: 10.1016/0012-1606(84)90189-1. [DOI] [PubMed] [Google Scholar]
  5. Caddy K. W., Biscoe T. J. Structural and quantitative studies on the normal C3H and Lurcher mutant mouse. Philos Trans R Soc Lond B Biol Sci. 1979 Oct 11;287(1020):167–201. doi: 10.1098/rstb.1979.0055. [DOI] [PubMed] [Google Scholar]
  6. Cohen J., Woodhams P. L., Balázs R. Preparation of viable astrocytes from the developing cerebellum. Brain Res. 1979 Feb 9;161(3):503–514. doi: 10.1016/0006-8993(79)90679-6. [DOI] [PubMed] [Google Scholar]
  7. Del Cerro M., Swarz J. R. Prenatal development of Bergmann glial fibres in rodent cerebellum. J Neurocytol. 1976 Dec;5(6):669–676. doi: 10.1007/BF01181580. [DOI] [PubMed] [Google Scholar]
  8. Denis-Donini S., Glowinski J., Prochiantz A. Glial heterogeneity may define the three-dimensional shape of mouse mesencephalic dopaminergic neurones. Nature. 1984 Feb 16;307(5952):641–643. doi: 10.1038/307641a0. [DOI] [PubMed] [Google Scholar]
  9. Fujita S. Quantitative analysis of cell proliferation and differentiation in the cortex of the postnatal mouse cerebellum. J Cell Biol. 1967 Feb;32(2):277–287. doi: 10.1083/jcb.32.2.277. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Furie M. B., Rifkin D. B. Proteolytically derived fragments of human plasma fibronectin and their localization within the intact molecule. J Biol Chem. 1980 Apr 10;255(7):3134–3140. [PubMed] [Google Scholar]
  11. Hatten M. E. Embryonic cerebellar astroglia in vitro. Brain Res. 1984 Apr;315(2):309–313. doi: 10.1016/0165-3806(84)90166-4. [DOI] [PubMed] [Google Scholar]
  12. Hatten M. E., Francois A. M. Adhesive specificity of developing cerebellar cells on lectin substrata. Dev Biol. 1981 Oct 15;87(1):102–113. doi: 10.1016/0012-1606(81)90064-6. [DOI] [PubMed] [Google Scholar]
  13. Hatten M. E., Furie M. B., Rifkin D. B. Binding of developing mouse cerebellar cells to fibronectin: a possible mechanism for the formation of the external granular layer. J Neurosci. 1982 Sep;2(9):1195–1206. doi: 10.1523/JNEUROSCI.02-09-01195.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hatten M. E., Liem R. K. Astroglial cells provide a template for the positioning of developing cerebellar neurons in vitro. J Cell Biol. 1981 Sep;90(3):622–630. doi: 10.1083/jcb.90.3.622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hatten M. E., Liem R. K., Mason C. A. Defects in specific associations between astroglia and neurons occur in microcultures of weaver mouse cerebellar cells. J Neurosci. 1984 Apr;4(4):1163–1172. doi: 10.1523/JNEUROSCI.04-04-01163.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hatten M. E., Liem R. K., Mason C. A. Two forms of cerebellar glial cells interact differently with neurons in vitro. J Cell Biol. 1984 Jan;98(1):193–204. doi: 10.1083/jcb.98.1.193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hatten M. E., Sidman R. L. Cell reassociation behavior and lectin-induced agglutination of embryonic mouse cells from different brain regions. Exp Cell Res. 1978 Apr;113(1):111–125. doi: 10.1016/0014-4827(78)90092-7. [DOI] [PubMed] [Google Scholar]
  18. Liem R. Simultaneous separation and purification of neurofilament and glial filament proteins from brain. J Neurochem. 1982 Jan;38(1):142–150. doi: 10.1111/j.1471-4159.1982.tb10865.x. [DOI] [PubMed] [Google Scholar]
  19. Lim R. Glia maturation factor. Curr Top Dev Biol. 1980;16:305–322. [PubMed] [Google Scholar]
  20. MIALE I. L., SIDMAN R. L. An autoradiographic analysis of histogenesis in the mouse cerebellum. Exp Neurol. 1961 Oct;4:277–296. doi: 10.1016/0014-4886(61)90055-3. [DOI] [PubMed] [Google Scholar]
  21. McCarthy K. D., de Vellis J. Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue. J Cell Biol. 1980 Jun;85(3):890–902. doi: 10.1083/jcb.85.3.890. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Morrison R. S., de Vellis J. Growth of purified astrocytes in a chemically defined medium. Proc Natl Acad Sci U S A. 1981 Nov;78(11):7205–7209. doi: 10.1073/pnas.78.11.7205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mudge A. W. Schwann cells induce morphological transformation of sensory neurones in vitro. Nature. 1984 May 24;309(5966):367–369. doi: 10.1038/309367a0. [DOI] [PubMed] [Google Scholar]
  24. Pettmann B., Sensenbrenner M., Labourdette G. Isolation of a glial maturation factor from beef brain. FEBS Lett. 1980 Sep 8;118(2):195–199. doi: 10.1016/0014-5793(80)80217-1. [DOI] [PubMed] [Google Scholar]
  25. Raff M. C., Miller R. H., Noble M. A glial progenitor cell that develops in vitro into an astrocyte or an oligodendrocyte depending on culture medium. Nature. 1983 Jun 2;303(5916):390–396. doi: 10.1038/303390a0. [DOI] [PubMed] [Google Scholar]
  26. Raff M. C., Mirsky R., Fields K. L., Lisak R. P., Dorfman S. H., Silberberg D. H., Gregson N. A., Leibowitz S., Kennedy M. C. Galactocerebroside is a specific cell-surface antigenic marker for oligodendrocytes in culture. Nature. 1978 Aug 24;274(5673):813–816. [PubMed] [Google Scholar]
  27. Rakic P. Neuron-glia relationship during granule cell migration in developing cerebellar cortex. A Golgi and electronmicroscopic study in Macacus Rhesus. J Comp Neurol. 1971 Mar;141(3):283–312. doi: 10.1002/cne.901410303. [DOI] [PubMed] [Google Scholar]
  28. Rakic P., Sidman R. L. Sequence of developmental abnormalities leading to granule cell deficit in cerebellar cortex of weaver mutant mice. J Comp Neurol. 1973 Nov 15;152(2):103–132. doi: 10.1002/cne.901520202. [DOI] [PubMed] [Google Scholar]
  29. Ranscht B., Clapshaw P. A., Price J., Noble M., Seifert W. Development of oligodendrocytes and Schwann cells studied with a monoclonal antibody against galactocerebroside. Proc Natl Acad Sci U S A. 1982 Apr;79(8):2709–2713. doi: 10.1073/pnas.79.8.2709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Salton S. R., Richter-Landsberg C., Greene L. A., Shelanski M. L. Nerve growth factor-inducible large external (NILE) glycoprotein: studies of a central and peripheral neuronal marker. J Neurosci. 1983 Mar;3(3):441–454. doi: 10.1523/JNEUROSCI.03-03-00441.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Salzer J. L., Bunge R. P. Studies of Schwann cell proliferation. I. An analysis in tissue culture of proliferation during development, Wallerian degeneration, and direct injury. J Cell Biol. 1980 Mar;84(3):739–752. doi: 10.1083/jcb.84.3.739. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Salzer J. L., Williams A. K., Glaser L., Bunge R. P. Studies of Schwann cell proliferation. II. Characterization of the stimulation and specificity of the response to a neurite membrane fraction. J Cell Biol. 1980 Mar;84(3):753–766. doi: 10.1083/jcb.84.3.753. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sensenbrenner M., Devilliers G., Bock E., Porte A. Biochemical and ultrastructural studies of cultured rat astroglial cells: effect of brain extract and dibutyryl cyclic AMP on glial fibrillary acidic protein and glial filaments. Differentiation. 1980;17(1):51–61. doi: 10.1111/j.1432-0436.1980.tb01081.x. [DOI] [PubMed] [Google Scholar]
  34. Sidman R. L., Rakic P. Neuronal migration, with special reference to developing human brain: a review. Brain Res. 1973 Nov 9;62(1):1–35. doi: 10.1016/0006-8993(73)90617-3. [DOI] [PubMed] [Google Scholar]
  35. Sommer I., Schachner M. Monoclonal antibodies (O1 to O4) to oligodendrocyte cell surfaces: an immunocytological study in the central nervous system. Dev Biol. 1981 Apr 30;83(2):311–327. doi: 10.1016/0012-1606(81)90477-2. [DOI] [PubMed] [Google Scholar]
  36. Swarz J. R., Oster-Granite M. L. Presence of radial glia in foetal mouse cerebellum. J Neurocytol. 1978 Jun;7(3):301–312. doi: 10.1007/BF01176995. [DOI] [PubMed] [Google Scholar]
  37. Trenkner E., Hatten M. E., Sidman R. L. Effect of ether-soluble serum components in vitro on the behavior of immature cerebellar cells in weaver mutant mice. Neuroscience. 1978;3(11):1093–1100. doi: 10.1016/0306-4522(78)90127-6. [DOI] [PubMed] [Google Scholar]
  38. Trenkner E., Sidman R. L. Histogenesis of mouse cerebellum in microwell cultures. Cell reaggregation and migration, fiber and synapse formation. J Cell Biol. 1977 Dec;75(3):915–940. doi: 10.1083/jcb.75.3.915. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Trimmer P. A., Reier P. J., Oh T. H., Eng L. F. An ultrastructural and immunocytochemical study of astrocytic differentiation in vitro: changes in the composition and distribution of the cellular cytoskeleton. J Neuroimmunol. 1982 Jun;2(3-4):235–260. doi: 10.1016/0165-5728(82)90058-3. [DOI] [PubMed] [Google Scholar]
  40. Willinger M., Margolis D. M., Sidman R. L. Neuronal differentiation in cultures of weaver (wv) mutant mouse cerebellum. J Supramol Struct Cell Biochem. 1981;17(1):79–86. doi: 10.1002/jsscb.380170109. [DOI] [PubMed] [Google Scholar]
  41. Wood P. M., Bunge R. P. Evidence that sensory axons are mitogenic for Schwann cells. Nature. 1975 Aug 21;256(5519):662–664. doi: 10.1038/256662a0. [DOI] [PubMed] [Google Scholar]

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