Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1981 Feb;78(2):1283–1287. doi: 10.1073/pnas.78.2.1283

Requirement for nonoligodendrocyte cell signals for enhanced myelinogenic gene expression in long-term cultures of purified rat oligodendrocytes.

S Bhat, E Barbarese, S E Pfeiffer
PMCID: PMC319993  PMID: 6165009

Abstract

Comparisons have been made of myelinogenic activities in fetal rat brain mixed primary cultures and cultures of isolated oligodendrocytes of comparable age. The specific activities of the sulfatide synthesis, 2',3'-cyclic-nucleotide 3'-phosphohydrolase (2',3'-cyclic-nucleotide 3'-phosphodiesterase, EC 3.1.4.37), and accumulation of myelin basic protein, when expressed per mg of protein, were as high (or generally higher) in isolated oligodendrocyte cultures as in comparable mixed primary cultures at 29 days. However, when these data were analyzed per oligodendrocyte, it became apparent that the isolated oligodendrocytes were substantially less active than their mixed culture counterparts. The results suggest the necessity of nonologodendrocyte positive signals for the optimal expression by oligodendrocytes of myelin-related differentiated functions. The isolation method involves the selection of oligodendrocytes by shaking them from primary cultures of rat brain, followed by the lysis of other contaminating cells in a balanced salt solution at pH 7.2. More than 99% of the isolated cells are viable, at least initially divide, and can be cultured for at least 60 days. The oligodendrocytes selected in this way were characterized by: (i) morphology, (ii) immunofluorescence labeling by antibodies to myelin basic protein and galactosylceramide, and (iii) biochemical analyses for myelin basic protein, activity of 2',3'-cyclic-nucleotide 3'-phosphohydrolase, and sulfogalactosylceramide synthesis.

Full text

PDF
1283

Images in this article

1284Image
on p.1284
1284Image
on p.1284
1284Image
on p.1284
1285Image
on p.1285
1285Image
on p.1285
1285Image
on p.1285

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aguayo A. J., Epps J., Charron L., Bray G. M. Multipotentiality of Schwann cells in cross-anastomosed and grafted myelinated and unmyelinated nerves: quantitative microscopy and radioautography. Brain Res. 1976 Mar 5;104(1):1–20. doi: 10.1016/0006-8993(76)90643-0. [DOI] [PubMed] [Google Scholar]
  2. Barbarese E., Braun P. E., Carson J. H. Identification of prelarge and presmall basic proteins in mouse myelin and their structural relationship to large and small basic proteins. Proc Natl Acad Sci U S A. 1977 Aug;74(8):3360–3364. doi: 10.1073/pnas.74.8.3360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Barbarese E., Carson J. H., Braun P. E. Accumulation of the four myelin basic proteins in mouse brain during development. J Neurochem. 1978 Oct;31(4):779–782. doi: 10.1111/j.1471-4159.1978.tb00110.x. [DOI] [PubMed] [Google Scholar]
  4. Barbarese E., Carson J. H., Braun P. E. Subcellular distribution and structural polymorphism of myelin basic protein in normal and Jimpy mouse brain. J Neurochem. 1979 May;32(5):1437–1446. doi: 10.1111/j.1471-4159.1979.tb11082.x. [DOI] [PubMed] [Google Scholar]
  5. Bunge R. P. Glial cells and the central myelin sheath. Physiol Rev. 1968 Jan;48(1):197–251. doi: 10.1152/physrev.1968.48.1.197. [DOI] [PubMed] [Google Scholar]
  6. Dawson G., Kernes S. M. Mechanism of action of hydrocortisone potentiation of sulfogalactosylceramide synthesis in mouse oligodendroglioma clonal cell lines. J Biol Chem. 1979 Jan 10;254(1):163–167. [PubMed] [Google Scholar]
  7. Dawson G., Stoolmiller A. C. Comparison of the ganglioside composition of established mouse neuroblastoma cell strains grown in vivo and in tissue culture. J Neurochem. 1976 Jan;26(1):225–226. doi: 10.1111/j.1471-4159.1976.tb04466.x. [DOI] [PubMed] [Google Scholar]
  8. Fry J. M., Lehrer G. M., Bornstein M. B. Sulfatide synthesis: inhibition by experimental allergic encephalomyelitis serum. Science. 1972 Jan 14;175(4018):192–194. doi: 10.1126/science.175.4018.192. [DOI] [PubMed] [Google Scholar]
  9. Glastris B., Pfeiffer S. E. Mammalian membrane marker enzymes: sensitive assay for 5'-nucleotidase and assay for mammalian 2',3'-cyclic-nucleotide-3'-phosphohydrolase. Methods Enzymol. 1974;32:124–131. doi: 10.1016/0076-6879(74)32015-0. [DOI] [PubMed] [Google Scholar]
  10. Kruger L., Maxwell D. S. Electron microscopy of oligodendrocytes in normal rat cerebrum. Am J Anat. 1966 Mar;118(2):411–435. doi: 10.1002/aja.1001180207. [DOI] [PubMed] [Google Scholar]
  11. Kurihara T., Tsukada Y. The regional and subcellular distribution of 2',3'-cyclic nucleotide 3'-phosphohydrolase in the central nervous system. J Neurochem. 1967 Dec;14(12):1167–1174. doi: 10.1111/j.1471-4159.1967.tb06164.x. [DOI] [PubMed] [Google Scholar]
  12. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  13. 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]
  14. McKhann G. M., Ho W. The in vivo and in vitro synthesis of sulphatides during development. J Neurochem. 1967 Jul;14(7):717–724. doi: 10.1111/j.1471-4159.1967.tb10305.x. [DOI] [PubMed] [Google Scholar]
  15. Mori S., Leblond C. P. Electron microscopic identification of three classes of oligodendrocytes and a preliminary study of their proliferative activity in the corpus callosum of young rats. J Comp Neurol. 1970 May;139(1):1–28. doi: 10.1002/cne.901390102. [DOI] [PubMed] [Google Scholar]
  16. Pfeiffer S. E., Wechsler W. Biochemically differentiated neoplastic clone of Schwann cells. Proc Natl Acad Sci U S A. 1972 Oct;69(10):2885–2889. doi: 10.1073/pnas.69.10.2885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Poser C. M. Dysmyelination revisited. Arch Neurol. 1978 Jul;35(7):401–408. doi: 10.1001/archneur.1978.00500310003001. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Raine C. S., Poduslo S. E., Norton W. T. The ultrastructure of purified preparations of neurons and glial cells. Brain Res. 1971 Mar 19;27(1):11–24. doi: 10.1016/0006-8993(71)90368-4. [DOI] [PubMed] [Google Scholar]
  20. Raine C. S., Traugott U., Iqbal K., Snyder D. S., Cohen S. R., Farooq M., Norton W. T. Encephalitogenic properties of purified preparations of bovine oligodendrocytes tested in guinea pigs. Brain Res. 1978 Feb 17;142(1):85–96. doi: 10.1016/0006-8993(78)90178-6. [DOI] [PubMed] [Google Scholar]
  21. 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]
  22. Shortman K., Williams N., Adams P. The separation of different cell classes from lymphoid organs. V. Simple procedures for the removal of cell debris. Damaged cells and erythroid cells from lymphoid cell suspensions. J Immunol Methods. 1972 May;1(3):273–287. doi: 10.1016/0022-1759(72)90005-1. [DOI] [PubMed] [Google Scholar]
  23. Silberberg D., Benjamins J., Herschkowitz N., McKhann G. M. Incorporation of radioactive sulphate into sulphatide during myelination in cultures of rat cerebellum. J Neurochem. 1972 Jan;19(1):11–18. doi: 10.1111/j.1471-4159.1972.tb01248.x. [DOI] [PubMed] [Google Scholar]
  24. Snyder D. S., Raine C. S., Farooq M., Norton W. T. The bulk isolation of oligodendroglia from whole rat forebrain: a new procedure using physiologic media. J Neurochem. 1980 Jun;34(6):1614–1621. doi: 10.1111/j.1471-4159.1980.tb11252.x. [DOI] [PubMed] [Google Scholar]
  25. Sternberger N. H., Itoyama Y., Kies M. W., Webster H deF Immunocytochemical method to identify basic protein in myelin-forming oligodendrocytes of newborn rat C.N.S. J Neurocytol. 1978 Apr;7(2):251–263. doi: 10.1007/BF01217922. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES