Variation in content and function of non-neuronal cells in the outgrowth of sympathetic ganglia from embryos of differing age

D. Roufa, M. B. Bunge, M. I. Johnson, C. J. Cornbrooks

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Abstract

Studies on cellular interactions in developing nervous system are greatly facilitated by the availability of tissue culture preparations that contain single or combined populations of neurons and non-neuronal cells (NNCs). Using superior cervical ganglia (SCG) from early E15 rats on air-dried collagen, we were able to prepare (1) cultures containing neurons along with Schwann cells (SCs) as the only NNC type present without the use of antimitotic treatment and (2) cultures containing only neurons, following brief antimitotic treatment. Light-microscopic observation of E15 outgrowth showed a uniform population of flattened cells, unlike that of E20 cultures, which contained a mixture of spindle-shaped and flattened cells. Autoradiograms following [3H]thymidine administration to E15 cultures revealed a striking gradient of nuclear labeling: Only a few cells were labeled near the explant and nearly all cells were labeled at the growth front. This was in marked contrast to E20 cultures, in which nuclei were labeled throughout the outgrowth. The conclusion that the labeling gradient is explained by the presence of SCs without other NNC types in E15 cultures was confirmed by immunocytochemical studies. Anti-laminin antibodies stain only those extracellular matrix components related to the SC surface, whereas anti-fibronectin antibodies stain fibroblast-related components (Cornbrooks et al., 1983a). Anti-laminin antibodies stained cell surfaces in both E15 and E20 outgrowth. E15 outgrowth did not stain with anti-fibronectin antibodies although marked staining was obtained in E20 cultures. Electron microscopy confirmed the presence of only SCs in E15, and of both SCs and fibroblasts in E20 outgrowth. Thus, it appears that there is a narrow developmental window in which the ganglia contain neurons and SCs but relatively few fibroblast components; cultures prepared from ganglia at this stage form outgrowth containing only neurites and SCs without antimitotic treatment. Surprisingly, neither SC ensheathment nor SC basal lamina formation was normal in E15 and E20 outgrowth. When either E15 or E20 SCG SCs were transplanted onto dorsal root ganglion neurons free of endogenous SCs, however, the sensory neurites were typically ensheathed or myelinated and basal lamina appeared 9 d later, identifying the SCG NNCs as functionally competent SCs. The dramatically improved function of SCs relating to sensory axon suggests that these neurons have a more direct influence on SC function than do autonomic neurons; SCs accompanying autonomic neurons may be more influenced by nonaxonal environmental conditions, which, in the culture dish, include the presence of substratum and medium components. In summary, early E15 SCG cultures provide not only a highly convenient source of populations of neurons or neurons plus SCs (without fibroblasts) but also, along with E20 cultures, offer an opportunity to further study factors required for SC differentiation.

Original languageEnglish (US)
Pages (from-to)790-802
Number of pages13
JournalJournal of Neuroscience
Volume6
Issue number3
DOIs
StatePublished - 1986

ASJC Scopus subject areas

  • Neuroscience(all)

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