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The Journal of Neuroscience, October 1, 2000, 20(19):7370-7376

Culture in Reduced Levels of Oxygen Promotes Clonogenic Sympathoadrenal Differentiation by Isolated Neural Crest Stem Cells

Sean J. Morrison^{1, 3}, Marie Csete², Andrew K. Groves⁴, William Melega⁵, Barbara Wold², and David J. Anderson^{1, 2}

¹ Howard Hughes Medical Institute and ² Division of Biology, California Institute of Technology, Pasadena, California 91125, ³ Departments of Internal Medicine and Cell and Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109-0934, ⁴ Department of Cell and Molecular Biology, House Ear Institute, Los Angeles, California 90057, and ⁵ Department of Pharmacology, University of California Los Angeles, Los Angeles, California 90095

Isolated neural crest stem cells (NCSCs) differentiate to autonomic neurons in response to bone morphogenetic protein 2 (BMP2) in clonal cultures, but these neurons do not express sympathoadrenal (SA) lineage markers. Whether this reflects a developmental restriction in NCSCs or simply inappropriate culture conditions was not clear. We tested the growth and differentiation potential of NCSCs at ~5% O₂, which more closely approximates physiological oxygen levels. Eighty-three percent of p75⁺P₀ cells isolated from embryonic day 14.5 sciatic nerve behaved as stem cells under these conditions, suggesting that this is a nearly pure population. Furthermore, addition of BMP2 plus forskolin in decreased oxygen cultures elicited differentiation of thousands of cells expressing tyrosine hydroxylase, dopamine--hydroxylase, and the SA lineage marker SA-1 in nearly all colonies. Such cells also synthesized and released dopamine and norepinephrine. These data demonstrate that isolated mammalian NCSCs uniformly possess SA lineage capacity and further suggest that oxygen levels can influence cell fate. Parallel results indicating that reduced oxygen levels can also promote the survival, proliferation, and catecholaminergic differentiation of CNS stem cells (Studer et al., 2000) suggests that neural stem cells may exhibit a conserved response to reduced oxygen levels.

Key words: sympathoadrenal; neural crest; stem cell; hypoxia; oxygen; sympathetic neuron; dopamine; noradrenergic; autonomic differentiation; cell fate determination

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Copyright © 2000 Society for Neuroscience  0270-6474/00/20197370-07$05.00/0

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