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The Journal of Neuroscience, November 1, 1999, 19(21):9445-9458

Differentiation of Mammalian Vestibular Hair Cells from Conditionally Immortal, Postnatal Supporting Cells

Patrick Lawlor, Walter Marcotti, Marcelo N. Rivolta, Corné J. Kros, and Matthew C. Holley

Department of Physiology, School of Medical Sciences, University of Bristol, Bristol, BS8 1TD, United Kingdom

We provide evidence from a newly established, conditionally immortal cell line (UB/UE-1) that vestibular supporting cells from the mammalian inner ear can differentiate postnatally into more than one variant of hair cell. A clonal supporting cell line was established from pure utricular sensory epithelia of H2k^btsA58 transgenic mice 2 d after birth. Cell proliferation was dependent on conditional expression of the immortalizing gene, the "T" antigen from the SV40 virus. Proliferating cells expressed cytokeratins, and patch-clamp recordings revealed that they all expressed small membrane currents with little time-dependence. They stopped dividing within 2 d of being transferred to differentiating conditions, and within a week they formed three defined populations expressing membrane currents characteristic of supporting cells and two kinds of neonatal hair cell. The cells expressed several characteristic features of normal hair cells, including the transcription factor Brn3.1, a functional acetylcholine receptor composed of 9 subunits, and the cytoskeletal proteins myosin VI, myosin VIIa, and fimbrin. Immunofluorescence labeling and electron microscopy showed that the cells formed complex cytoskeletal arrays on their upper surfaces with structural features resembling those at the apices of normal hair cells. The cell line UB/UE-1 provides a valuable in vitro preparation in which the expression of numerous structural and physiological components can be initiated or upregulated during early stages of mammalian hair cell commitment and differentiation.

Key words: mouse; vestibular; utricle; hair cells; epithelial cells; conditional immortalization; tsA58; differentiation; development; potassium current; inward rectifier

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Copyright © 1999 Society for Neuroscience  0270-6474/99/19219445-14$05.00/0

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