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FM1-43 dye behaves as a permeant blocker of the hair-cell mechanotransducer channel

Gale, J.E., Marcotti, W., Kennedy, H.J., Kros, C.J. and Richardson, G.P. (2001) FM1-43 dye behaves as a permeant blocker of the hair-cell mechanotransducer channel. The Journal of Neuroscience, 21 (18). pp. 7013-7025. ISSN 1529-2401

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Abstract

Hair cells in mouse cochlear cultures are selectively labeled by brief exposure to FM1-43, a styryl dye used to study endocytosis and exocytosis. Real-time confocal microscopy indicates that dye entry is rapid and via the apical surface. Cooling to 4°C and high extracellular calcium both reduce dye loading. Pretreatment with EGTA, a condition that breaks tip links and prevents mechanotransducer channel gating, abolishes subsequent dye loading in the presence of calcium. Dye loading recovers after calcium chelation with a time course similar to that described for tip-link regeneration. Myo7a mutant hair cells, which can transduce but have all mechanotransducer channels normally closed at rest, do not label with FM1-43 unless the bundles are stimulated by large excitatory stimuli. Extracellular perfusion of FM1-43 reversibly blocks mechanotransduction with half-blocking concentrations in the low micromolar range. The block is reduced by high extracellular calcium and is voltage dependent, decreasing at extreme positive and negative potentials, indicating that FM1-43 behaves as a permeant blocker of the mechanotransducer channel. The time course for the relief of block after voltage steps to extreme potentials further suggests that FM1-43 competes with other cations for binding sites within the pore of the channel. FM1-43 does not block the transducer channel from the intracellular side at concentrations that would cause complete block when applied extracellularly. Calcium chelation and FM1-43 both reduce the ototoxic effects of the aminoglycoside antibiotic neomycin sulfate, suggesting that FM1-43 and aminoglycosides enter hair cells via the same pathway.

Item Type: Article
Copyright, Publisher and Additional Information: © 2001 The Society for Neuroscience. Reproduced in accordance with the publisher's self-archiving policy.
Keywords: hair cell, cochlea, mechanotransduction, ion channel, endocytosis, aminoglycosides, myosin VIIA, FM1-43
Institution: The University of Sheffield
Academic Units: The University of Sheffield > Faculty of Science (Sheffield) > School of Biological Sciences (Sheffield) > Department of Biomedical Science (Sheffield)
Depositing User: Ms Suzannah Rockett
Date Deposited: 28 Aug 2008 17:51
Last Modified: 04 Jun 2014 09:51
Published Version: http://www.jneurosci.org/cgi/reprint/21/18/7013
Status: Published
Publisher: HighWire Press
Refereed: Yes
URI: http://eprints.whiterose.ac.uk/id/eprint/4465

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