Student Theses and Dissertations

Date of Award

2011

Document Type

Thesis

RU Laboratory

Hudspeth Laboratory

Keywords

auditory hair bundles, stereocilia, sensory hair cells, glycocalyx, cellular adhesion, hearing

Abstract

The sensory hair cells of the inner ear are exquisitely sensitive machines that translate the broad dynamic range of sound intensities in our auditory landscape into the electrical language of neurons. The mechanosensitive organelle of the hair cell is the hair bundle, a cluster of linked, finger-like, membrane-ensheathed projections, stereocilia, emerging from the cellʼs apical surface. As a structure, the hair bundle is highly conserved, changing little yet performing many functions throughout the vertebrate evolutionary tree. The mechanosensitivity of the hair bundle is achieved by the tension-gating of mechanosensitive channels joined to proteinaceous tip links that connect the distal tips of neighboring stereocilia along the axis of mechanosensitivity. When the hair bundle is deflected and the distal tips of stereocilia shear in relation to one another, tension is applied to the tip links causing the mechanotransduction channels to open. This allows cations to flow in and depolarize the cell membrane triggering synaptic release at the base of the cell, and consequently sending the information to the brain. The cell membranes in the hair bundle face a difficult task when the bundle oscillates in response to sound. For efficient auditory mechanotransduction, it is essential that all stereocilia move nearly in unison, shearing at their distal tips yet maintaining contact without membrane fusion, yet the mechanism producing this cohesion is unknown nor have physical forces associated with it ever been measured. The mechanism I have tested in my doctoral work is that of counterion-mediated tethering of negatively charged sugars on opposing stereociliary membranes. Using capillary electrophoresis, I demonstrated that the stereociliary glycocalyx acts as a negatively charged polymer brush, necessary for the soundness of the glyco-tethering hypothesis. I found by force-fiber photomicrometry that when the distal tips of stereocilia were brought together they formed elastic attachments in a manner dependent on the presence of N-linked sugars and the surrounding ionic environment. Ca2+- and Mg2+-mediated attachments varied in their strength and susceptibility to overcharging, though Mg2+ played a larger role in the observed adhesion. Both partial deglycosylation and removal of divalent ions from surrounding solutions dramatically reduced adhesiveness. During the process of adhesion between the distal tips of stereocilia, chaotic stick-slip friction was observed and appeared qualitatively similar to stick-slip associated with earthquakes. Together, these results indicate that stereocilia are likely to form glycan- and divalent ionmediated attachments to one another that may provide the necessary cohesion for auditory hair bundles. This indicates the importance of the glycocalyx for hearing, and more generally, the biomechanics of cellular adhesion.

Comments

A thesis presented to the faculty of The Rockefeller University in partial fulfillment of the requirements for the degree of Doctor of Philosophy.

Permanent URL

http://hdl.handle.net/10209/393

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