University of Virginia Health System

The auditory and vestibular systems of the vertebrate inner ear possess sensory cells that transduce the appropriate stimuli (sound in the cochlea, linear acceleration in the saccule and utricule and rotational velocity in the semicircular canals) into receptor potentials. These "hair cells", so called because of their apical microvilli structure, are the mechanoreceptors of the inner ear.  Mechanical stimuli applied to the bundle of microvilli stretch extracellular links, pulling open mechano-sensitive transducer channels.  Opening of transducer channels allows the passage of cations, primarily potassium and calcium, which generates a receptor current.  The receptor current activates rapidly (within microseconds).  For sustained stimuli the current decays or adapts over 10-100 milliseconds.  Adaptation has been studied biophysically in turtle and frog hair cells.  Our aim is to understand and characterize hair cell transduction and adaptation in mammals and to identify the molecular elements involved.

We use a glass micropipette or fluid puff to deflect the sensory bundles. Receptor currents can be recorded in vitro using a whole-cell voltage-clamp. Hair cells respond to extremely small deflections of a few tens of nanometers with currents as large as few hundred picoamps

II-SENSORY SIGNALLING IN DEVELOPING VESTIBULAR HAIR CELLS

Despite recent advances in the genetics of hearing loss, there has been little progress toward understanding the pathophysiology of congenital hearing and balance dysfunction. Many of these disorders are thought to be due to degeneration, lack or malfunction of the hair cells of the inner ear sensory epithelia. An obvious prerequisite to understanding prenatal disorders is knowledge of how hair cells develop in the normal state. In mice, hair cells begin to appear around E12 and hair bundles are first observed around E13. We are interested to determine precisely how and when hair cells acquire the remarkable functions of mechano-transduction and sensory signaling.

Studying the development of hair cell transduction, we have demonstrated the concurrent acquisition of transduction elements and a rapid, all-or-nothing onset of fully functional mechanosensitivity between E16 and E17 (Géléoc and Holt, 2003). The precisely defined developmental timeline of transduction acquisition has provided an opportunity for identification of the genes required for normal function. Using real-time PCR we have searched for potential mechanotransducer channels and tested them functionally using si-RNAs carried by viral vectors. This work has demonstrated that TRPA1 is a potential candidate for the channel (Nature 2004). We are now studying the expression pattern of other molecules which are or might be part of the mechanotransduction complex.

Studying the development of hair cell function, we also investigate the expresssion of voltage dependent conductances and their molecular correlates. We have recently described how ionic conductances are acquired in mouse embryonic hair cells (J. Neurosci. 2004). Our work has demonstrated that hair cells start differentiating into type I and type II cells at around E18. We are now focused on the molecular identification of the various conductances.

Acquisition of the low-voltage-activated conductance, GK,L, at approximately E18

• G.S.G. Géléoc, G.W.T. Lennan, G.P. Richardson, and C.J. Kros:  A quantitative comparison of mechano-electrical transduction in vestibular and auditory hair cells of neonatal mice (1997) Proceedings of the Royal Society London, 264: 611-621.
• G.S.G. Géléoc, S.O. Casalotti, A. Forge and J.F. Ashmore:  A sugar transporter as the outer hair cell motor. (1999) Nature neuroscience, 8: 713-719.
• W. Marcotti, G.S.G. Géléoc, G.W.T. Lennan  and C.J. Kros:  Developmental expression of an inwardly rectifying potassium conductance in inner and outer hair cells along the mouse cochlea. (1999) European Journal of Physiology, Pflüger Arch, 439: 113-122.
• G.S.G. Géléoc and J.R. Holt:  Auditory amplification: Outer hair cells pres the issue. (2003) Trends in Neuroscience, 26(3):115-117.
• G.S.G. Géléoc and J.R. Holt:  Developmental acquisition of sensory transduction in hair cells of the mouse inner ear. (2003) Nature Neuroscience, Online 09-14-03 doi:10.1038/nn1120.
• D.P Corey, J. Garcia-Anoveros, J.R. Holt, K.Y. Kwan, S.Y. Lin, M.A. Vollrath, A. Amalfitano, E. Cheung., B.H. Derfler, A. Duggan, G.S.G. Géléoc, P. Gray, M.P. Hoffman, N. Hopkins, H.L. Rehm, D. Tamasauskas, and D.S. Zhang: TRPA1 is a candidate for the mechanosensitive transduction channel of vertebrate hair cells. (2004) Nature, 432:723-730.
• G.S.G. Géléoc, Jessica Risner and J.R. Holt: Developmental Acquisition of Voltage Dependent Conductances and Sensory Signaling in Hair Cells of the Embryonic Mouse Inner Ear. (2004), J. Neuroscience, 24:11148-11159.