Research Projects
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1. Dominant-negative inhibition of ion channel function. Our lab has developed a powerful strategy for linking the expression of ion channel genes with physiologically defined conductances. We use adenoviral vectors to transfect organotypic cultures of the mouse inner ear. Transfected hair cells are identified by expression of green fluorescent protein (GFP). We can examine control (GFP-negative) and neighboring transfected cells (GFP-positive) and compare the current profiles measured from each cell. Below is a representative figure that shows currents recorded from control and transfected type I hair cells of the mouse utricle. In this case the vector carried the dominant-negative form of the potassium channel gene KCNQ4. For more details see Holt et al., J. Neurosci., 2007.
2. Transduction and adaptation in cochlear hair cells. Since the mouse is a powerful genetic system we have been interested to characterize transduction and adaptation in inner and outer hair cells of the mouse cochlea. We have designed a new system for stimulating cochlear hair cells and used it successfully to evoke robust transduction and adaptation in both inner and outer hair cells. See Stauffer and Holt, J Neurophysiol., 2007 for more details.
3. The role of Myosin 1c in hair cell adaptation. Together with our collaborators we have been studying the function of myosin 1c in hair cells using a novel chemical-genetic strategy. A targeted mutation in the ATP binding pocket of myosin 1c allows for chemical inhibition of the mutant protein but not the wild-type form. We have examined hair cell adaptation in transgenic and knock-in mice that express the mutant form of myosin 1c and find that introduction of the selective inhibitor blocks adaptation in vestibular hair cells of transgenic and knock-in mice but not wild-type mice. For more information see Holt et al., Cell, 2002 and Stauffer et al., Neuron, 2005.
5. In vitro transfection of human hair cells. We have developed a novel model system to study gene therapy compounds that may be developed to treat inner ear disorders in humans. This system is based on inner ear tissue harvested from human patients. All patients offered their consent and were selected for this study because they had already lost inner function on the affected side due to a specific brain tumor growing on the 8th cranial nerve. During the procedure the surgeons were able to remove the inner ear tissue. In the lab we developed the necessary techniques to keep the tissue alive for up to five days. We have used the tissue to study the human hair cells and transfect them with wild-type genes that are known to cause deafness when mutated. We suggest that this model system will be a valuable proving ground for testing potential gene therapy strategies and will thus facilitate the translation of basic discoveries from the lab into effective treatments in the clinic. For more information see Kesser et al., Gene Therapy, 2007.
6. Functional development of hair cells. For this project we are interested to identify when during development hair cells acquire their unique properties of mechanotransduction and sensory signaling. In the mouse vestibular system we have identified a developmental window between embryonic day 16 and 17 during which mechanosensitivity is acquired. See Géléoc and Holt., Nature Neuroscience, 2003 for details. We have also examined the functional maturation of voltage-dependent conductances in the basolateral hair cell membrane. For more information see Geleoc et al., J. Neurosci., 2004.
If you have questions about these or any other projects please feel free to email Jeff Holt (jeffholt@virginia.edu) or Gwen Géléoc (gg3h@virginia.edu).
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