The goal of the Section on Developmental Neuroscience is to determine the factors that play a role in the development of the sensory epithelium of the mammalian cochlea, the organ of Corti. The organ of Corti is comprised of a highly rigorous pattern of cells and its development is crucial for normal hearing. During the last year, members of the laboratory concentrated on several different issues related to the development of the organ of Corti. A key step in the development of the mammalian organ of Corti is the establishment of appropriate bundle polarity. Since sensory hair cells are only sensitive to vibrations that lead to deflections of the stereociliary bundle in a single direction, the establishment of bundle orientation or polarity is crucial for the normal perception of sound. Based on the known functions of Wnt proteins, we began a project to examine the possible role of Wnts in determination of bundle orientation. Results indicate that Wnt-7a is expressed in a gradient throughout the outer hair cell region. Addition of exogenous Wnt-7a protein to developing cochlear cultures leads to bundle misorientation, while inhibition of a downstream target of Wnt-7a, Calmodulin Kinase II, also leads to bundle misorientation. These results suggest that developing hair cells may use a gradient of Wnt-7a protein to appropriately orient their stereociliary bundles. The PKC signaling pathway has been shown to play a key role in many aspects of cell signaling. To determine whether PKC might play a role in development of the mammalian cochlea, cochlear cultures were exposed to PKC activators or inhibitors. Results indicate that inhibition of PKC leads to a significant increase in the number of cells that develop as inner hair cells while activation of PKC results in a decrease in the number of inner hair cells. Analysis of the levels of mRNA for a key hair cell gene, Math1, indicates that inhibition of PKC leads to an increase in Math1 mRNA within 24 hours. These results suggest that PKC may play a key role in regulating the development of the mammalian organ of Corti. A second set of important cell types within the mammalian organ of Corti are the pillar cells that are located between the inner and outer hair cells. Inner and outer pillar cells form the medial and lateral walls of the tunnel of Corti, a structure that is required for normal hearing. The factors that play a role in pillar cell formation are poorly understood. To begin to identify factors that could play a role in pillar cell formation, we inhibited the activation of fibroblast growth factor receptor 3 (FGFr3). Previous studies have suggested that FGFr3 is expressed in pillar cells and may play a role in their development. Using a pharmacological antagonist of FGFr3, we were able to demonstrate that inhibition of FGFr3 leads to a disruption in the development of cells as pillar cells. Results also indicated that constant stimulation of FGFr3 is required for the ongoing development of cells as pillar cells. Finally, the organ of Corti is characterized by a rigorous pattern of specific cell types. However, the factors that play a role in the establishment of this pattern are poorly understood. To begin to identify the timing of different patterning events, we isolated different regions of the developing cochlear duct at early developmental time points. Results indicate that interactions between developing cochlear epithelial cells and underlying mesenchymal cells play a key role in establishing the cellular pattern of the organ of Corti at early developmental time points. Moreover, the coordinated timing of the development of the entire organ is controlled through intrinsic factors within the epithelium. In collaborative projects, the laboratory participated in two projects related to understanding the role of thyroid hormone during the development of the inner ear. In collaboration with Sheue-Yann Chang at NCI and with Andrew Griffith at NIDCD we have analyzed the phenotype in mice that are transgenic for a mutated form of the Thyroid Hormone receptor beta gene that leads to a syndrome called """"""""Resistance to Thyroid Hormone (RTH)"""""""" in humans. Results indicate that the cochleae from these animals appear normal at birth, but by three weeks of age severe defects develop in the tectorial membrane that result in profound deafness. In a second collaboration with Douglas Forrest at Mt. Sinai Hospital in New York, we have analyzed the effects of complete deletion of both thyroid receptors. Results indicate severe defects in the tectorial membranes from these animals by as early as postnatal day 8. In adults, the tectorial membrane defects persist and there is degeneration of the organ of Corti as well. The results of these projects strongly implicate thyroid hormone as an important factor in the development of the tectorial membrane as well as in the overall maintenance of the cellular structure of the organ of Corti.
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