Cytoarchitecture of the mouse organ of corti from base to apex, determined using in situ two-photon imaging.
ABSTRACT: The cells in the organ of Corti are highly organized, with a precise 3D microstructure hypothesized to be important for cochlear function. Here we provide quantitative data on the mouse organ of Corti cytoarchitecture, as determined using a new technique that combines the imaging capabilities of two-photon microscopy with the autofluorescent cell membranes of the genetically modified mTmG mouse. This combination allowed us to perform in situ imaging on freshly excised tissue, thus minimizing any physical distortions to the tissue that extraction from the cochlea and chemical fixation and staining might have caused. 3D image stacks of the organ of Corti were obtained from base to apex in the cochlear duct, from which 3D lengths and relative angles for inner and outer hair cells, Deiters' cells, phalangeal processes, and inner and outer pillars were measured. In addition, intercellular distances, diameters, and stereocilia shapes were obtained. An important feature of this study is the quantitative reporting of the longitudinal tilts of the outer hair cells towards the base of the cochlea, the tilt of phalangeal processes towards the apex, and Deiters' cells that collectively form a Y-shaped building block that is thought to give rise to the lattice-like organization of the organ of Corti. The variations of this Y-shaped element along the cochlear duct and between the rows of outer hair cells are shown with the third row morphologically different from the other rows, and their potential importance for the cochlear amplifier is discussed.
Project description:According to the generally accepted theory of mammalian cochlear mechanics, the fluid in the cochlear scalae interacts with the elastic cochlear partition to generate transversely oscillating displacement waves that propagate along the cochlear coil. Using a computational model of cochlear segments, a different type of propagating wave is reported, an elastic propagating wave that is independent of the fluid-structure interaction. The characteristics of the propagating wave observed in the model, such as the wavelength, speed, and phase lag, are similar to those observed in the living cochlea. Three conditions are required for the existence of the elastic propagating wave in the cochlear partition without fluid-interaction: 1), the stiffness gradient of the cochlear partition; 2), the elastic longitudinal coupling; and 3), the Y-shaped structure in the organ of Corti formed by the outer hair cell, the Deiters cell, and the Deiters cell phalangeal process. The elastic propagating waves in the cochlear partition disappeared without the push-pull action provided by the outer hair cell and Deiters cell phalangeal process. The results suggest that the mechanical feedback of outer hair cells, facilitated by the organ of Corti microstructure, can control the tuning and amplification by modulating the cochlear traveling wave.
Project description:The cochlea performs frequency analysis and amplification of sounds. The graded stiffness of the basilar membrane along the cochlear length underlies the frequency-location relationship of the mammalian cochlea. The somatic motility of outer hair cell is central for cochlear amplification. Despite two to three orders of magnitude change in the basilar membrane stiffness, the force capacity of the outer hair cell's somatic motility, is nearly invariant over the cochlear length. It is puzzling how actuators with a constant force capacity can operate under such a wide stiffness range. We hypothesize that the organ of Corti sets the mechanical conditions so that the outer hair cell's somatic motility effectively interacts with the media of traveling waves-the basilar membrane and the tectorial membrane. To test this hypothesis, a computational model of the gerbil cochlea was developed that incorporates organ of Corti structural mechanics, cochlear fluid dynamics, and hair cell electro-physiology. The model simulations showed that the micro-mechanical responses of the organ of Corti are different along the cochlear length. For example, the top surface of the organ of Corti vibrated more than the bottom surface at the basal (high frequency) location, but the amplitude ratio was reversed at the apical (low frequency) location. Unlike the basilar membrane stiffness varying by a factor of 1700 along the cochlear length, the stiffness of the organ of Corti complex felt by the outer hair cell remained between 1.5 and 0.4 times the outer hair cell stiffness. The Y-shaped structure in the organ of Corti formed by outer hair cell, Deiters cell and its phalange was the primary determinant of the elastic reactance imposed on the outer hair cells. The stiffness and geometry of the Deiters cell and its phalange affected cochlear amplification differently depending on the location.
Project description:The mammalian inner ear detects sound with the organ of Corti, an intricately patterned region of the cochlea in which one row of inner hair cells and three rows of outer hair cells are surrounded by specialized supporting cells. The organ of Corti derives from a prosensory domain that runs the length of the cochlear duct and is bounded by two nonsensory domains, Kölliker's organ on the neural side and the outer sulcus on the abneural side. Although much progress has been made in identifying the signals regulating organ of Corti induction and differentiation, less is known about the mechanisms that establish sensory and nonsensory territories in the cochlear duct. Here, we show that a gradient of bone morphogenetic protein (BMP) signaling is established in the abneural-neural axis of the cochlea. Analysis of compound mutants of Alk3/6 type I BMP receptors shows that BMP signaling is necessary for specification of the prosensory domain destined to form the organ of Corti. Reduction of BMP signaling in Alk3/6 compound mutants eliminates both the future outer sulcus and the prosensory domain, with all cells expressing markers of Kölliker's organ. BMP4 upregulates markers of the future outer sulcus and downregulates marker genes of Kölliker's organ in cochlear organ cultures in a dose-dependent manner. Our results suggest BMP signaling is required for patterning sensory and nonsensory tissue in the mammalian cochlea.
Project description:The organ of Corti has evolved a panoply of cells with extraordinary morphological specializations to harness, direct, and transduce mechanical energy into electrical signals. Among the cells with prominent apical specializations are hair cells and nearby supporting cells. At the apical surface of each hair cell is a mechanosensitive hair bundle of filamentous actin (F-actin)-based stereocilia, which insert rootlets into the F-actin meshwork of the underlying cuticular plate, a rigid organelle considered to hold the stereocilia in place. Little is known about the protein composition and development of the cuticular plate or the apicolateral specializations of organ of Corti supporting cells. We show that supervillin, an F-actin cross-linking protein, localizes to cuticular plates in hair cells of the mouse cochlea and vestibule and zebrafish sensory epithelia. Moreover, supervillin localizes near the apicolateral margins within the head plates of Deiters' cells and outer pillar cells, and proximal to the apicolateral margins of inner phalangeal cells, adjacent to the junctions with neighboring hair cells. Overall, supervillin localization suggests this protein may shape the surface structure of the organ of Corti.
Project description:In the mammalian cochlea, small vibrations of the sensory epithelium are amplified due to active electro-mechanical feedback of the outer hair cells. The level of amplification is greater in the base than in the apex of the cochlea. Theoretical studies have used longitudinally varying active feedback properties to reproduce the location-dependent amplification. The active feedback force has been considered to be proportional to the basilar membrane displacement or velocity. An underlying assumption was that organ of Corti mechanics are governed by rigid body kinematics. However, recent progress in vibration measurement techniques reveals that organ of Corti mechanics are too complicated to be fully represented with rigid body kinematics. In this study, two components of the active feedback are considered explicitly-organ of Corti mechanics, and outer hair cell electro-mechanics. Physiological properties for the outer hair cells were incorporated, such as the active force gain, mechano-transduction properties, and membrane RC time constant. Instead of a kinematical model, a fully deformable 3D finite element model was used. We show that the organ of Corti mechanics dictate the longitudinal trend of cochlear amplification. Specifically, our results suggest that two mechanical conditions are responsible for location-dependent cochlear amplification. First, the phase of the outer hair cell's somatic force with respect to its elongation rate varies along the cochlear length. Second, the local stiffness of the organ of Corti complex felt by individual outer hair cells varies along the cochlear length. We describe how these two mechanical conditions result in greater amplification toward the base of the cochlea.
Project description:Hair cell death is a major cause of hearing impairment. Preservation of surface barrier upon hair cell loss is critical to prevent leakage of potassium-rich endolymph into the organ of Corti and to prevent expansion of cellular damage. Understanding of wound healing in this cytoarchitecturally complex organ requires ultrastructural 3D visualization. Powered by the serial block-face scanning electron microscopy, we penetrate into the cell biological mechanisms in the acute response of outer hair cells and glial-like Deiters' cells to ototoxic trauma in vivo. We show that Deiters' cells function as phagocytes. Upon trauma, their phalangeal processes swell and the resulting close cellular contacts allow engulfment of apoptotic cell debris. Apical domains of dying hair cells are eliminated from the inner ear sensory epithelia, an event thought to depend on supporting cells' actomyosin contractile activity. We show that in the case of apoptotic outer hair cells of the organ of Corti, elimination of their apices is preceded by strong cell body shrinkage, emphasizing the role of the dying cell itself in the cleavage. Our data reveal that the resealing of epithelial surface by junctional extensions of Deiters' cells is dynamically reinforced by newly polymerized F-actin belts. By analyzing Cdc42-inactivated Deiters' cells with defects in actin dynamics and surface closure, we show that compromised barrier integrity shifts hair cell death from apoptosis to necrosis and leads to expanded hair cell and nerve fiber damage. Our results have implications concerning therapeutic protective and regenerative interventions, because both interventions should maintain barrier integrity.
Project description:The mammalian organ of Corti of the inner ear is a highly sophisticated sensory end organ responsible for detecting sound. Noggin is a secreted glycoprotein, which antagonizes bone morphogenetic proteins 2 and 4 (Bmp2 and Bmp4). The lack of this antagonist causes increased rows of inner and outer hair cells in the organ of Corti. In mice, Bmp2 is expressed transiently in nascent cochlear hair cells. To investigate whether Noggin normally modulates the levels of Bmp2 for hair cell formation, we deleted Bmp2 in the cochlear hair cells using two cre strains, Foxg1(cre/+) and Gfi1(cre/+). Bmp2 conditional knockout cochleae generated using these two cre strains show normal hair cells. Furthermore, Gfi1(cre/+);Bmp2(lox/-) mice are viable and have largely normal hearing. The combined results of Noggin and Bmp2 mutants suggest that Noggin is likely to regulate other Bmps in the cochlea such as Bmp4.
Project description:The field of cochlear mechanics has been undergoing a revolution due to recent findings made possible by advancements in measurement techniques. While it has long been assumed that basilar-membrane (BM) motion is the most important determinant of sound transduction by the inner hair cells (IHCs), it turns out that other parts of the sensory epithelium closer to the IHCs, such as the reticular lamina (RL), move with significantly greater amplitude for weaker sounds. It has not been established how these findings are related to the complex cytoarchitecture of the organ of Corti between the BM and RL, which is composed of a lattice of asymmetric Y-shaped elements, each consisting of a basally slanted outer hair cell (OHC), an apically slanted phalangeal process (PhP), and a supporting Deiters' cell (DC). Here, a computational model of the mouse cochlea supports the hypothesis that the OHC micromotors require this Y-shaped geometry for their contribution to the exquisite sensitivity and frequency selectivity of the mammalian cochlea. By varying only the OHC gain parameter, the model can reproduce measurements of BM and RL gain and tuning for a variety of input sound levels. Malformations such as reversing the orientations of the OHCs and PhPs or removing the PhPs altogether greatly reduce the effectiveness of the OHC motors. These results imply that the DCs and PhPs must be properly accounted for in emerging OHC regeneration therapies.
Project description:The organ of Corti in the cochlea is a two-cell layered epithelium: one cell layer of mechanosensory hair cells that align into one row of inner and three rows of outer hair cells interdigitated with one cell layer of underlying supporting cells along the entire length of the cochlear spiral. These two types of epithelial cells are derived from common precursors in the four- to five-cell layered primordium and acquire functionally important shapes during terminal differentiation through the thinning process and convergent extension. Here, we have examined the role of Six1 in the establishment of the auditory sensory epithelium. Our data show that prior to terminal differentiation of the precursor cells, deletion of Six1 leads to formation of only a few hair cells and defective patterning of the sensory epithelium. Previous studies have suggested that downregulation of Sox2 expression in differentiating hair cells must occur after Atoh1 mRNA activation in order to allow Atoh1 protein accumulation due to antagonistic effects between Atoh1 and Sox2. Our analysis indicates that downregulation of Sox2 in the differentiating hair cells depends on Six1 activity. Furthermore, we found that Six1 is required for the maintenance of Fgf8 expression and dynamic distribution of N-cadherin and E-cadherin in the organ of Corti during differentiation. Together, our analyses uncover essential roles of Six1 in hair cell differentiation and formation of the organ of Corti in the mammalian cochlea.
Project description:Supporting cells in the cochlea play critical roles in the development, maintenance, and function of sensory hair cells and auditory neurons. Although the loss of hair cells or auditory neurons results in sensorineural hearing loss, the consequence of supporting cell loss on auditory function is largely unknown. In this study, we specifically ablated inner border cells (IBCs) and inner phalangeal cells (IPhCs), the two types of supporting cells surrounding inner hair cells (IHCs) in mice in vivo. We demonstrate that the organ of Corti has the intrinsic capacity to replenish IBCs/IPhCs effectively during early postnatal development. Repopulation depends on the presence of hair cells and cells within the greater epithelial ridge and is independent of cell proliferation. This plastic response in the neonatal cochlea preserves neuronal survival, afferent innervation, and hearing sensitivity in adult mice. In contrast, the capacity for IBC/IPhC regeneration is lost in the mature organ of Corti, and consequently IHC survival and hearing sensitivity are impaired significantly, demonstrating that there is a critical period for the regeneration of cochlear supporting cells. Our findings indicate that the quiescent neonatal organ of Corti can replenish specific supporting cells completely after loss in vivo to guarantee mature hearing function.