Visualization of the mechanosensory machinery in live mammalian auditory hair cells

The inner ear detects sound-induced vibrations through deflection of rigid projections at the apex of mechanosensory hair cells, known as stereocilia. Stereocilia of a hair cell form the “hair bundle”. Sound is detected when stereocilia are deflected and tug on tiny extracellular tip links that gate...

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Autores:
Galeano Naranjo, Carolina
Tipo de recurso:
Fecha de publicación:
2021
Institución:
Universidad Nacional de Colombia
Repositorio:
Universidad Nacional de Colombia
Idioma:
eng
OAI Identifier:
oai:repositorio.unal.edu.co:unal/81543
Acceso en línea:
https://repositorio.unal.edu.co/handle/unal/81543
https://repositorio.unal.edu.co/
Palabra clave:
660 - Ingeniería química::666 - Cerámica y tecnologías afines
Piezoelectricidad
Transducción
Scanning ion conductance microscopy
Super-resolution
Live-cell imaging
Cochlear hair cells
Stereocilia
Mechanotransduction
: Microscopía de conductancia Iónica de barrido
Visualización de células vivas
Células ciliadas del oído
Estereocilios
Mecanotransducción
Rights
openAccess
License
Atribución-NoComercial-SinDerivadas 4.0 Internacional
id UNACIONAL2_23e87508806ae6963c9888d61b3c032d
oai_identifier_str oai:repositorio.unal.edu.co:unal/81543
network_acronym_str UNACIONAL2
network_name_str Universidad Nacional de Colombia
repository_id_str
dc.title.eng.fl_str_mv Visualization of the mechanosensory machinery in live mammalian auditory hair cells
dc.title.translated.spa.fl_str_mv Visualización de la maquinaria mecanosensorial en celulas ciliadas vivas
title Visualization of the mechanosensory machinery in live mammalian auditory hair cells
spellingShingle Visualization of the mechanosensory machinery in live mammalian auditory hair cells
660 - Ingeniería química::666 - Cerámica y tecnologías afines
Piezoelectricidad
Transducción
Scanning ion conductance microscopy
Super-resolution
Live-cell imaging
Cochlear hair cells
Stereocilia
Mechanotransduction
: Microscopía de conductancia Iónica de barrido
Visualización de células vivas
Células ciliadas del oído
Estereocilios
Mecanotransducción
title_short Visualization of the mechanosensory machinery in live mammalian auditory hair cells
title_full Visualization of the mechanosensory machinery in live mammalian auditory hair cells
title_fullStr Visualization of the mechanosensory machinery in live mammalian auditory hair cells
title_full_unstemmed Visualization of the mechanosensory machinery in live mammalian auditory hair cells
title_sort Visualization of the mechanosensory machinery in live mammalian auditory hair cells
dc.creator.fl_str_mv Galeano Naranjo, Carolina
dc.contributor.advisor.none.fl_str_mv Frolenkov, Gregory
Vélez Ortega, A. Catalina
Vásquez Araque, Neil Aldrin
dc.contributor.author.none.fl_str_mv Galeano Naranjo, Carolina
dc.contributor.researchgroup.spa.fl_str_mv Frolenkov Laboratory
dc.subject.ddc.spa.fl_str_mv 660 - Ingeniería química::666 - Cerámica y tecnologías afines
topic 660 - Ingeniería química::666 - Cerámica y tecnologías afines
Piezoelectricidad
Transducción
Scanning ion conductance microscopy
Super-resolution
Live-cell imaging
Cochlear hair cells
Stereocilia
Mechanotransduction
: Microscopía de conductancia Iónica de barrido
Visualización de células vivas
Células ciliadas del oído
Estereocilios
Mecanotransducción
dc.subject.other.none.fl_str_mv Piezoelectricidad
Transducción
dc.subject.proposal.eng.fl_str_mv Scanning ion conductance microscopy
Super-resolution
Live-cell imaging
Cochlear hair cells
Stereocilia
Mechanotransduction
dc.subject.proposal.spa.fl_str_mv : Microscopía de conductancia Iónica de barrido
Visualización de células vivas
Células ciliadas del oído
Estereocilios
Mecanotransducción
description The inner ear detects sound-induced vibrations through deflection of rigid projections at the apex of mechanosensory hair cells, known as stereocilia. Stereocilia of a hair cell form the “hair bundle”. Sound is detected when stereocilia are deflected and tug on tiny extracellular tip links that gate mechanotransduction channels. Any study of the potential dynamic changes within this mechano-electrical transduction (MET) machinery is currently challenging due to small sizes of their components, in a range of 5-200 nm. Conventional label-free optical microscopy has a best resolution of ~200 nm. Therefore, it is impossible to visualize the MET apparatus in live cells. Super-resolution imaging has been only of a limited use due to the requirement of fluorescent labeling. Therefore, ultrastructural details of the MET apparatus in the auditory hair cells have been investigated mostly with electron microscopy (EM). Unfortunately, EM techniques require chemical or cryofixation of the sample, making the study of dynamic processes in the MET machinery nearly impossible or extremely labor intensive. In theory, scanning probe techniques, such as atomic force microscopy (AFM), have enough resolution to visualize ultrastructural details in live cells. However, the efforts to image hair cell bundles with AFM were not too encouraging, since stereocilia bundles are usually damaged after even the slightest contact of the AFM probe with them. The hopping probe ion conductance microscopy (HPICM) is an alternative non-contact scanning probe technique capable of performing time lapse imaging of the surface of live cells with a complex topography, with single nanometer resolution and without making physical contact with the sample. The HPICM uses an electrical current passing through a glass nanopipette to detect the cell surface in close vicinity to the pipette, while a 3D-positioning piezoelectric system scans the surface and generates its image. The goal of this project was to optimize HPICM for visualization of nanoscale structures on the surface of the stereocilia in live auditory hair cells. We were able to visualize stereocilia bundles in live inner hair cells including the links interconnecting stereocilia (~5 nm in diameter). We were also able to obtain time-lapse imaging of these bundles. We believe that this is an important step toward our goal of studying ultrastructural changes in the mechanosensory machinery of live hair cells with HPICM.
publishDate 2021
dc.date.issued.none.fl_str_mv 2021
dc.date.accessioned.none.fl_str_mv 2022-06-08T21:24:12Z
dc.date.available.none.fl_str_mv 2022-06-08T21:24:12Z
dc.type.spa.fl_str_mv Trabajo de grado - Maestría
dc.type.driver.spa.fl_str_mv info:eu-repo/semantics/masterThesis
dc.type.version.spa.fl_str_mv info:eu-repo/semantics/acceptedVersion
dc.type.content.spa.fl_str_mv Text
dc.type.redcol.spa.fl_str_mv http://purl.org/redcol/resource_type/TM
status_str acceptedVersion
dc.identifier.uri.none.fl_str_mv https://repositorio.unal.edu.co/handle/unal/81543
dc.identifier.instname.spa.fl_str_mv Universidad Nacional de Colombia
dc.identifier.reponame.spa.fl_str_mv Repositorio Institucional Universidad Nacional de Colombia
dc.identifier.repourl.spa.fl_str_mv https://repositorio.unal.edu.co/
url https://repositorio.unal.edu.co/handle/unal/81543
https://repositorio.unal.edu.co/
identifier_str_mv Universidad Nacional de Colombia
Repositorio Institucional Universidad Nacional de Colombia
dc.language.iso.spa.fl_str_mv eng
language eng
dc.relation.references.spa.fl_str_mv Beurg, M., Fettiplace, R., Nam, J. H., & Ricci, A. J. (2009). Localization of inner hair cell mechanotransducer channels using high-speed calcium imaging. Nat Neurosci, 12(5), 553-558. https://doi.org/10.1038/nn.2295
Conchello, J. A., & Lichtman, J. W. (2005). Optical sectioning microscopy. Nat Methods, 2(12), 920-931. https://doi.org/10.1038/nmeth815
Dufrene, Y. F. (2008). Towards nanomicrobiology using atomic force microscopy. Nat Rev Microbiol, 6(9), 674-680. https://doi.org/10.1038/nrmicro1948
Engström, H., & Engström, B. (1978). Structure of the hairs on cochlear sensory cells. Hearing Research, 1, 49-66.
Fettiplace, R. (2017). Hair Cell Transduction, Tuning, and Synaptic Transmission in the Mammalian Cochlea. Compr Physiol, 7(4), 1197-1227. https://doi.org/10.1002/cphy.c160049
Fettiplace, R., & Hackney, C. M. (2006). The sensory and motor roles of auditory hair cells. Nat Rev Neurosci, 7(1), 19-29. https://doi.org/doi: 10.1038/nrn1828
Frolenkov, G. I., Belyantseva, I. A., Friedman, T. B., & Griffith, A. J. (2004). Genetic insights into the morphogenesis of inner ear hair cells. Nat Rev Genet, 5(7), 489-498. https://doi.org/10.1038/nrg1377
Furness, D. N., & Hackney, C. M. (1985). Cross-links between stereocilia in the guinea pig cochlea. Hearing Research, 18, 177-188.
Galeano-Naranjo, C., Velez-Ortega, A. C., & Frolenkov, G. I. (2021). Stereocilia Bundle Imaging with Nanoscale Resolution in Live Mammalian Auditory Hair Cells. J Vis Exp(167). https://doi.org/10.3791/62104
Gavara, N., Manoussaki, D., & Chadwick, R. S. (2011). Auditory mechanics of the tectorial membrane and the cochlear spiral. Curr Opin Otolaryngol Head Neck Surg, 19(5), 382-387. https://doi.org/10.1097/MOO.0b013e32834a5bc9
Goodyear, R. J., Marcotti, W., Kros, C. J., & Richardson, G. P. (2005). Development and properties of stereociliary link types in hair cells of the mouse cochlea. J Comp Neurol, 485(1), 75-85. https://doi.org/10.1002/cne.20513
Hadi, S., Alexander, A. J., Velez-Ortega, A. C., & Frolenkov, G. I. (2020). Myosin-XVa Controls Both Staircase Architecture and Diameter Gradation of Stereocilia Rows in the Auditory Hair Cell Bundles. J Assoc Res Otolaryngol, 21(2), 121-135. https://doi.org/10.1007/s10162-020-00745-4
Hansma, P. K., Drake, B., Marti, O., Gould, S. A., & Prater, C. B. (1989). The scanning ion-conductance microscope. Science, 243(4891), 641-643. https://doi.org/10.1126/science.2464851
Ivanchenko, M. V., Cicconet, M., Jandal, H. A., Wu, X., Corey, D. P., & Indzhykulian, A. A. (2020). Serial scanning electron microscopy of anti-PKHD1L1 immuno-gold labeled mouse hair cell stereocilia bundles. Sci Data, 7(1), 182. https://doi.org/10.1038/s41597-020-0509-4
Jacobs, R. A., & Hudspeth, A. J. (1990). Ultrastructural correlates of mechanoelectrical transduction in hair cells of the bullfrog’s internal ear. Cold Spring Harbor Symposia on Quantitative Biology, 55, 547-561. https://doi.org/10.1101/sqb.1990.055.01.053
Kachar, B., Parakkal, M., Kurc, M., Zhao, Y., & Gillespie, P. G. (2013). High-resolution structure of hair-cell tip links. Proceedings of the National Academy of Sciences, 110(29), 12155-12155. https://doi.org/10.1073/pnas.1311228110
Kazmierczak, M., Harris, S. L., Kazmierczak, P., Shah, P., Starovoytov, V., Ohlemiller, K. K., & Schwander, M. (2015). Progressive Hearing Loss in Mice Carrying a Mutation in Usp53. J Neurosci, 35(47), 15582-15598. https://doi.org/10.1523/JNEUROSCI.1965-15.2015
Korchev, Y. E., Bashford, C. L., Milovanovic, M., Vodyanoy, I., & Lab, M. J. (1997). Scanning Ion Conductance Microscopy of Living Cells. Biophys Journal, 73, 653-658. https://doi.org/10.1016/S0006-3495(97)78100-1
Korchev, Y. E., Milovanovic, M., Bashford, C. L., Bennett, D. C., Sviderskaya, E. V., Vodyanoy, I., & Lab, M. J. (1997). Specialized scanning ion-conductance microscope for imaging of living cells. Journal of microscopy, 188, 17-23.
Krey, J. F., & Gillespie, P. G. (2012). Molecular Biology of Hearing and Balance. Basic neurochemistry, 916-927. https://doi.org/10.1016/B978-0-12-374947-5.00053-5
Langer, M. G., Koitschev, A., Haase, H., Rexhausen, U., Hörber, J. K., & Ruppersberg, J. P. (2000). Mechanical stimulation of individual stereocilia of living cochlear hair cells by atomic force microscopy. Ultramicroscopy, 82, 269-278. https://doi.org/10.1016/s0304-3991(99)00136-9
Lehnhard, E., & Lehnhardt, M. (2003). Study Letter 2: Functional Anatomy, Physiology and Pathology of the Auditory System. COMENIUS 2.1 ACTION: Qualification of educational staff working with hearing-impaired children (QESWHIC)
Metlagel, Z., Krey, J. F., Song, J., Swift, M. F., Tivol, W. J., Dumont, R. A., Thai, J., Chang, A., Seifikar, H., Volkmann, N., Hanein, D., Barr-Gillespie, P. G., & Auer, M. (2019). Electron cryo-tomography of vestibular hair-cell stereocilia. J Struct Biol, 206(2), 149-155. https://doi.org/10.1016/j.jsb.2019.02.006
Novak, P., Li, C., Shevchuk, A. I., Stepanyan, R., Caldwell, M., Hughes, S., Smart, T. G., Gorelik, J., Ostanin, V. P., Lab, M. J., Moss, G. W., Frolenkov, G. I., Klenerman, D., & Korchev, Y. E. (2009). Nanoscale live-cell imaging using hopping probe ion conductance microscopy. Nat Methods, 6(4), 279-281. https://doi.org/10.1038/nmeth.1306
Pickles, J. O., Comis, S. D., & Osborne, M. P. (1984). Cross-links between stereocilia in the guinea pig organ of Corti, and their possible relation to sensory transduction. Hearing Research, 15(2), 103-112. https://doi.org/10.1016/0378-5955(84)90041-8
Salt, A. N., & Hirose, K. (2018). Communication pathways to and from the inner ear and their contributions to drug delivery. Hear Res, 362, 25-37. https://doi.org/10.1016/j.heares.2017.12.010
Shevchuk, A., Tokar, S., Gopal, S., Sanchez-Alonso, J. L., Tarasov, A. I., Velez-Ortega, A. C., Chiappini, C., Rorsman, P., Stevens, M. M., Gorelik, J., Frolenkov, G. I., Klenerman, D., 57 & Korchev, Y. E. (2016). Angular Approach Scanning Ion Conductance Microscopy. Biophys J, 110(10), 2252-2265. https://doi.org/10.1016/j.bpj.2016.04.017
Sigal, Y. M., Zhou, R., & Zhuang, X. (2018). Visualizing and discovering cellular structures with super-resolution microscopy. Science, 361, 880-887.
Sutter Instrument. (2010). P-2000 Micropipette Puller Operation Manual. https://www.sutter.com/manuals/P-2000_OpMan.pdf
Velez-Ortega, A. C., Freeman, M. J., Indzhykulian, A. A., Grossheim, J. M., & Frolenkov, G. I. (2017). Mechanotransduction current is essential for stability of the transducing stereocilia in mammalian auditory hair cells. Elife, 6. https://doi.org/10.7554/eLife.24661
Velez-Ortega, A. C., & Frolenkov, G. I. (2016). Visualization of Live Cochlear Stereocilia at a Nanoscale Resolution Using Hopping Probe Ion Conductance Microscopy. Methods Mol Biol, 1427, 203-221. https://doi.org/10.1007/978-1-4939-3615-1_12
Vélez-Ortega, A. C., & Frolenkov, G. I. (2019). Building and repairing the stereocilia cytoskeleton in mammalian auditory hair cells. Hear Res, 376, 47-57. https://doi.org/10.1016/j.heares.2018.12.012
Waldchen, S., Lehmann, J., Klein, T., van de Linde, S., & Sauer, M. (2015). Light-induced cell damage in live-cell super-resolution microscopy. Sci Rep, 5, 15348. https://doi.org/10.1038/srep15348
Wangemann, P. (2006). Supporting sensory transduction: cochlear fluid homeostasis and the endocochlear potential. J Physiol, 576(0022-3751 (Print)), 11-21. https://doi.org/10.1113/jphysiol.2006.112888
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dc.format.extent.spa.fl_str_mv xvi, 58 páginas
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dc.publisher.spa.fl_str_mv Universidad Nacional de Colombia
dc.publisher.program.spa.fl_str_mv Medellín - Ciencias - Maestría en Ciencias - Biotecnología
dc.publisher.department.spa.fl_str_mv Escuela de biociencias
dc.publisher.faculty.spa.fl_str_mv Facultad de Ciencias
dc.publisher.place.spa.fl_str_mv Medellín
dc.publisher.branch.spa.fl_str_mv Universidad Nacional de Colombia - Sede Medellín
institution Universidad Nacional de Colombia
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spelling Atribución-NoComercial-SinDerivadas 4.0 Internacionalhttp://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/openAccesshttp://purl.org/coar/access_right/c_abf2Frolenkov, Gregoryddb35dc57b3a87a7b6a6ea8c776f4233600Vélez Ortega, A. Catalinac41f5a44f65126c38868f648538fdbe6Vásquez Araque, Neil Aldrinb738c93743b300229ba7dce362dc8bc0600Galeano Naranjo, Carolina663e69405e3b5debac197b2e02dbff64Frolenkov Laboratory2022-06-08T21:24:12Z2022-06-08T21:24:12Z2021https://repositorio.unal.edu.co/handle/unal/81543Universidad Nacional de ColombiaRepositorio Institucional Universidad Nacional de Colombiahttps://repositorio.unal.edu.co/The inner ear detects sound-induced vibrations through deflection of rigid projections at the apex of mechanosensory hair cells, known as stereocilia. Stereocilia of a hair cell form the “hair bundle”. Sound is detected when stereocilia are deflected and tug on tiny extracellular tip links that gate mechanotransduction channels. Any study of the potential dynamic changes within this mechano-electrical transduction (MET) machinery is currently challenging due to small sizes of their components, in a range of 5-200 nm. Conventional label-free optical microscopy has a best resolution of ~200 nm. Therefore, it is impossible to visualize the MET apparatus in live cells. Super-resolution imaging has been only of a limited use due to the requirement of fluorescent labeling. Therefore, ultrastructural details of the MET apparatus in the auditory hair cells have been investigated mostly with electron microscopy (EM). Unfortunately, EM techniques require chemical or cryofixation of the sample, making the study of dynamic processes in the MET machinery nearly impossible or extremely labor intensive. In theory, scanning probe techniques, such as atomic force microscopy (AFM), have enough resolution to visualize ultrastructural details in live cells. However, the efforts to image hair cell bundles with AFM were not too encouraging, since stereocilia bundles are usually damaged after even the slightest contact of the AFM probe with them. The hopping probe ion conductance microscopy (HPICM) is an alternative non-contact scanning probe technique capable of performing time lapse imaging of the surface of live cells with a complex topography, with single nanometer resolution and without making physical contact with the sample. The HPICM uses an electrical current passing through a glass nanopipette to detect the cell surface in close vicinity to the pipette, while a 3D-positioning piezoelectric system scans the surface and generates its image. The goal of this project was to optimize HPICM for visualization of nanoscale structures on the surface of the stereocilia in live auditory hair cells. We were able to visualize stereocilia bundles in live inner hair cells including the links interconnecting stereocilia (~5 nm in diameter). We were also able to obtain time-lapse imaging of these bundles. We believe that this is an important step toward our goal of studying ultrastructural changes in the mechanosensory machinery of live hair cells with HPICM.El oído interno detecta vibraciones inducidas por el sonido a través de la desviación de proyecciones rígidas en la superficie apical de las células ciliadas mecanosensoriales, conocidas como estereocilios. Los estereocilios de una célula ciliada forman el "haz de cilios". El sonido se detecta cuando los estereocilios se desvían y tiran de unas pequeñas uniones extracelulares que abren los canales de mecano-transducción. El estudio de posibles cambios dinámicos de la maquinaria de transducción mecano-eléctrica (MET) es actualmente un desafío, debido al tamaño de sus estructuras, en un rango de 5-200 nm. La microscopía óptica convencional sin etiquetas fluorescentes (fluorescent tags) tiene una resolución máxima de ~ 200 nm. Debido a esto, es imposible visualizar el aparato MET en células vivas. Por otro lado, la microscopia de superresolución ha tenido un uso limitado debido a que es necesario el uso de moléculas fluorescentes. Por lo tanto, los detalles ultraestructurales del aparato MET en las células ciliadas auditivas se han investigado principalmente con microscopía electrónica (EM). Desafortunadamente, las técnicas EM requieren de fijación química o criogénica de la muestra, lo que hace que el estudio de los procesos dinámicos en la maquinaria MET sea casi imposible o extremadamente laborioso. En teoría, las técnicas de exploración con sonda, como la microscopía de fuerza atómica (AFM por sus sigla en inglés), tienen suficiente resolución para visualizar detalles ultraestructurales en células vivas. Sin embargo, los esfuerzos para obtener imágenes de los haces de células ciliadas con AFM no han sido demasiado alentadores, ya que los haces de estereocilios generalmente se dañan al mínimo contacto con la sonda de AFM. Como alternativa, la microscopía de conductancia iónica con sonda de salto (HPICM por sus sigla en inglés) es una técnica de sonda de exploración sin contacto capaz de realizar imágenes de lapso de tiempo (time lapse) de la superficie de células vivas con una topografía compleja, con una resolución de un solo nanómetro y sin hacer contacto físico con la muestra. El HPICM utiliza una corriente eléctrica que pasa a través de una nanopipeta de vidrio para detectar la superficie de la célula en las proximidades de la pipeta, mientras que un sistema piezoeléctrico de posicionamiento 3D escanea la superficie y genera su imagen. El objetivo de este proyecto fue optimizar el HPICM para la visualización de estructuras a nanoescala en la superficie de los estereocilios en células ciliadas auditivas vivas. Se lograron visualizar haces de estereocilios en células ciliadas internas vivas, incluidos los enlaces que interconectan los estereocilios (~ 5 nm de diámetro). También se pudieron obtener imágenes de lapso de tiempo de estos haces de cilios. Se considera, que este es un paso importante hacia el objetivo de estudiar los cambios ultraestructurales en la maquinaria mecano-sensorial de las células ciliadas vivas con HPICM. (texto tomado de la fuente)MaestríaMaestría en Ciencias - BiotecnologíaÁrea curricular Biotecnologíaxvi, 58 páginasapplication/pdfengUniversidad Nacional de ColombiaMedellín - Ciencias - Maestría en Ciencias - BiotecnologíaEscuela de biocienciasFacultad de CienciasMedellínUniversidad Nacional de Colombia - Sede Medellín660 - Ingeniería química::666 - Cerámica y tecnologías afinesPiezoelectricidadTransducciónScanning ion conductance microscopySuper-resolutionLive-cell imagingCochlear hair cellsStereociliaMechanotransduction: Microscopía de conductancia Iónica de barridoVisualización de células vivasCélulas ciliadas del oídoEstereociliosMecanotransducciónVisualization of the mechanosensory machinery in live mammalian auditory hair cellsVisualización de la maquinaria mecanosensorial en celulas ciliadas vivasTrabajo de grado - Maestríainfo:eu-repo/semantics/masterThesisinfo:eu-repo/semantics/acceptedVersionTexthttp://purl.org/redcol/resource_type/TMBeurg, M., Fettiplace, R., Nam, J. H., & Ricci, A. J. (2009). Localization of inner hair cell mechanotransducer channels using high-speed calcium imaging. Nat Neurosci, 12(5), 553-558. https://doi.org/10.1038/nn.2295Conchello, J. A., & Lichtman, J. W. (2005). Optical sectioning microscopy. Nat Methods, 2(12), 920-931. https://doi.org/10.1038/nmeth815Dufrene, Y. F. (2008). Towards nanomicrobiology using atomic force microscopy. Nat Rev Microbiol, 6(9), 674-680. https://doi.org/10.1038/nrmicro1948Engström, H., & Engström, B. (1978). Structure of the hairs on cochlear sensory cells. Hearing Research, 1, 49-66.Fettiplace, R. (2017). Hair Cell Transduction, Tuning, and Synaptic Transmission in the Mammalian Cochlea. Compr Physiol, 7(4), 1197-1227. https://doi.org/10.1002/cphy.c160049Fettiplace, R., & Hackney, C. M. (2006). The sensory and motor roles of auditory hair cells. Nat Rev Neurosci, 7(1), 19-29. https://doi.org/doi: 10.1038/nrn1828Frolenkov, G. I., Belyantseva, I. A., Friedman, T. B., & Griffith, A. J. (2004). Genetic insights into the morphogenesis of inner ear hair cells. Nat Rev Genet, 5(7), 489-498. https://doi.org/10.1038/nrg1377Furness, D. N., & Hackney, C. M. (1985). Cross-links between stereocilia in the guinea pig cochlea. Hearing Research, 18, 177-188.Galeano-Naranjo, C., Velez-Ortega, A. C., & Frolenkov, G. I. (2021). Stereocilia Bundle Imaging with Nanoscale Resolution in Live Mammalian Auditory Hair Cells. J Vis Exp(167). https://doi.org/10.3791/62104Gavara, N., Manoussaki, D., & Chadwick, R. S. (2011). Auditory mechanics of the tectorial membrane and the cochlear spiral. Curr Opin Otolaryngol Head Neck Surg, 19(5), 382-387. https://doi.org/10.1097/MOO.0b013e32834a5bc9Goodyear, R. J., Marcotti, W., Kros, C. J., & Richardson, G. P. (2005). Development and properties of stereociliary link types in hair cells of the mouse cochlea. J Comp Neurol, 485(1), 75-85. https://doi.org/10.1002/cne.20513Hadi, S., Alexander, A. J., Velez-Ortega, A. C., & Frolenkov, G. I. (2020). 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EVESURBIFBPUiBMQSBTRUNSRVRBUsONQSBHRU5FUkFMLiAqTEEgVEVTSVMgQSBQVUJMSUNBUiBERUJFIFNFUiBMQSBWRVJTScOTTiBGSU5BTCBBUFJPQkFEQS4gCgpBbCBoYWNlciBjbGljIGVuIGVsIHNpZ3VpZW50ZSBib3TDs24sIHVzdGVkIGluZGljYSBxdWUgZXN0w6EgZGUgYWN1ZXJkbyBjb24gZXN0b3MgdMOpcm1pbm9zLiBTaSB0aWVuZSBhbGd1bmEgZHVkYSBzb2JyZSBsYSBsaWNlbmNpYSwgcG9yIGZhdm9yLCBjb250YWN0ZSBjb24gZWwgYWRtaW5pc3RyYWRvciBkZWwgc2lzdGVtYS4KClVOSVZFUlNJREFEIE5BQ0lPTkFMIERFIENPTE9NQklBIC0gw5psdGltYSBtb2RpZmljYWNpw7NuIDE5LzEwLzIwMjEK

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