Problems in Cochlear Micromechanics

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The motor protein prestin is presumed to densely populate the plasma membrane of the outer hair cells in the mammalian cochlea. Older electron microscopy (EM) studies indicated the presence of a dense array of particles with uniform size of about 10nm in diameter. It was presumed that these must be the motor proteins, but no identification was ever performed. Here, a novel cell-free protocol for imaging outer hair cell native plasma membranes was developed. We used an atomic force microscope (AFM) to image the intracellular domain of a plasma membrane that was attached to a mica substrate under physiological conditions. Imaging revealed the presence of a dense array of particles whose size was consistent with those observed by EM. In addition, the particles were organized in a specific pattern, rather than a random distribution. The pattern allowed speculation as to the oligomeric state of the protein in-situ. The particles were unambiguously identified by the introduction of anti-prestin antibodies that bound to the protein particles while maintaining their organizational patterns.