Problems in Cochlear Micromechanics

运动蛋白prestin被认为密集地分布在哺乳动物耳蜗外毛细胞的质膜上。较早的电子显微镜（EM）研究表明存在密集的颗粒阵列，直径约为10nm。据推测，这些一定是运动蛋白，但从未进行过鉴定。本文提出了一种新的无细胞外毛细胞原生质膜成像方法。我们使用原子力显微镜（AFM）对生理条件下附着在云母基质上的质膜的胞内结构域进行了成像。成像显示存在密集的颗粒阵列，其大小与EM观察到的一致。此外，颗粒以特定的模式组织，而不是随机分布。这种模式可以推测蛋白质的原位低聚状态。通过引入抗prestin抗体，这些颗粒被明确地识别出来，这些抗体与蛋白质颗粒结合，同时保持其组织模式。

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.