Section of Physiological Acoustics and Communication, Faculty of Medicine, Eberhard Karls University Tübingen, Tübingen, Germany.
Eur J Neurosci. 2021 Aug;54(3):4755-4767. doi: 10.1111/ejn.15331. Epub 2021 Jul 5.
The plasma-membrane marker FM1-43 was employed to reveal the relative significance of different types of endocytic and transcytic mechanisms in outer hair cells (OHCs) of the guinea-pig cochlea. A double-barrel local perfusion system was used to label independently the apical or synaptic pole of the isolated OHC to study mechanisms of vesicle uptake at the poles and of vesicle trafficking along and across the cell. Treatment with an inhibitor of macropino- and phagocytosis, phenylarsine oxide, or of clathrin-mediated endocytic activity, concanavalin A, significantly reduced the dye uptake at both the apical and the synaptic poles, indicating the presence of both clathrin-independent and clathrin-mediated processes at both poles. However, measurement of uptake speed in the presence of the inhibitors suggested that clathrin-independent processes contribute more extensively to endocytosis at the basal pole than the apical pole. Treatment with an inhibitor of myosin VI, 2,4,6-triiodophenol, significantly delayed both the apicobasal and the basoapical fluorescence signals. However, treatment with an inhibitor of kinesin, monastrol, or of dynein, ciliobrevin D, significantly delayed the signals only in the basoapical direction. The myosinVI inhibitor, but neither the kinesin nor dynein inhibitors, significantly delayed the signals to the subsurface cisternae. That is, myosin VI carries vesicles in both longitudinal directions as well as radially to the subsurface cisternae, whereas kinesin and dynein participate primarily in basoapical trafficking. This fundamental information is essential for elucidating recycling mechanisms of specific proteins involved in establishing, controlling and maintaining the electromechanical action of OHCs and, therefore, is vital for understanding auditory perception.
荧光标记 FM1-43 被用来揭示豚鼠耳蜗外毛细胞（OHC）中不同类型内吞和跨细胞机制的相对重要性。采用双管局部灌流系统分别标记分离的 OHC 的顶极或突触极，以研究极部囊泡摄取和囊泡沿细胞和跨细胞转运的机制。用巨胞饮和吞噬作用抑制剂苯胂酸钠或网格蛋白介导的内吞作用抑制剂伴刀豆球蛋白 A 处理，显著减少了顶极和突触极的染料摄取，表明两种无网格蛋白和网格蛋白介导的过程都存在于两极。然而，在抑制剂存在下测量摄取速度表明，无网格蛋白过程比顶极更广泛地参与基底极的内吞作用。肌球蛋白 VI 抑制剂 2,4,6-三碘苯甲酚处理显著延迟了顶端到基底和基底到顶端的荧光信号。然而，用微管蛋白抑制剂 monastrol 或动力蛋白抑制剂 ciliobrevin D 处理仅显著延迟了基底到顶端的信号。肌球蛋白 VI 抑制剂，但不是微管蛋白或动力蛋白抑制剂，显著延迟了向亚表面质膜下腔的信号。也就是说，肌球蛋白 VI 携带囊泡在纵向和径向两个方向上向亚表面质膜下腔，而微管蛋白和动力蛋白主要参与基底到顶端的转运。这些基本信息对于阐明参与建立、控制和维持 OHC 机电作用的特定蛋白质的再循环机制至关重要，因此对于理解听觉感知至关重要。
