Different rates of endocytic activity and vesicle transport from the apical and synaptic poles of the outer hair cell

Abstract

Background. Intense endocytic activity at the apex of outer hair cells (OHCs)-the electromechanical cells of the cochlea-has been demonstrated using the vital plasmamembrane marker FM1-43 and confocal laserscanning microscopy. Vesicular traffic toward the cell nucleus to distinct locations of the endoplasmic reticulum has also been shown. Objective. The current study characterizes the dynamics of endocytic activity, as well as apicobasal and basoapical trafficking, using a local perfusion technique that we recently developed and published to visualize bidirectional trafficking in isolated bipolar cells. Materials and methods. The fluorescent plasma-membranemarkers FM1-43 (10 µM) and FM4-64 (10 µM), together with a fluidphase marker, Lucifer yellow (50 µM), were used to label endocytosed vesicles in isolated OHCs of the guinea pig cochlea. Targets of endocytosed vesicles were examined with a fluorescentmarker of subsurface cisternae, DiOC6 (0.87 µM). Single-and two-photon confocal laser-scanningmicroscopy was used to visualize labeled vesicles. Results. The plasma-membrane markers presentedmore intense vesicle internalization at the synaptic pole than at the apical pole of the OHC. Intracellular basoapical vesicle trafficking was faster than apicobasal trafficking. Vesicles endocytosed at the synaptic pole were transcytosed to the endoplasmic reticulum system. An intracellular Lucifer yellow signal was not detected. Conclusion. The larger endocytic fluorescent signals in the synaptic pole and the faster basoapical trafficking imply that membrane internalization and vesicle trafficking aremore efficient at the synaptic pole than at the apical pole of the OHC.