Microdomains Displace Laterally and Rotate in the Plasma Membrane of Cochlear Outer Hair Cells

Abstract

Guinea pig outer hair cells (OHC) are cylindrical, with a near constant diameter of ∼8 μm and length ranging from 20 to 90 μm. OHC's lateral plasma membrane is literally packed with motor (prestin) and associated proteins organized in microdomains. These membrane microdomains are connected by hundreds of 25-nm long “pillars” to cytoskeletal microdomains of up to 10 μm2 composed by long, parallel actin filaments cross-linked by shorter spectrin tetramers. Membrane potential-dependent conformational changes in prestin molecules result in reversible changes of cell length (shortening and elongation) following cycle-by-cycle electrical stimulation. We labeled the lateral surface of isolated guinea pig OHCs with polystyrene microspheres (φ=0.5 μm) and, using 1,000 fps high-resolution video recording, investigated their movement simultaneously at the apical, middle and basal region of cells stimulated with a 50 Hz external alternating electrical field. Under electrical stimulation microspheres attached to non-motile cells or the basal region of OHCs didn’t move, whereas those at the middle and apical regions of the OHCs’ lateral wall showed robust back and forth displacements. During stimulation the directions of microspheres’ trajectories changed from random to parallel to the electrical field with angular speeds of up to 6 rad/s, and were back to random after 5 min without stimulation. Microspheres responses were affected by changes in plasma membrane cholesterol levels and cytoskeleton integrity, and by inhibitors of OHC motor response. We concluded that membrane microdomains are able to shift and rotate in the plane of the lateral plasma membrane of cochlear OHCs, and membrane lipid composition as well as the membrane skeleton regulates their dynamics.