Fig. 4. Development of the afferent innervation in the cochlea during neurite reorganization (P0-P6) in mice. TMRD- and peripherin-labeled neurites undergo refinement (P0-P3) and retraction (P3-P6), as demonstrated by maximum intensity projections of confocal images from cochlear whole-mount preparations. (A,D,G) TMRD labeling (red) of type I afferent nerve fibers. These neurites innervate the IHCs between P0 and P6. Peripherin (green) immunolabeling of type II afferent nerve fibers (B,E,H) and merged images (C,F,I) demonstrate the independence and the relationship between TMRD and peripherin labeling. (A-C) At P0, both TMRD- and peripherin-labeled neurites travel along the OHCs in an unorganized pattern. (D-F) Both TMRD- and peripherin-labeled neurites formed three rows of outer spiral bundles beneath the OHCs in an organized pattern. The inner spiral plexus, associated with the IHCs, exhibited only TMRD labeling, but not peripherin labeling, at P3. (G-I) TMRD labeling was reduced under the OHCs compared with the strong representation of peripherin-expressing neurites in the outer spiral bundle region. Scale bars 10 µm. SGNI, type I spiral ganglion neurons; SGNII, type II spiral ganglion neurons.