Spiral Ganglion Cell Density Analysis

The rats were sacrificed at W1 and W2 after click stimulation; then, left cochlear sectioning was performed along the paramodiolar axis, followed by hematoxylin/eosin staining. After the aforementioned fixation procedure was executed, the cochleae were decalcified in 10% ethylene-diamine tetra-acetic acid (EDTA) for 4 weeks at 4°C, dehydrated, embedded in paraffin, and sectioned serially (4 μm thick) parallel to the modiolar axis [20 (link)]. The specimens were observed with the spiral ganglion cells counted under a light microscope (Olympus BX51) and photographed (Olympus U-25ND6), and the digital images were saved. After the image collected, we used ImageJ software to encircle the exterior of spiral ganglion cells, then calculate the density of spiral ganglion cells by defining control group as 100%, and then calculate it for other groups proportionally.