Figure 3

Changes of euchromatin/heterochromatin ratios and evidence of histone deacetylation in the glial cells of the injured auditory nerve.

(a) Alterations of nuclear ultrastructure in glial cells within Rosenthal’s canal (RC) and the osseous spiral lamina (OSL). In the control ears, glial cells (white arrows), which wrap around the perikarya of type I neurons and their processes, had nuclei with prominent electron-dense heterochromatin (patches of dark coloration). 1–3 days after ouabain exposure, glial cell nuclei exhibit less heterochromatin but more of the lightly colored euchromatin (black arrows). (b) A schematic diagram showing the area of the OSL and RC in the peripheral mouse auditory nerve. (c) Quantitative analysis of nuclear subdomains showed a significant decrease of heterochromatin percentage in the glial cell nuclei of injured auditory nerves (n = 6 per group) as compared to controls (n = 11 per group; Student’s unpaired t-test; *p < 0.05; **p < 0.01). (d) Injured auditory nerves showed a decline in HDAC 1 (top row) and HDAC2 (bottom row) immunoreactivity. The majority of glial cells in control auditory nerve were positive for HDACs (green), whereas numerous glial cells within an injured auditory nerve were negative for HDAC (arrows) or showed a reduced level of HDAC (arrow heads) in the cytoplasm. Right panels show the enlarged images of the boxed areas. (e) Quantitative analysis of HDAC expressed cells were conducted at 3 days after ouabain exposure (n = 3–4 per group; Student’s unpaired t-test; *p < 0.05). Note that that glial cells showed only a modest reduction in the amount of HDAC2⁺ cells but a larger reduction in HDAC1⁺ cells after ouabain exposure for 3 days. Scale bars, 2 μm in (a); 10 μm in (d).