Fig. 3
Quantification of CX3CR1 + macrophage density in the cochlea following cochlear implantation. Cochlear implantation was performed in CX3CR1+/GFP Thy1+/YFP mice, fed on chow with 1200 ppm of PLX-5622 (PLX) or control chow (No PLX) for 7 days. Following surgery, mice were continued with respective chow (PLX or No PLX). Starting on post-operative day 7, mice within stimulation cages, connected to the CI processor were stimulated for 5 h per day, 5 days a week. Mice were euthanized at 10, 28 and 56 days post-CI and imaged. Image analysis was performed in IMARIS image analysis software. In 30-µm-thick mid-modiolar sections, CX3CR1 + macrophage cells were counted on maximum intensity z-projections of 3D confocal image stacks. The outline of Rosenthal’s canal (RC) and lateral wall at the base, middle, and apex of the cochlea, scala tympani of the base of the cochlea, and modiolus was traced and the volume of each area was measured. CX3CR1 + macrophages were counted using an automated counting system in IMARIS image analysis software aided by custom-made macros. The density of macrophages with visible, Hoechst + nuclei was calculated per 105 µm3 in each area mentioned. Values derived from every region of the cochlea for an individual animal were averaged together from 3 sections; “n” is the total number of mice used in the study. A–H Density of macrophage in different locations of cochlea in different treatment groups. Number of cochlea analyzed in this study are as follows: at day 10 post-CI, implanted ‘no PLX’ (n = 4), implanted PLX, (n = 3), contralateral ‘no PLX’ (n = 3), contralateral PLX (n = 5); at day 28 post-CI, implanted ‘no PLX’ (n = 6), implanted PLX, (n = 5), contralateral ‘no PLX’ (n = 3), contralateral PLX (n = 4); at day 56 post-CI, implanted ‘no PLX’ (n = 3), implanted PLX, (n = 6), contralateral ‘no PLX’ (n = 3), contralateral PLX (n = 4). Error bars indicate SEM