Baseline and Endpoint auditory tests (ABR and DPOAE): Macrophage ablation followed by partial repopulation protected against cisplatin-induced hearing loss and OHC dysfunction (Experiment 1) (Manuscript: Fig. 2; Fig. S2-1, Fig. S2-2_Supp Fig 5-3)

Prior to the start of PLX3397 chow pre-treatment and at the end of the cisplatin administration protocol, mice underwent two types of auditory tests: auditory brainstem responses (ABRs; measure hearing sensitivity) and distortion product otoacoustic emissions (DPOAEs; indirectly measure outer hair cell function). These data suggest that partial macrophage ablation protected against cisplatin-induced hearing loss and OHC dysfunction (Experiment 1).For ABR measurements, subcutaneous needle electrodes (Rhythmlink, Columbia, SC, USA) were placed behind left pinna of the test ear (reference), vertex (active), and near the tail of the mouse (ground). Tone-burst stimuli (Cos2, 3 msec, 0.5 msec rise/fall in alternating polarity) were presented at a rate of 29.9/sec at 8, 11.2, 16, 22.4, 32, and 40 kHz starting at 90dB SPL. At each sound level, 1024 waveforms were averaged, amplified (20x), and filtered (HP: 300Hz, LP: 3kHz, NT: 60Hz). At near-threshold sound pressure levels (SPL), the ABR waveforms were recorded twice, and two waveforms were superimposed for comparison. ABR threshold was defined as the lowest stimulus intensity that resulted in a reproducible waveform displaying identifiable peaks.DPOAEs were measured in response to two primary pure tones, f1 and f2, generated by Multi Field 1 speakers. Two primary tones were presented at 6 frequency pairs, where f2 corresponded to ABR test frequencies (f2 = 8, 11.2, 16, 22.4, 32, 40kHz; f2/f1 = 1.25). The sound level was increased in 5dB steps from 30dB to 90dB. At each sound level, 512 responses were averaged. DPOAE at 2f1-f2 was recorded in the mouse inner ear canal using an ER-10B+ microphone (Etymotic, Elk Grove Village, IL, USA) connected to a modified pipette tip to fit the mouse external ear canal. Biological noise floors and amplitudes were calculated for each treatment group and plotted relative to each other.在PLX3397饲料预处理启动前及顺铂给药方案结束时，小鼠接受了两类听觉测试：听性脑干反应（auditory brainstem responses, ABR，用于评估听力灵敏度）与畸变产物耳声发射（distortion product otoacoustic emissions, DPOAEs，间接反映外毛细胞（outer hair cell, OHC）功能）。本实验数据表明，部分巨噬细胞清除可减轻顺铂诱导的听力损失与外毛细胞功能损伤（实验1）。 针对听性脑干反应的检测，将皮下针电极（Rhythmlink，美国南卡罗来纳州哥伦比亚市）分别放置于受试耳左耳耳廓后方（参考电极）、颅顶点（活动电极）以及小鼠尾部附近（接地电极）。采用升余弦波形声短突发刺激（时长3ms，升降沿0.5ms，极性交替），以29.9次/秒的速率呈现，测试频率涵盖8、11.2、16、22.4、32及40kHz，初始声压级设为90dB SPL。每个声压级下，对1024个波形进行平均、放大（20倍）及滤波（高通300Hz，低通3kHz，陷波60Hz）。在近阈值声压级下，重复记录两次ABR波形，并将两个波形叠加以进行比对。ABR阈值被定义为可引出可重复且具有可识别峰的最低刺激强度。 畸变产物耳声发射的检测采用Multi Field 1扬声器生成的两个主纯音f1与f2完成。设置6组频率对，其中f2与ABR测试频率一致（f2=8、11.2、16、22.4、32及40kHz；f2/f1=1.25）。声压级以5dB为步长从30dB递增至90dB，每个声压级下对512次响应进行平均。采用连接定制移液管尖端以适配小鼠外耳道的ER-10B+麦克风（Etymotic，美国伊利诺伊州埃尔克格罗夫村），在小鼠内耳道中记录2f1-f2成分的畸变产物耳声发射。为每个处理组计算生物学本底噪声与振幅，并进行组间相对绘图分析。