Fig. 5: Reduced vesicle replenishment and slowed endocytic membrane retrieval in Otof-ΔC IHCs.

a Representative trace collected from the double-pulse stimulation revealed a notable ΔC_m after two consecutive 20 ms depolarizations. The ratio of ΔC_m2/ΔC_m1 was calculated to quantify synaptic vesicle replenishment. Significantly reduced ΔC_m2/ΔC_m1 was found in Otof-ΔC IHCs at 50 and 100 ms only (lower). b A train of twenty successive short depolarization pulse (duration 5 ms, interpulse interval 10 ms) induced C_m increments (upper panel) with IHCs from Otof-ΔC mice showed a cumulative decline in ΔC_m. c Endocytosis was assessed by determining the decrease in C_m after 200 ms depolarizing pulses and fitting with a linear function. Otof-ΔC IHCs showed a slower linear Cm recovery after a 200 ms depolarization. d Comparison of ABR wave I amplitudes with different numbers of averaging repeats showed no change in WT but a reduction in wave I amplitude in Otof-ΔC mice, suggesting a slower recovery in triggering auditory nerve fiber firing. e Increments from 50 to 12800 ABR averages were applied to a fixed stimulus condition of 16 kHz, 90 dB SPL. As the number of averages increased, a cumulative reduction in the ABR wave I amplitude recapitulated the train pulse stimulus results from IHC patch-clamp recordings. Age-matched and sex-matched littermate WT controls were used for all experiments. Statistical analysis by two-side unpaired t test or Mann–Whitney test with significance indicated (a, c) and two-way ANOVA followed by the Bonferroni post hoc test with significance indicated (b, e). All data, statistical test used and p values can be found in the source data file.