Ultrahigh-frequency hearing-associated freezing behavior is preserved in 1-month-old (1M) Prestin-KO mice. A Paradigm of sound-cue associated freezing behavior. Pure-tone sound at 16 kHz or 63 kHz played by a TDT ES1 (Free Field) electrostatic speaker is used as the conditioned stimulus, and foot-shock was used as the unconditioned stimulus. B Representative examples of locomotion of control mice, 1-month (1M), and 2-month (2M) Prestin-KO mice before (gray, 30 s), during (red, 10 s), and after (blue, 30 s) the pure-tone sound cue. The mice had been trained to pair either the 16-kHz or the 63-kHz cue with the foot-shock-induced freezing. Dots indicate the location of a mouse every 0.5 s. Note that the 2M Prestin-KO mouse reacts to the 16 kHz but not the 63 kHz cue at 90 dB SPL. C Percentage freezing time with the 16-kHz cue at 90 dB SPL. Pre-cue vs post-cue, paired t-test test, Prestin-KO 1-month (1M), t9 = 13.7, ****P <0.0001; Prestin-KO 2-month (2M), t4 = 9.477, ***P = 0.0009; WT, t9 = 27.57, ****P <0.0001. D Percentage freezing time with the 16-kHz cue at 60 dB SPL. Pre-cue vs Post-cue, paired t-test test, Prestin-KO 1M, t4 = 1.0, P = 0.3739; WT, t5 = 3.796, *P = 0.0127. E Percentage freezing time with the 63-kHz cue at 90 dB SPL. Pre-cue vs Post-cue, paired t-test test, Prestin-KO 1M, t9 = 14.07, ****P <0.0001; Prestin-KO 2M, t7 = 0.7977, P = 0.4512; WT, t9 = 15.81, ****P <0.0001. In (C–E), data are presented as the mean ± SD, and N numbers are shown in panels.