Sound-evoked olivocochlear activation in unanesthetized mice

Abstract

Genetic tools available for the mouse make it a powerful model to study the modulation of cochlear function by descending control systems. Suppression of distortion product otoacoustic emission (DPOAE) amplitude by contralateral acoustic stimulation (CAS) provides a robust tool for noninvasively monitoring the strength of descending modulation, yet investigations in mice have been performed infrequently and only under anesthesia, a condition likely to reduce olivocochlear activation. Here, we characterize the contralateral olivocochlear reflex in the alert, unanesthetized mouse. Head-fixed mice were restrained between two closed acoustic systems, while an artifact rejection protocol minimized contamination from self-generated sounds and movements. In mice anesthetized with pentobarbital, ketamine or urethane, CAS at 80 dB SPL evoked, on average, a <1-dB change in DPOAE amplitude. In contrast, the mean CAS-induced DPOAE suppression in unanesthetized mice was nearly 8 dB. Experiments in mice with targeted deletion of the α9 subunit of the nicotinic acetylcholine receptor confirmed the contribution of the medial olivocochlear efferents to this phenomenon. These findings demonstrate the utility of the CAS assay in the unanesthetized mouse and highlight the adverse effects of anesthesia when probing the functional status of descending control pathways within the auditory system.

FIG. 1.

Movement artifacts during DPOAE measurements in unanesthetized mice. A Animal movement introduced artifacts (lighter points) in the primary (filled triangles) or noise floor (open triangles) levels. B Percentage of discarded trials for each animal tested in the unanesthetized case, before dynamic recalibration. Values represent the mean of three runs, once at each tested frequency. C Example mouse in which runs with (hatching) and without dynamic recalibration were interleaved. Dynamic recalibration decreased the percentage of discarded trials (dotted line) without affecting the maximum CAS-induced DPOAE suppression (solid line).

FIG. 2.

Amount of CAS-evoked DPOAE suppression depends on state of arousal. A Contra-noise suppression of the DPOAE (f₂ = 16 kHz, L₂ = 25 dB SPL) under ketamine/xylazine anesthesia. Gray box indicates timing of contralateral broadband noise presentation (80 dB SPL). B Contra-noise suppression of DPOAE from the same mouse before anesthetization. The same stimulus conditions are used for (A) and (B). C Mean change in DPOAE amplitude relative to the pre-CAS baseline (n = 8 for unanesthetized and ketamine/xylazine groups, n = 4 for urethane–xylazine and pentobarbital groups). Unanesthetized and ketamine/xylazine groups were the same eight animals. In (C) only, each point is an average of five consecutive time points; shaded areas indicate SEM.

FIG. 3.

CAS-evoked DPOAE suppression is stronger and more stereotyped in unanesthetized mice. A Dot density graph of the maximum CAS-induced DPOAE change in individual mice at three ipsilateral f₂ frequencies. Pre- and post-CAS refer to measurements made at f₂ = 16 kHz before and after CAS, respectively, to indicate fluctuations in baseline DPOAE amplitude. B Absolute values of CAS-induced maximum DPOAE change in individual mice shown as open circles; bars represent the median. C Absolute value of the maximum CAS-induced DPOAE suppression during the first and second halves of noise presentation for each mouse.

FIG. 4.

CAS-induced DPOAE suppression in the unanesthetized mouse is largely, though not completely, mediated by the MOC system. A Example trace from an unanesthetized α9^−/− mouse (f₂ = 16 kHz, L₂ = 35 dB SPL, CAS level = 80 dB). B Example trace from an unanesthetized α9 ^+/+ mouse (f₂ = 16 kHz, L₂ = 33 dB SPL, CAS level = 80 dB). Missing trials were discarded due to movement artifact. C Maximum DPOAE suppression during CAS in CBA/CaJ mice, a9^+/+ mice, and a9^−/− mice, all in the unanesthetized condition.