**5.1 Frequency specificity in the Medial Olivocochlear System (MOCS)**

It is now well established that the sensitivity and frequency tuning of the peripheral auditory system is influenced by the cochlear efferent neural pathways (Guinan Jr, 2006). Activation of the MOCS by acoustic stimulation of the contralateral ear has been shown to suppress sensitivity of the cochlea, for example by reduction in cochlear nerve action potential amplitude (Fex, 1962). It is considered that this effect is mediated by suppression of the cochlear amplifier effect of OHC activity (Siegel & Kim, 1982). It is likely that relatively specific stimulus conditions are required for efferents to play a role in hearing (M. C. Liberman, 1988), but despite intensive investigation, the nature of this role remains unclear. Further assessment of how the MOCS is activated by different stimuli should improve understanding of this issue (Maison, Micheyl, Andéol, Gallégo, & Collet, 2000).

Tonotopicity of the MOCS has been clearly demonstrated in recordings from single olivocochlear fibers in the cat and guinea pig (Brown, 1989; Cody & Johnstone, 1982; M. Liberman & Brown, 1986). In these studies, efferent neural tuning curves were derived by measuring firing rate in response to contralateral tones of different frequency, and were found to have a shape and sharpness similar to cochlear afferent tuning curves. In addition, horseradish peroxidase injection was used to reveal the projection of some fibers, and in all cases they terminated on OHCs at a cochlear position where afferent

Contralateral Suppression of Otoacoustic Emissions:

Canadian Council on Animal Care.

**5.1.1.3 Contralateral stimulus** 

**5.1.1.4 Analysis of results** 

variable).

**5.1.1.2 Real time DPOAE measurement** 

Working Towards a Simple Objective Frequency Specific Test for Hearing Screening 35

Xylazine 1.3mg/kg). Five animals were studied twice, typically with an interval of >4 weeks between recording sessions. Thus in total, 15 recording sessions were completed. All studies were approved by the local Animal Care Committee, following the guidelines of the

DPOAEs were measured in real time with a Vivo 600 DPR device (Vivosonic Inc., Toronto, ON). In contrast to conventional OAE techniques which employ signal averaging to extract the signal from noise, this technique uses digital filtering and signal modeling. The continuous real-time signal is ideally suited to the detection of changes in OAE amplitude, such as those produced by contralateral stimuli (James et al., 2005). Primary frequencies were set at f2/f1= 1.22 for values of f2 between 1.6 and 8.0 kHz, with intensities of L1 = 70dB and L2 = 65dB. DPOAEs were measured at 2f1-f2. The OAE probe, in a conforming soft plastic cuff, was inserted into the external auditory meatus by straightening the soft tissues to allow the probe to abut the lateral aspect of the bony meatus (approximately 13mm from the tympanic membrane). Multiple recordings of up to three minutes duration were made in each session. All recordings were made in a sound-attenuating booth. The DPOAE probe was calibrated in the ear canal by the device and calibration confirmed in a 2ml coupler using an SR760 FFT Spectrum Analyzer (Stanford Research Systems, Sunnyvale, CA) and a

precision CR: 511D Acoustic Calibrator (Cirrus Research plc, North Yorkshire, U.K.).

return to pre-stimulus levels (typically > 300ms longer than CAS duration).

An intermittent pure tone stimulus was applied to the contralateral ear using an ER-2 transducer with a foam ear-insert (Etymotic Research Inc., IL). 60 different CAS frequencies were tested between 0.6 – 17 kHz. Sweep direction from high to low, or low to high frequency of contralateral stimulation was changed between sweeps to control for any gradual drift in DPOAE level that might occur during a recording period. CAS intensity was set at 50 dB SPL as a previous study had shown the threshold for a response to be around 30dB SPL while acoustic cross talk occurred at intensities of ≥70 dB SPL (using noise floor measures and recordings in cadaveric chinchilla). Stimulus duration was set at 0.5s with rise / fall times of 4 ms. The interval between stimuli was long enough to allow DPOAE levels to

DPOAE signals were recorded in real time, and level changes occurring in synchrony with contralateral stimulation were noted. Subsequent analysis was performed on the recorded real time trace and on averaged data, using VivoAnalysis software (Vivosonic Inc., ON), based on LabVIEW 5.1 data acquisition software (National Instruments, TX). Averaging was synchronized with the start of the CAS and was used to smooth the data and remove nonsynchronous or spontaneous variation in the DPOAE signal. Averaged data were used to measure the magnitude of the DPOAE response to CAS from the baseline (no contralateral stimulation condition) to maximum OAE change (i.e. at asymptotic level). Frequency response curves to indicate tuning of contralateral suppression were plotted with magnitude of suppression (dependent variable) versus frequency of CAS tone (independent

neurons have a characteristic frequency (CF) similar to that measured in the cochlear efferent.

Frequency specificity of MOCS activity can also be detected when recording the response of inner hair cells and auditory nerve fibers to acoustic stimulation. For example in cats, the response of single cochlear afferent fibers to tone pips is suppressed by simultaneously applying tone pips to the contralateral ear. This suppression is maximal when the contralateral tone is similar to the characteristic frequency of the afferent fiber (Murata, Tanahashi, Horikawa, & Funai, 1980; Warren III & Liberman, 1989a; Warren III & Liberman, 1989b). Similarly, when recording the compound action potential induced by tone pips with a round window electrode, maximum suppression is induced by contralateral tone pips of similar frequency (M. C. Liberman, 1989).

As OAEs are generated by OHC activity, they may provide a more direct and non-invasive insight into the effect of the MOCS on its target cells than neural recordings. In human subjects, suppression of spontaneous OAEs is maximal with a CAS tone at a frequency close to the spontaneous OAE (Mott, Norton, Neely, & Bruce Warr, 1989). In addition to suppression, a frequency shift of spontaneous OAEs is caused by CAS and interestingly this is maximal with a CAS about 3/8 to 1/2 octaves below the spontaneous OAE frequency. OAEs evoked by tone pips can be suppressed by contralateral narrow band noise, suppression being maximal with CAS frequencies close to the frequency of the tone pip (Veuillet, Collet, & Duclaux, 1991).

Contralateral suppression of OAEs has not been widely used to investigate MOCS frequency specificity in animal models. A systematic study in the barn owl produced frequency response functions in which DPOAE suppression was plotted as a function of CAS frequency (Manley, Taschenberger, & Oeckinghaus, 1999). This showed maximal suppression with CAS similar to primary frequencies. Extrapolation of these findings to other models is limited by the variability of DPOAE levels and the additional types of efferent fiber which are present in birds.

The purpose of the present study was to investigate the frequency specificity of the MOCS in the chinchilla. In this species there has been a report of difficulty in detecting MOCS change in response to contralateral stimulation (Azeredo et al., 1999). On the other hand, electrical stimulation of the olivocochlear bundle in the floor of the fourth ventricle elicits OAE suppression (Siegel & Kim, 1982). In our present study, the suppressive effect on DPOAEs of contralateral pure tone stimuli is investigated with real-time recording of the DPOAE.
