Acta Otolaryngol (Stockh) 1989; Suppl. 467: 131-137

CHAPTER 14

Middle Ear Vibration and Sound Pressure Measurements in the Isolated Cochlea Preparation

SHYAM M. KHANNA,’ AKE FLOCK,’ MATS ULFENDAHL’ and WILLEM F. DECRAEMER3 ‘College of Physicians & Surgeons, Columhio University. New York. USA. ‘Deportment of Physiology I I , Karolinska Instituter, Stockholm. Sweden, ond ’Uniwrsitv of Antwerp, Rijksuniwrsituir Centrum Antwerpen. Belgium

INTRODUCTION

A wide variety of auditory functions and responses are measured in terms of the sound pressure at some reference location in the ear canal. Another common measure that has been used to specify the mechanical input to the ear is the vibration of the malleus at the umbo.

These two input measures were utilized in the isolated cochlea preparation of the guinea pigs temporal bone (Ulfendahl, Flock and Khanna. 1989a. b) so that the observed cellular responses could be related to the classical measures of input. In this preparation there are several special conditions that affect the transfer of the input acoustical signal from the ear canal to the organ of Corti. The effect of each of these must be accounted for to correctly relate the cellular vibrations to the acoustical or mechanical input signal. In the isolated cochlea the middle ear structures and a portion of the ear canal were left intact so that acoustical stimulation could be used for measurement of cellular vibration. The temporal bone is immersed to keep the opened turn in the tissue culture medium. As a consequence the middle ear cavity and the medial surface of the tympanic membrane are also immersed in the fluid.

Due to fluid immersion and opening the cochlea in our preparation the following conditions may differ: (i) The sound pressure measured in the ear canal may be different due to the loading of the tympanic membrane by the immersion fluid in the middle ear; (ii) The mechanical characteristics of the middle ear may be altered by the fluid load on the medial surface of the tympanic membrane and by the immersion of the other middle ear structures; (iii) The inner ear impedance as seen by the middle ear may be different since the scala vestibuli is opened at the fourth turn; (iv) For a given middle ear vibration amplitude the alternating pressure in the scala vestibuli may be different due to the opened fourth turn and by the fluid load on the round window in the middle ear cavity. Attempts were made to assess the magnitude of some of these effects.

METHODS

The methods used for the preparation, mounting, observation and vibration measurements in response to a sound stimulus have been described elsewhere (Ulfendahl, Flock and Khanna, 1989a. b; Lund and Khanna, 1989: Willemin, Khanna and Dandliker. 1989; Koester et al., 1989).

Sound pressure was generated in the ear canal using a Sokolich acoustical driver connected to the ear canal with rubber tubing approximately 25 cm in length. The sound pressure was measured using a Sokolich probe microphone system (Sokolich, 1977). Attenuation and phase shift introduced by the tube could not be measured with our present system.

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132 S. M. Khanna et al.

1.0 d UJ

: -

1 . I , , I

Fig. I . Sound pressure level in the ear canal as a function of fre- quency. Electrical input applied to the acoustic system was maintained constant at 1 .O V amplitude. Solid and dashed lines show data collected in two different experiments. The aver- age sound pressure across the frequency range was approxi- mately 0.3 Pa.

SOUND PRESSURE MEASUREMENTS

Sound pressure level was measured in the ear canal as a function of frequency, while constant input voltage ( 1 V) w a s applied to the acoustic transducer. Pressure measure- ments were corrected for the probe tube response. Results from two experiments are shown in Figure 1. The average pressure produced was about 0.3 PdV. For a 1 volt input this corresponded to an average SPL over the frequency range of 83.5 dB. The pressure throughout the frequency range was within +3 dB of this value. The maximum sound pressure level used in our experiments was 97.5 dB (5V input).

The phase of the sound pressure measurements with respect to the electrical input to the driver is shown in Figure 2. The phase lag increases approximately linearly with frequen- cy; the average slope of the phase curve is -0.328"/Hz. This corresponds to a time delay of 9.11 X seconds between the point at which the pressure is measured and the electrical

Fig. 2. Phase of the sound pres- sure as a function of frequency. Solid and dashed lines show phase corresponding to the am- plitude data shown in Figure 1. Phase angles were measured with respect to the phase of the electrical input to the transduc- er. The average slope of the phase curve is -0.32WHz.

FREQUENCY (Hr)

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Middle ear response 133

6 a t; k W

W P z t i n

z + K

>

E! a m

I I , , . I I I . . , I

25 50 100 500 1000 2000

Fig. 3. Vibration amplitude at the tip of the malleus measured as a function of frequency for a I volt input applied to the acoustic system. The middle ear cavity was opened and immersed in the tissue culture medium. The re- sponse is relatively flat below 170 Hz. For frequencies above 250 Hz the amplitude decreases with an average slope of ap- proximately 20 dB per octave.

FREQUENCY (Hz)

input. This delay includes the propagation time of the acoustical signal between the driver and the microphone tip and the phase response of the low-pass filters.

MIDDLE EAR MEASUREMENTS

The middle ear vibrations are normally measured looking from the outer ear canal perpendicularly at the umbo. In our preparation, the ear canal is closed as it is connected to the sound system. Access is available only to the underside of the malleus through the opening made in the bulla cavity and this access does not allow us to look at the malleus perpendicularly. Malleus vibrations can therefore be measured only at an oblique angle from the underside.

The vibration amplitude of the malleus for a I volt input to the acoustic transducer is shown in Figure 3 for three different ears. The three response amplitudes show a shallow

Fig. 4. Phase response corre- sponding to the amplitude re- sponse shown in Figure 3. The average slope of the phase c u m is -0.663'/Hz.

FREQUENCY (Hz)

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- c E t Y

w

; 9 5

z l- 0 a a m >

Fig. 5 . Amplitude of malleus vi- bration before (solid line) and af- ter opening the cochlea (dotted line). Cochlear opening slightly increases the vibration ampli- tude in the region 50 to 300 Hz and in the region 400 to ROO Hz. This suggests a decrease in the cochlear impedance as a conse- quence of opening the scala ves- tibuli in the apical turn.

FREQUENCY (Hz)

resonance peak between 100 and 200 Hz; above 250 Hz the average response drops off rapidly (about -20 dB/oct). On this declining slope three resonances are observed at about 800, 1300 and 1900 Hz. The relative phase between the malleus vibrations and the electrical input signal is plotted in Figure 4. The average slope of the phase curve is -0.663"/Hz. This delay is larger than that measured for the sound pressure at the end of the sound tube (-0.328"/Hz, Figure 2). This additional delay is mainly due to sound propagation through the rubber tube connecting the end of the sound transducer to the cochlea preparation.

Measurements of the vibration amplitude of the malleus tip at 100 dB SPL in the middle ear of a living guinea pig (Manley and Johnstone, 1974: closed-sound system, open-. air filled bulla, perpendicular viewing angle) showed a smooth curve falling off with a slope of about 8 dB/oct above 500 Hz. The average slope observed in our measurements above 250 Hz is much higher. This sharp drop-off may simply be due to the fact that our measure- ments have been made in a viewing direction that is oblique. Changes in the middle ear response with viewing angle have been observed in the middle ear of the living cat (unpublished observations). The other possibility is that the increase in impedance due to the fluid load on the tympanic membrane may cause a steeper roll-off of the amplitude response. The average level of vibration at the frequencies before the steep drop-off in our measurements ( ~ 2 5 0 Hz) is 6.OxlO-' f l a . The data of Manley and Johnstone (1974) correspond to a level of 2 . 2 ~ lo-' &Pa at 100 Hz. This means that both have about the same order of magnitude. It is somewhat suprising that vibration amplitude is higher when the middle ear is filled with fluid than with air.

Phase data on the umbo vibration of the living guinea pig are not available in literature. Therefore we cannot compare our phase data.

EFFECT OF OPENING THE COCHLEA

The bony cap of the cochlea is removed to gain access to the apical turn. and therefore the apical end of the scala tympani is open to the fluid in the test chamber. To investigate if the opening affects the impedance of the cochlea and in turn affects the response measured at

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Middle ear response 135

2.0 I I I l l l ~ l I l ~ l r l ~ l l ~ l " " . l l r l l l l l 1 l " 1 1

1.5 - -

-360 - fs

s w a

Q -720 - J Fig. 6. Phase of the malleus vi-

bration before (solid line) and af- ter opening the cochlea (dotted line). Cochlea opening increases the phase of the response very slightly. The changes are less than 10".

-1000 -

- 1 3 5 ~ 0 " " ' " ' ' " " " " " L " ' ~ ' 500 1000 1500 2Ooo FREQUENCY (Hz)

.5

the malleus, malleus vibrations were measured before and after the opening of the cochlea. To open the cochlea the temporal bone preparation had to be removed from the measuring system and consequently the viewing direction and location of measurement were not exactly the same as before. The results of two measurements are shown in Figure 5 . There is a slight increase in vibration amplitude below 800 Hz. The phase curves in Figure 6 show a slight increase in phase delay. The ratio of vibration amplitude before and after opening the cochlea is shown in Figure 7. The ratio varies between I . 1 and I . 2 . The change in vibration amplitude is of the order of 10%' when the cochlea is opened.

- - Fig. 7. Ratio of the malleus vi- bration amplitude measured be- fore and after opening the co- chlea.

THE EFFECT O F GLUTARALDEHYDE

Glutaraldehyde acts as a cross linking fixative expected to stop active processes and alter passive mechanical properties in the experimental preparation. As part of the inner ear

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1000 1500 50 100 500 FREOUENCY (Hz)

Fig. 8. Effect of adding glutaral- dehyde to the tissue culture so- lution. Solid line shows malleus vibration amplitude vs. frequen- cy before the addition of glutar- aldehyde. Dotted line 18 min- utes after the addition. Dashed line 74 minutes after the addi- tion, and dashed dotted line af- ter 150 min. Addition of glutaral- dehyde stiffens the middle ear system and reduces the vibra- tion amplitude. Below 400 Hz the amplitude continues to de- crease with time. Reduction is by a factor of nearly 6. Above 300 Hz changes in vibration modes of the tympanic mem- brane produce maxima and minima in the malleus response.

experiments, glutaraldehyde treated preparations were used to establish baseline proper- ties of the cellular response in the absence of active processes (Flock, Khanna and Ulfendahl, 1989). The effect of adding glutaraldehyde to the tissue culture solution (2.5 % concentration) on the middle ear response is illustrated in Figure 8. Measurements were made before adding glutaraldehyde (solid line) and then 18 min (dotted line), 74 min (dashed line), and 150 min (dashed-dotted line) following the addition. As a result of the stiffening effect of the glutaraldehyde the vibration amplitude of the malleus decreases with time while most of the resonances are shifted to slightly higher frequencies. At higher frequencies supplementary resonances are introduced. The reduction in middle ear vibra- tion amplitude will affect the magnitude of the nonlinear components of the micromechani- cal response. This reduction in stimulus level should be taken into account when interpret- ing the results of glutaraldehyde experiments (Flock, Khanna and Ulfendahl, 1989).

rn W W U B Fig. 9. Phase of the malleus re-

sponses corresponding to the amplitude responses shown in Figure 5 . Same symbols are used to indicate time. The phase lag below 600 Hz and above 850 Hz is decreased by the addition of glutaraldehyde.

FREOUENCY (Hz)

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Middle eur response 137

The phase of the middle ear response is also affected by addition of glutaraldehyde. Responses before and after addition are shown in Figure 9. Supplementary resonances make the curve look much less smooth, the overall phase delay is generally reduced.

CONCLUSIONS

Immersing the temporal bone preparation in tissue culture medium causes the bulla cavity to be filled with liquid. The measurements of the malleus tip vibration under this condition were carried out to test whether the applied sound pressures produce a mechanical input to the middle ear comparable to that obtained when the middle ear is filled with air. For frequencies below 750 Hz the mechanical input is within + I S to -10 dB of a normal middle ear response.

Opening the cochlea affects the middle ear vibrations only slightly. The increase was about 10%.

The impedance at the oval window consists of at least two components; (i) the imped- ance due to the footplate of the stapes and its suspending annular ligament, and (ii) the cochlear impedance. At low frequencies the stapes components are dominant. It is thus not possible to assess from our present measurements the change in the cochlear imped- ance due to the opening of the cochlea.

The main conclusion from these experiments is that the change in transmission charac- teristics from the ear canal to the stapes in the working range of frequencies below 1 kHz is quite modest and therefore the sound pressure levels used in our experiments are compa- rable to those used in an intact living ear.

ACKNOWLEDGEMENTS This reasearch was supported by a program project grant NS22334 from NlDCD and by the Emil Capita Foundation.

REFERENCES Flock A. Khanna SM. Ulfendahl M (1989). Effects of glutaraldehyde and metabolic inhibitors on the

vibratory responses in the isolated cochlea. Acta Otolaryngol (Stockh) Suppl 467: 20-219. Koester CJ. Khanna SM. Rosskothen H, Tackaberry RB (1989). Incident light optical sectioning

microscope for visualization of cellular structures in the inner ear. Acta Otolaryngol (Stockh) Suppl 467: 27-33.

h n d DT. Khanna SM ( 1989). A digital system for the generation of acoustic stimuli and the analysib of cellular vibration data. Acta Otolaryngol (Stockh) Suppl 467: 77-89.

Manley GA. Johnstone BM (1974). Middle-ear function in the guinea pig. J Acoust SOC Am 56:

Sokolich WG (1977). Improved acoustic system for auditory research. J Acoust SOC Am 62: S12. Ulfendahl M. Flock A, Khanna SM (1989a). A temporal bone preparation for the study of cochlear

Ulfendahl M. Flock A. Khanna SM ( 1989b). Isolated cochlea preparation for the study of cellular

Willemin JF. Khanna SM. Dandliker R ( 1989). Heterodyne interferometer for cellular vibration

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micromechanics at the cellular level. Hearing Res 40: 55-64.

vibrations and motility. Acta Otolaryngol (Stockh) Suppl 467: 9 1-96,

measurement. Acta Otolaryngol (Stockh) Suppl 467: 3542.

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