Exaggerated Startle Reflex to Low Frequency Tone Bursts in C57BL/6J Mice with Progressive Hearing Loss: A Longitudinal Aging Study
Jim Ison, Peter Rivoli, Jason Moore, and Paul Allen
Brain and Cognitive Sciences, University of Rochester, Rochester NY 14627 USA
30
20
Late ASR (v-units)
24 kHz
1400
1200
1000
600
200
0
0
12
16
20
24 28 32 36
Age (weeks)
40
44
48
Figure 2: At Week 6 ABR thresholds (3 to 48 kHz in order) were 51;
27; 8; 25; and 26 dB SPL. The overall loss of sensitivity appears to
result from two linear processes, the first having a slope of less than
1 dB/week, the second a slope of 3 dB/week or more; however the
calculated slope of the second process is in part an artifact of its
onset being variable across mice (see individual mice, Figure 5)
Loss of 90 dB ABR P1/N1 amplitude from
6 to 28 weeks of age, across frequency
6
5
10
8
8
12
16
20
24 28 32 36
Age (weeks)
2000
1500
ASR 6
ABR 12
ABR 24
ABR 48
100
80
60
1000
40
500
20
S#973
4
0
2
0
10
0
-2
3
4
5
Latency (ms)
6
3
2
1
6 8 10 12 14 16 18 20 22 24 26 28
Age (weeks)
Figure 3: Note parallel changes in P1/N1 amplitude, compared to
threshold changes. Neither ABR thresholds nor amplitudes correlated
with the early ASR across mice. (Inset is an example of the replicated
12 kHz 90 dB ABR waveform at 6 weeks of age in S#973)
44
48
52
20
30
40
Weeks of age
2000
100
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1500
60
1000
40
500
20
S#982
0
0
10
20
30
40
Weeks of age
50
3000
2500
2000
100
80
60
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20
S#976
0
10
50
ASR 6
ABR 12
ABR 24
ABR 48
ASR 6
ABR 12
ABR 24
ABR 48
4000
3500
3000
2500
2000
1500
1000
500
0
20
30
40
Weeks of age
50
ASR 6
ABR 12
ABR 24
ABR 48
100
80
60
1500
40
1000
20
500
S#980
0
0
10
20
30
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Weeks of age
ABR Threshold (dB SPL)
3k, 90dB
6k, 90dB
12k 90dB
24k, 90dB
32k, 90dB
48k, 80dB
40
Figure 4: At 10 weeks of age the most vigorous ASR is for 12 kHz,
as might be expected from the ABR thresholds and amplitudes. The
ASR to 3 and 6 kHz, relative to 24 kHz, is better than their relative
ABR levels. The ASR first declined across all frequencies, similar to
the Figure 2 ABR, but then increased beyond Week 23 for 3 kHz
and 6 kHz stimuli (similar effects were obtained for lower ES levels).
6
2
4
4
52
ASR (v-units)
8
ASR (v-units)
4
ASRL = ASRE+600 (R2 = 0.49) *
A
0
400
10
2000
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1600
1400
1200
1000
800
600
400
200
0
800
50
Figure 5: Data from four mice - note amplitude gain from early to late
ASR, and also that 24, 32, and 48 kHz ABR losses always preceded
ASR gain, while ASR gain always preceded 12 kHz ABR losses.
ASR Residual (v-units)
40
1600
300
200
200
400
600
800
Early ASR (v-units)
1000
r = +0.60*
100
0
-100
-200
B
-300
46 48 50 52 54 56 58 60
Mean Early 24 to 48 kHz ABR (dB SPL)
ASR Res./Residual (v-units)
ASR Amplitude (V-units)
50
12 kHz
ABR Threshold (dB SPL)
ABR stimuli and recording: Beginning at 6 or 8 weeks of age, and
continuing to 54 weeks, fortnightly ABR tests (and DPOAE tests, not
reported here) were interspersed with ASR tests. Measurements were
made from Avertin anaesthetized mice using needle electrodes at the
vertex and mastoid, and hind limb. Tone-pip stimuli (5ms; 3, 6, 12, 24, 32,
48 kHz; 10-90 dB SPL) were generated and ABR waveforms measured
(11/s, duplicates of 100 reps) using TDT System II and 3 DSP
workstations in conjunction with BioSig. Stimulus level was reduced by
10 dB then the threshold (visible 4.5 complex) bracketed in 5 dB steps.
60
1800
ABR Threshold (dB SPL)
Figure 1: The mouse is
shown in the test cage
located directly under the ES
speaker. The pedestal fits
onto a stiff platform to which
an accelerometer is attached.
70
6 kHz
ABR Threshold (dB SPL)
Startle stimuli and recording: The mice
were held in a wire cage mounted to a
platform over an accelerometer (See
Figure 1). Startle eliciting stimuli (ES)
were delivered from overhead: 15 ms
duration + 5 ms rt/ft; 0, 60, 70, 80, 90,
100, 110 dB; 3, 6, 12, and 24 kHz; 8 trials
each condition; at 15 s intervals.
80
2000
ASR (v-units)
Subjects: C57BL/6J mice (n = 12
survivors of 16) were tested fortnightly
from 10 weeks to 52 weeks of age (ASR),
from 6 to 54 weeks (ABR).
90
Initial ASR correlates with later ASR: and ABR
threshold losses correlate with the residuals
3 kHz
ASR (v-units)
Methods
3 Khz [ABR(dB 3kHz) = -0.7 x Weeks]
6 kHz
12 kHz [ABR(dB 12kHz) = -2.9 x Weeks]
24 kHz
32 kHz
48 kHz
ABR Potential (micro-V)
The C57BL/6J mouse cochlea progressively degenerates beginning at
about 2 months of age. Some data suggest a small initial functional
deficit across the spectrum (and see Figure 2), but the major loss in
sensitivity appears first for high frequency hearing then gradually
spreads to lower frequencies (see Figure 2 again, and also Figure 3). In
a prior between-group study (JARO, 2003) the loss of high frequency
hearing (ABR thresholds) was associated with a progressive increase in
the amplitude of the acoustic startle reflex (ASR) to low-frequency tone
bursts even as the ASR to high-frequency tones declined. Here we
replicate these findings in a 10-month long longitudinal design that
allows better tracking of the within-subject association of ABR measures
of hearing sensitivity with increased low-frequency ASR. This
experiment confirms the prior data in its showing that individual
differences in hyper-reflexia are related to the degree of hearing loss,
and extends them in showing that the effect is present in every mouse.
The strength and uniformity of hyper-reflexia across mice suggests the
utility of this animal model for studying the functional effects of the
neural reorganization that accompanies hearing loss in C57 mice.
Initial loss in 110 dB ASR in all frequencies,
then gains at 3 and 6 kHz beyond 23 weeks
Loss of ABR thresholds from 6 to 54 weeks
of age, across frequency
ABR Potential (micro-Volts)
Overview
Mean (SEM) ABR Threshold Increment (dB SPL)
ARO 2005
#430
200
100
r = +0.61*
0
-100
-200
-300
C
-400
30
35
40
45
50
55
Mean Late 12 kHz ABR (dB SPL)
Figure 6: In A, the late (>23 wk) ASR levels (3 & 6 k, 100 & 110 dB)
correlate with the early (10-12 wk) ASR level. This accounts for about
50% of the late ASR variance between mice. Next, in B, the overall
early (10-30 wk) high frequency ABR thresholds (24-48 kHz) account
for 36% of the residual variance in the late ASR left over from A.
Finally, in C, the late (> 30 wk) 12 kHz ABR threshold accounts for
about 36% of the variance in the residual late ASR variance left over
from B. These data support the hypothesis that low-frequency ASR
hyperreflexia results from hearing loss at higher frequencies .
jison@bcs.rochester.edu; Supported by NIA Grant #AG09524