Reversing pathological neural activity using
Navzer D. Engineer1,2, Jonathan R. Riley1, Jonathan D. Seale1, Will A. Vrana1, Jai A. Shetake1, Sindhu P. Sudanagunta1,
Michael S. Borland1& Michael P. Kilgard1
Brain changes in response to nerve damage or cochlear trauma can
for many types of chronic pain and tinnitus1–3. Several studies have
reported that the severity of chronic pain and tinnitus is correlated
tions5. However, there is as yet no direct evidence for a causal role of
plasticity in the generation of pain or tinnitus. Here we report evid-
ence that reversing the brain changes responsible can eliminate the
Exposure to intense noise degrades the frequency tuning of auditory
cortex neurons and increases cortical synchronization. Repeatedly
pairing tones with brief pulses of vagus nerve stimulation completely
eliminated the physiological and behavioural correlates of tinnitus in
end of therapy. This method for restoring neural activity to normal
may be applicable to a variety of neurological disorders.
Damage to the peripheral nervous system causes plasticity in mul-
tiple regions of the central nervous system. Significant changes have
been reported in map organization, spontaneous activity, neural syn-
chronization and stimulus selectivity2. The ideal method of testing
whether map plasticity or some other form of plasticity is directly
responsible for chronic pain and tinnitus would be to reverse the plas-
ticity and evaluate the perceptual consequence.
Recent attempts to use sensory exposure or discrimination training
have provided some temporary relief6,7. Although the clinical benefits
were limited, these studies provide some support for the hypothesis
that neural plasticity could be used to treat these conditions. It is
possible that a long-lasting reversal of the pathological plasticity in
these patients would provide significant relief.
with electrical stimulation of the cholinergic nucleus basalis generates
this method could be used to reverse the effect of pathological plastic
clinical use. We have developed a less invasive method for generating
sensory inputs, and have demonstrated a potential clinical application.
tic changes. The efficacy of VNS in enhancing plasticity seems to lie in
be used to drive neural plasticity that would reverse the behavioural
correlate of tinnitus in noise-exposed rats. The first set of experiments
confirms that repeatedly pairing a single tone frequency with VNS is
sufficient to generate specific and long-lasting changes in cortical maps.
The rationale for our tinnitus therapy is that increasing the number of
cortical neurons tuned to frequencies other than the tinnitus frequency
tinnitus in noise-exposed rats.
In our first set of experiments, we sought to evaluate whether pairing
in the frequency representation in the cortex, as we found for nucleus
rats) or a 19-kHz, 50-dBSPL tone (n55 rats) for 20 days (SPL, sound
pressure level), 300 times per day in normal-hearing rats with cuff elec-
trodes implanted on the left cervical vagus nerve (Methods). The VNS–
tone pairing procedure was identical to earlier tone pairing procedures
with nucleus basalis, ventral tegmentum or locus coeruleus stimulation
that generate long-lasting map plasticity8,11,12. VNS parameters (30Hz,
0.8mA) were similar to the parameters used in previous rat and human
VNS studies, except that the duration of stimulation and the widths of
individual pulses were reduced by 60-fold and fivefold, respectively
was sufficient to reduce the amplitude of the cortical electroencephalo-
hours after the last VNS–tone pairing session, we used standard micro-
electrode mapping techniques to document frequency map plasticity.
VNS–tone pairing caused a 70–79% increase in the number of primary
tone frequency (Fig. 1). This result confirms our hypothesis that VNS–
tone pairing can be used to direct map plasticity lasting more than 24h.
Pairing VNS with sensory stimuli is a potentially attractive method
epilepsy or depression13. By pairing tones with brief trains of VNS, we
have been able to alter cortical frequency maps significantly in rats
using only 1%of theVNSthatis deliveredclinically(that is, 30s every
5min, 24h per day) for epilepsy treatment in humans.
Having demonstrated that VNS can be used to generate specific and
long-lasting map plasticity, in our second set of studies we sought to
pathological plasticity and eliminate tinnitus. Exposure to intense, high-
frequency noise is known to generate an overrepresentation of mid-
and synchronization of auditory neurons14–16. We induced noise
trauma by exposing rats to 1h of 115-dBSPL, octave-band noise
centred at 16kHz (ref. 17; Methods). Auditory brainstem responses
were used to confirm the effects of the noise exposure on hearing
threshold, including temporary deafness for frequencies above 8kHz
and a long-lasting increase of auditory brainstem response thresholds
as many A1 recording sites were tuned to frequencies between 2 and
4kHz in comparison with naive controls (3567% versus 1462%,
P,0.05), and very few neurons responded to frequencies above
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23kHz (1.761% versus 11.563%, P,0.01). The average frequency
bandwidth of A1 neurons increased by 21% (1.7560.04 versus
1.4760.03 octaves at 10dB above threshold, P,0.00001), and the
averagenumberof spikesevoked bya tone withineach site’s receptive
field increased by 30% (4.360.1 versus 3.360.1, P,0.00001). The
average spontaneous rate increased by 23% (17.760.6 versus
14.360.4 Hz, P,0.00001). The degree of synchronization during
silence measured using the correlation coefficient between multiunit
versus 0.1960.01 synchronous spikes per secondof silence,P,0.05;
have been proposed to be directly responsible for tinnitus2,19. Earlier
studies using several different methods have documented that noise
exposure can generate behavioural correlates of tinnitus near the low-
frequency edge of the noise trauma17,20–22. However, few studies have
directly compared neurophysiology and behavioural observations
from the same animals20,23. It was therefore of great interest to us to
relate noise-induced plasticity to perceptual disturbances.
Each of the eighteen noise-exposed rats used in this study was
centred on 8 or 10kHz, but showed no impairment when the gap
occurred in narrowband noise centred on 2 or 4kHz or in broadband
noise (Fig. 2, 4 weeks after exposure). Several studies have concluded
that a frequency-specific impairment in gap detection is a likely sign
that noise-exposed rats experience a mid-frequency tinnitus percept
which fills the silent gaps17,23(Methods and Supplementary Figs 6–9).
Although it is not possibleto evaluatethe subjective experience of rats
definitively, the gap impairment has been taken as a possible beha-
vioural correlate of tinnitus.
Map distortion and tuning curve broadening (but not changes in
spontaneous activity or synchronization) were significantly correlated
with the degree of gap impairment in untreated noise-exposed rats
(R.0.7 (Pearson correlation coefficient), P,0.05, n58 sham rats;
Figs 3a, b and 4a–d and Supplementary Fig. 13). These correlations
must be interpreted with caution because any variability in the initial
cochlear trauma could generate a correlation between neural and
behavioural changes even in the absence of a causal relationship.
document the reversal of the gap detection impairment.
We speculated that pairing VNS with randomly interleaved pure
tones that span the rat hearing range, but exclude the overrepresented
frequencies, could decrease the cortical representation of the excluded
frequencies24. We also expected that pairing multiple tonefrequencies
with VNS (‘VNS/multiple tone’ pairing) would increase frequency
stimulation experiments25. We quantified behavioural and physio-
logical correlates of tinnitus in noise-exposed rats and then tested
whether pairing VNS with multiple tone frequencies could reverse
the pathological plasticity and eliminate the perceptual disturbance in
VNS was repeatedly paired with multiple pure tones 300 times per
day for 18 days in seven noise-exposed rats with impaired gap detec-
tion for mid-frequency sounds (Methods). Because we found that gap
impairment occurred at 8–10kHz, we selected the frequency of each
randomly interleaved tone to be 1.3, 2.2, 3.7, 17.8 or 29.9kHz. This
pairing procedure was chosen because previous studies suggest it
would reduce the cortical response to mid-frequency tones, increase
frequency selectivity and decrease cortical synchronization2,25. After
ten days of therapy, each of the seven rats showed a significant startle
reduction in cued trials relative to uncued trials for every frequency
tested (P,0.05; Fig. 2a and Supplementary Fig. 9a). Thus, pairing of
sure, which suggests that the rats’ presumed tinnitus was no longer
Putative tinnitus frequency
Gap detection (% startle suppression)
Figure 2 | VNS/multiple tone pairing eliminates the behavioural correlate
of tinnitus. Four weeks after noise exposure, each of the rats in both groups
was unable to detect a gap in one or more of the narrowband noises tested
four weeks after noise exposure is the putative tinnitus frequency for each rat.
For both groups, gap detection at the putative tinnitus frequency was
significantly impaired in comparison to broadband noise (P,0.05). The gap
detection at the non-tinnitus frequency is based on gap detection in 16-kHz
narrowband noise. a, Gap detection at the putative tinnitus frequency (dotted
line) improved significantly after ten days of VNS–tone pairing, and the
improvement persisted at least until the acute physiology experiment (n55
rats). b, Thesham group(n59 rats) continued tobeimpaired. Twosham rats
didnotcontribute data atthenon-tinnitusfrequencybecause they showed gap
impairments at 16kHz (as well as 8 and 10kHz) four weeks after noise
exposure. Black and grey horizontal bars represent duration of VNS and sham
therapy, respectively. Asterisks represent significant differences (P,0.05) in
gap detection at the putative tinnitus frequency between VNS therapy and
sham therapy rats. Error bars, s.e.m.
Percentage of sites
Characteristic frequency (kHz)
VNS + 9 kHz
VNS + 19 kHz
Figure 1 | VNS–tone pairing causes map plasticity. Repeatedly pairing VNS
with a tone increases the number of A1 recordings sites tuned to the paired
frequency. a, VNS was paired with a 9-kHz tone 6,000 times over 20 days in
eight rats. b, VNS was paired with a 19-kHz tone in five rats. This group heard
4-kHz tones equally often but without VNS pairing. Asterisks indicate
significant (P,0.05) increases in the fraction of A1 sites with characteristic
frequencies near the paired tone. Error bars, s.e.m. This result in normal-
distortions induced by exposure to intense noise.
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impairment in their ability to detect gaps in the putative tinnitus fre-
tively) did not show a significant startle reduction in cued trials
(P.0.05; Supplementary Fig. 9b) for at least one of the frequencies
tested at each time point.
ment in gap detection was also eliminated when measured one day,
one week and three weeks after the end of the therapy. This impair-
ment was maintained in all three control groups at every time point
tested (Supplementary Figs 9 and 10). Theseresults indicate that pair-
ing VNS with multiple tone frequencies is sufficient to eliminate the
gap impairment induced by noise exposure (Supplementary Fig. 11).
behavioural correlate of chronic tinnitus.
apy, we evaluated the physiological properties of the auditory cortex of
was due to renormalization of the auditory cortex. After VNS/multiple
tone pairing, most of the A1 properties that were degraded by noise
exposure returned to normal levels. For example, the proportion of A1
tinguishable from that in naive controls after VNS/multiple tone treat-
Figs 12 and 13a). The proportion of A1 neurons responding to 4-kHz,
rats and returned to normal levels in rats that had received the therapy
three weeks earlier (naive, 45.465.0%; sham, 74.167.6%; therapy,
49.166.6%;Figs3 (white circles) and 4a).The degree of low-frequency
map distortion was positively correlated with the degree of gap impair-
ment observed in individual rats (Fig. 4b and Supplementary Fig. 13b).
The percentage of cortex responding to 8-kHz, 30-dBSPL tones (Fig. 3,
(R250.51, P50.006). These results support the earlier hypothesis that
changes in cortical maps are causally related to tinnitus4,26.
VNS/multiple tone pairing reversed the increase in the width of fre-
quency tuning of A1 multiunit activity (that is, decreased frequency
selectivity) observed in noise-exposed rats (Fig. 4c). The bandwidth
(measured at 10, 20, 30 or 40dB above threshold) averaged across all
and Supplementary Fig.14), thus supporting theearlier hypothesis that
decreased frequency selectivity is causally related to tinnitus27.
Intensity (dB SPL)
Noise-exposed rats after sham therapy
Noise-exposed rats after VNS−tone therapy
Characteristic frequency (kHz)
Percentage of A1 responding
Figure 3 | VNS/multipletonepairingreversesmapdistortion. Theincreased
response of A1 neurons to tones following noise exposure is reversed by VNS/
multiple tone pairing. a, Colour indicatesthepercentage ofA1 neurons innaive
rats that respond to a tone of any frequency and intensity combination.
noise-exposed rats that received the VNS/multiple tone therapy. Black contour
Thewhite linesinc indicate significantdecreases(P,0.01)incomparisonwith
noise-exposed sham therapy rats. The filled white circles indicate the tone for
which the increase inthe number of cortical neuronswasgreatest, which isused
to quantify the degree of map distortion in Fig. 4a, b. The filled black circles
indicate the tone for which the proportional increase was greatest.
R2 = 0.36, P = 0.03
R2 = 0.65, P = 0.0002
R2 = 0.26, P = 0.04
R2 = 0.05, P = 0.4
R2 = 0.24, P = 0.054
0 2040 60
0 20 40 60
0 2040 60
Gap detection (%)
Figure 4 | Neurophysiological properties of naive, sham and therapy rats.
a, c, e, g, i, Noise exposure caused a significant map distortion (a), decreased
frequency selectivity (c), increased the tone-evoked response (e), increased the
multiple tone pairing returned each of these parameters, except spontaneous
tone-evoked response strength (f) were all correlated with the degree of gap
impairment in individual rats. h, j, Spontaneous activity (h) and synchronization
ability was quantified as the average gap detection at the putative tinnitus
frequency of each rat, averaged across the four time points collected after the
beginning of therapy (Fig. 2). Error bars, s.e.m. Asterisks represent significant
represent rats from the sham and therapy groups, respectively.
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observed in noise-exposed rats (Fig. 4e). The average number of spikes
evoked by tones within each site’s receptive field was weakly correlated
with the degree of impairment of gap detection (Fig. 4f), supporting the
Finally, VNS/multiple tone pairing also reversed the increase in cor-
the increase in cortical spontaneous activity observed in noise-exposed
the rate of spontaneous activity and the degree of gap impairment
activity and synchronization are not significantly correlated with beha-
vioural correlates of tinnitus in individual rats is consistent with earlier
it remains a possibility that these factors contribute to tinnitus.
Hearing loss, hyperacusis and tinnitus often result from noise expo-
Our results confirm that exposure to intense, high-frequency noise
causes pathological plasticity that is well correlated with the inability
to detect a gap in a mid-frequency, 65-dBSPL tone. Correlations alone
do not suggest that these changes cause tinnitus because another con-
founding factor (such as variability in the degree of cochlear trauma)
could cause both variables to be correlated without acausal connection.
By randomizing the treatment of rats with identical noise exposure, we
response to noise exposure. Thus, our observation that pairing multiple
tone frequencies with VNS can reverse both theneural and behavioural
correlates of tinnitus provides good evidence that abnormal activity in
tinnitus. In addition, neural correlates of hearing loss (tone thresholds)
and hyperacusis (rate level functions) were not correlated with gap
impairment in the rats tested (Supplementary Information). Thus, it is
reasonableto conclude that thegap impairments observed in this study
are primarily related to tinnitus.
VNS-directed plasticity represents a potentially powerful approach
to treating tinnitus. Unlike pharmaceutical approaches, this method
provides the possibility of generating long-lasting and stimulus-spe-
cific changes to neural circuits with minimal side effects. Our control
repeated association of VNS with tones, and not by VNS alone.
Additional studies are needed to determine whether the pairing of
thepathologicalplasticity associatedwith othercommonneurological
conditions, such as chronic pain and amblyopia.
neurophysiology techniques and analysis are described in Methods. The noise
exposure procedure, gap detection testing, and neurophysiology techniques were
identical to those in earlier reports8,17,25.
Full Methods and any associated references are available in the online version of
the paper at www.nature.com/nature.
Received 14 June; accepted 8 November 2010.
Published online 12 January 2011.
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Supplementary Information is linked to the online version of the paper at
Acknowledgements We would like to thank A. Kuzu, J. Omana, D. Vuppala, H. Rasul,
M. Fink, E. Hanacik, R. Miller and C. Walker for help with rat behavioural training. We
would also like to thank J. Eggermont, A. Møller, C. Bauer, J. Fritz, H. Reed, C. Engineer,
A. Reed, M. Brosch, R. Rennaker, R. Beitel, V. Miller, C. McIntyre, G. White, P. Pandya,
R. Tyler andD. deRidder for suggestions about earlier versions ofthe manuscript.This
work was supported by the James S. McDonnell Foundation, the Texas Advanced
Research Program, the National Institute for Deafness and other Communication
Disorders, and MicroTransponder Inc.
Author Contributions N.D.E., J.R.R., J.D.S., S.P.S. and M.S.B. did the behaviour training
sessions, noise exposure and auditory brainstem response recordings. N.D.E., J.R.R.,
surgeries. S.P.S. and N.D.E. did all the VNS implant surgeries. M.P.K. and N.D.E.
designed the experiments, wrote the manuscript and performed data analysis. All
authors discussed the paper and commented on the manuscript.
Author Information Reprints and permissions information is available at
www.nature.com/reprints. The authors declare competing financial interests: details
accompany the full-text HTML version of the paper at www.nature.com/nature.
Readers are welcome to comment on the online version of this article at
www.nature.com/nature. Correspondence and requests for materials should be
addressed to N.D.E. (firstname.lastname@example.org).
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As in humans, only the left vagus nerve was stimulated because the right vagus
nerve contains efferents that stimulate the sinoatrial node and can cause cardiac
brainstem responses (ABRs) and EEG. Each rat was given antibiotics to prevent
infection and a single dose of atropine and dexamethazone to reduce fluid accu-
mulations in the lungs immediately after completion of the surgery.
VNS stimulation parameters and single-tone pairing procedures. VNS was
delivered to unanaesthetized, unrestrained rats in a 25325325cm3wire cage,
located inside a 50360370cm3chamber lined with acoustic insulating foam. A
pilot study was conducted to determine the minimal VNS parameters that reliably
reduced EEG amplitude during slow-wave sleep (Supplementary Fig. 3). VNS para-
(,5kV). The impedance for three rats was unusually high after implantation and
stableacrossthe durationoftrainingforallotherrats. The500-mspure tonesbegan
nucleus basalis stimulation studies, stimulation beginning either 200ms before tone
onset or 50ms after tone onset generated indistinguishable map plasticity8.
VNS was delivered 300 times per day for 20 days, during a VNS–tone pairing
session that lasted 2.5h (Supplementary Figs 2). To prevent rats from anticipating
and the right auditory cortex was exposed to allow for high-density extracellular
(n55) was exposed to a 19-kHz, 50-dBSPL tone paired with VNS. During the
trials in which no VNS was delivered (50%), a 4-kHz, 50-dBSPL tone was pre-
sented. As a result, a 19- or 4-kHz tone was delivered every 15s. Frequency and
intensity calibrations were performed with an ACOPacific microphone (PS9200-
were presented from a speaker (Optimus) suspended 20cm above the wire cage.
Allpairedtoneshad a 5-msrise–falltime.The intensityofeverytonewasselected
to be approximately 20dBSPL above the rat hearing threshold.
Noise exposure and ABRs. Twenty-eight experimental and control rats were
barbiturate-anaesthetized and exposed to 16-kHz, 115-dBSPL, octave-band noise
for 1h (refs 17, 20). A single speaker was positioned 5cm from the left ear. No ear
plugs were used to restrict the noise exposure to one ear. Bilateral noise exposure
confirm cochlear trauma, elevated thresholds were quantified using ABRs in ten
(10ms long,2.5-ms rise–fall time) were deliveredat a rateof20Hz. Tone frequen-
cies were 4, 10, 16 and 32kHz in 10-dB steps from 0 to 85dBSPL. Tones were
The signals were filtered from 100 to 3,000Hz and recorded using BRAINWARE
step at which an ABR could be recognized (Supplementary Fig. 4).
Gap detection testing. The Turner gap detection method was used to assess a
6–8). This method has previously been cross-validated with a conditioned lever
suppression task20(R50.75) and a licking suppression task21. The gap detection
method was selected because it avoids the need for food or water deprivation,
electricshockormonthsofbehaviouraltraining17. Testingtookplace ina 20320
foam. The cage was placed on a startle platform (Lafayette Instrument Co.) that
Sounds were generated using System 3 hardware and software (Tucker-Davis
Technologies) and were delivered by a speaker (Tucker-Davis Technologies
FF1) mounted 20cm above the cage. Rats underwent gap detection testing with
20 and 24kHz at 65dBSPL (ref. 17). Startle responses were elicited by a 20-ms
burst of white noise at 100dBSPL. In 50% of trials, a 50-ms gap embedded in the
continuous soundserved asa warningofasubsequentstartlingnoiseandallowed
rats to reduce the amplitude of the response (Supplementary Fig. 7b). The gap in
the narrowband noise began 100ms before the onset of the broadband startling
noise. Rats underwent 30 trials during each session. The order of sessions with
each startle sound was 30–35s.
or 10kHz) did not serve as an effective warning, presumablybecause the ongoing
were not warned that a loud startling noise was coming and exhibited a strong
startle response (Supplementary Figs 7b and 8b). Gap detection was quantified as
oneminustheratioofthe startleamplitude whenthestartlingnoisewaspreceded
bya gapinthe65-dBSPL, continuousnarrowbandsoundto the startle amplitude
shows typical data from one noise-exposed rat for a session in which the noise
an 8-kHz tone served as the warning cue (right). The warning gap typically
reduced the startle amplitude by 60–70% (Supplementary Fig. 8a). In noise-
exposed rats, gaps in the narrowband noise centred near the low edge of the
trauma noise typically reduced the startle amplitude by less than 20%, which is
not a statistically significant reduction (Supplementary Fig. 8b). The same pro-
impairment four weeks after noise exposure is the putative tinnitus frequency for
each rat (Fig. 2).
study. Five rats were excluded from the study because they showed no detectable
startle response to the noise burst. Of the 31 remaining rats, three were excluded
noise exposure. Eighteen of these showed a statistically significant impairment in
the detection of gaps in one or both mid-frequency (8- or 10-kHz) narrowband
sounds tested, relative to gap detection before noise exposure (P,0.05). Three
rats were excluded from further study because they no longer showed a startle
response to the noise burst (that is, could no longer detect the startle stimulus).
Seven rats were excluded from further study because theyshowed no impairment
ments do not always result from noise exposure is consistent with human and
animal studies showing that although hearing loss is common in individuals with
tinnitus, the majority of individuals with hearing loss do not have tinnitus20,30,31.
in its ability to detect a gap in narrowband noise centred on 8kHz (16 of 18) or
10kHz (12 of 18). None of the 18 rats showed a significant impairment in the
ability to detect a gap in low-frequency narrowband noises (2 or 4kHz) or in
broadband noise (Fig. 2 and Supplementary Fig. 11). This result indicates that
these rats are able to respond normally to the startling noise burst and that the
mechanisms for modulating the startle response using silent gaps remain intact.
Our observation that noise-exposed rats can show gap detection impairments
centred at a single frequency or across a narrow range of frequencies is consistent
with clinical studies showingsignificant heterogeneity across subjects in the spec-
tral characteristics of the tinnitus percept22,32,33. Despite this heterogeneity, a large
fraction of tinnitus patients can match their tinnitus to a pitch and describe their
phantom sound as tonal22.
VNS tone delivery to noise exposed rats. Rats were tested for gap impairment
or experimental therapy. In the VNS/multiple tone paired group (n55 rats),
The tone frequencies paired with VNS in the therapy group were designed to
reduce the 8–10-kHz region of the frequency map. VNS was repeatedly paired
(300 trialsperday). Each tone waspresentedat,20dBabovethe normalhearing
threshold for that frequency. The tone-alonecontrol group was passivelyexposed
to thesametones onthe same schedule as used in the pairedgroup. A VNS-alone
tone pairing, rats were also tested on gap detection one and three weeks after the
end of therapy. At the end of three weeks (that is, 11 weeks after noise exposure),
multiunit responses were recorded from auditory cortex neurons from the ther-
Physiological and behavioural results from the tone-alone, VNS-alone and no-
therapy groups were statistically indistinguishable (Supplementary Fig. 10 and
physiological data not shown). Data from the three groups are combined and
referred to as sham controls in the main text.
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Neurophysiology. In this study, we recorded from a total of 1,492 sites in 21 rats Download full-text
in this report. We recorded 220 multiunit responses from A1 sites in noise-
exposed rats that received VNS/multiple tone pairing (n55). We also recorded
321 A1 sites from noise-exposed rats that did not receive VNS/tone pairing
(n58). The latter group included noise-exposed rats that received tones with
pooledto form a singledata setreferred toas thesham therapy group. During the
acute electrophysiology recordings, sounds were delivered in a foam-lined,
double-walled, sound-attenuated chamber using a speaker (Motorola 40-1221)
were recordedusingParylene-coatedtungstenelectrodesthatwere gluedtogether
(250-mm separation, 2MV at 1kHz; FHC) and lowered approximately 500mm
below the cortical surface. Frequency and intensity calibrations were performed
intensities from 0 to 75dB SPL (1,296 total stimuli). The tones (25-ms duration,
5-ms rise–fall time) were randomly interleaved and separated by 500ms. Tuning
software written in MATLAB v7.9 (Mathworks) to randomize the order of data
mental conditions of each rat during electrophysiology recordings.
Data analysis. Gap discrimination was quantified as the percentage inhibition of
the startle response when a gap (warning cue) preceded the starling noise relative
to the startle response when no gap was present17. Eight of 36 rats tested failed to
generate consistent startle responses and were excluded from the study before
noise exposure. Noise exposure eliminated the startle response in three of the
remaining 28 rats, and these rats were excluded from the study. Noise exposure
failed to generate any impairment in gap detection in seven of the remaining 25
VNS–tone paired rats and eight sham therapy rats). One rat died before neural
responses could be collected. Only behavioural responses (and EEG) were col-
lected from the remaining four rats (two treated rats and two shams) so that the
duration of the benefit could be estimated.
Sites were determined to be in A1 on the basis of continuous tonotopy. At
each A1 recording site, characteristic frequency, frequency bandwidth, response
in which the experimenter was blind to the experimental group and recording
location8. At each pair of simultaneously recorded A1 sites, neural synchrony
duringsilence (300s) was quantified as the cross-correlation function25. The peak
in the cross-correlation function (with or without subtraction of the shift
predictor) was also computed and gave similar results to Pearson correlation
coefficient (R). Map plasticity was quantified as the percent of A1 neurons with
a characteristic frequency in a given range or as the percent of A1 neurons
method of interpolation8,34. Frequency selectivity was quantified as the bandwidth
10, 20, 30or40dBabovethreshold. Results were similarregardlessoftheintensity
each tone within each site’s receptive field and as the spontaneous activity rate
All protocols and recording procedures comply with the NIH Guide for the
Care and Use of Laboratory Animals and were approved by the Institutional
Animal Care and Use Committee at the University of Texas at Dallas.
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34. Engineer, N. D. et al. Environmental enrichment improves response strength,
threshold, selectivity, and latency of auditory cortex neurons. J. Neurophysiol. 92,
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