Highlights

► Prepulse inhibition can be reliably and robustly measured using the Preyer reflex. ► The Preyer reflex is a more reliable response than the whole-body startle in guinea pigs. ► Salicylate impairs gap detection at specific background noise frequencies, indicating the presence of tinnitus. ► Measuring gap detection using the Preyer reflex is a suitable method for identifying tinnitus.

Tinnitus, the perception of sound in the absence of an external stimulus, is a particularly challenging condition to demonstrate in animals. In any animal model, objective confirmation of tinnitus is essential before we can study the neural changes that produce it. A gap detection method, based on prepulse inhibition of the whole-body startle reflex, is often used as a behavioural test for tinnitus in rodents. However, in the guinea pig the whole-body startle reflex is subject to rapid habituation and hence is not an ideal behavioural measure. By contrast, in this species the Preyer or pinna reflex is a very reliable indicator of the startle response and is much less subject to habituation. We have developed a novel adaptation of the gap detection paradigm, which uses the Preyer reflex to measure the startle response, rather than whole-body movement. Using this method, we have demonstrated changes in gap detection, in guinea pigs where tinnitus had been induced by the administration of a high dose of salicylate. Our data indicate that the Preyer reflex gap detection method is a reliable test for tinnitus in guinea pigs.

Cross-talk cancellation is a method for synthesising virtual auditory space using loudspeakers. One implementation is the “Optimal Source Distribution” technique [T. Takeuchi and P. Nelson, J. Acoust. Soc. Am. 112, 2786-2797 (2002)], in which the audio bandwidth is split across three pairs of loudspeakers, placed at azimuths of ±90°, ±15°, and ±3°, conveying low, mid and high frequencies, respectively. A computational simulation of this system was developed and verified against measurements made on an acoustic system using a manikin. Both the acoustic system and the simulation gave a wideband average cancellation of almost 25 dB. The simulation showed that when there was a mismatch between the head-related transfer functions used to set up the system and those of the final listener, the cancellation was reduced to an average of 13 dB. Moreover, in this case the binaural ITDs and ILDs delivered by the simulation of the OSD system often differed from the target values. It is concluded that only when the OSD system is set up with “matched” head-related transfer functions can it deliver accurate binaural cues.

Vocal communication is an important aspect of guinea pig behaviour and a large contributor to their acoustic environment. We postulated that some cortical areas have distinctive roles in processing conspecific calls. In order to test this hypothesis we presented exemplars from all ten of their main adult vocalizations to urethane anesthetised animals while recording from each of the eight areas of the auditory cortex. We demonstrate that the primary area (AI) and three adjacent auditory belt areas contain many units that give isomorphic responses to vocalizations. These are the ventrorostral belt (VRB), the transitional belt area (T) that is ventral to AI and the small area (area S) that is rostral to AI. Area VRB has a denser representation of cells that are better at discriminating among calls by using either a rate code or a temporal code than any other area. Furthermore, 10% of VRB cells responded to communication calls but did not respond to stimuli such as clicks, broadband noise or pure tones. Area S has a sparse distribution of call responsive cells that showed excellent temporal locking, 31% of which selectively responded to a single call. AI responded well to all vocalizations and was much more responsive to vocalizations than the adjacent dorsocaudal core area. Areas VRB, AI and S contained units with the highest levels of mutual information about call stimuli. Area T also responded well to some calls but seems to be specialized for low sound levels. The two dorsal belt areas are comparatively unresponsive to vocalizations and contain little information about the calls. AI projects to areas S, VRB and T, so there may be both rostral and ventral pathways for processing vocalizations in the guinea pig.

Reversible inactivation of the cortex by surface cooling is a powerful method for studying the function of a particular area. Implanted cooling cryoloops have been used to study the role of individual cortical areas in auditory processing of awake-behaving cats. Cryoloops have also been used in rodents for reversible inactivation of the cortex, but recently there has been a concern that the cryoloop may also cool non-cortical structures either directly or via the perfusion of blood, cooled as it passed close to the cooling loop. In this study we have confirmed that the loop can inactivate most of the auditory cortex without causing a significant reduction in temperature of the auditory thalamus or other subcortical structures. We placed a cryoloop on the surface of the guinea pig cortex, cooled it to 2°C and measured thermal gradients across the neocortical surface. We found that the temperature dropped to 20–24°C among cells within a radius of about 2.5 mm away from the loop. This temperature drop was sufficient to reduce activity of most cortical cells and led to the inactivation of almost the entire auditory region. When the temperature of thalamus, midbrain, and middle ear were measured directly during cortical cooling, there was a small drop in temperature (about 4°C) but this was not sufficient to directly reduce neural activity. In an effort to visualize the extent of neural inactivation we measured the uptake of thallium ions following an intravenous injection. This confirmed that there was a large reduction of activity across much of the ipsilateral cortex and only a small reduction in subcortical structures.

Tinnitus is an auditory phenomenon characterised by the perception of a sound in the absence of an external auditory stimulus. Chronic subjective tinnitus is almost certainly maintained via central mechanisms, and this is consistent with observed measures of altered spontaneous brain activity. A number of putative central auditory mechanisms for tinnitus have been proposed. The influential thalamocortical dysrhythmia model suggests that tinnitus can be attributed to the disruption of coherent oscillatory activity between thalamus and cortex following hearing loss. However, the extent to which this disruption specifically contributes to tinnitus or is simply a consequence of the hearing loss is unclear because the necessary matched controls have not been tested. Here, we rigorously test several predictions made by this model in four groups of participants (tinnitus with hearing loss, tinnitus with clinically normal hearing, no tinnitus with hearing loss and no tinnitus with clinically normal hearing). Magnetoencephalography was used to measure oscillatory brain activity within different frequency bands in a ‘resting’ state and during presentation of a masking noise. Results revealed that low-frequency activity in the delta band (1–4 Hz) was significantly higher in the ‘tinnitus with hearing loss’ group compared to the ‘no tinnitus with normal hearing’ group. A planned comparison indicated that this effect was unlikely to be driven by the hearing loss alone, but could possibly be a consequence of tinnitus and hearing loss. A further interpretative linkage to tinnitus was given by the result that the delta activity tended to reduce when tinnitus was masked. High-frequency activity in the gamma band (25–80 Hz) was not correlated with tinnitus (or hearing loss). The findings partly support the thalamocortical dysrhythmia model and suggest that slow-wave (delta band) activity may be a more reliable correlate of tinnitus than high-frequency activity.

The central nucleus of the inferior colliculus (IC) is organized into a series of fibro-dendritic laminae, orthogonal to the tonotopic progression. Many neurons have their dendrites confined to one lamina while others have dendrites that cross over a number of laminae. Here, we have used juxtacellular labeling in urethane anesthetized guinea pigs to visualize the cells with biocytin and have analyzed their response properties, in order to try and link their structure and function. Out of a sample of 38 filled cells, 15 had dendrites confined within the fibro-dendritic laminae and in 13 we were also able to reconstruct their local axonal tree. Based on dendritic morphology they were subdivided into flat or less flat; small, medium, or large; elongated or disk-shaped cells. Two of the elongated cells had many dendritic spines while the other cells had few or none. Twelve of the cells had their local axonal tree restricted to the same lamina as their dendrites while one cell had its dendrites in a separate lamina from the axon. The axonal plexus was more extensive (width 0.7–1.4 mm) within the lamina than the dendrites (width generally 0.07–0.53 mm). The intrinsic axons were largely confined to a single lamina within the central nucleus, but at least half the cells also had output axons with two heading for the commissure and five heading into the brachium. We were able to identify similarities in the physiological response profiles of small groups of our filled cells but none appeared to represent a homogeneous morphological cell type. The only common feature of our sample was one of exclusion in that the onset response, a response commonly recorded from IC cells, was never seen in laminar cells, but was in cells with a stellate morphology. Thus cells with laminar dendrites have a wide variety of physiological responses and morphological subtypes, but over 90% have an extensive local axonal tree within their local lamina.

Objective: We explored the relationship between audiogram shape and tinnitus pitch to answer questions arising from neurophysiological models of tinnitus: ‘Is the dominant tinnitus pitch associated with the edge of hearing loss?’ and ‘Is such a relationship more robust in people with narrow tinnitus bandwidth or steep sloping hearing loss?’ Design: A broken-stick fitting objectively quantified slope, degree and edge of hearing loss up to 16 kHz. Tinnitus pitch was characterized up to 12 kHz. We used correlation and multiple regression analyses for examining relationships with many potentially predictive audiometric variables. Study Sample: 67 people with chronic bilateral tinnitus (43 men and 24 women, aged from 22 to 81 years). Results: In this ample of 67 subjects correlation failed to reveal any relationship between the tinnitus pitch and the edge frequency. The tinnitus pitch generally fell within the area of hearing loss. The pitch of the tinnitus in a subset of subjects with a narrow tinnitus bandwidth (n = 23) was associated with the audiometric edge. Conclusions: Our findings concerning subjects with narrow tinnitus bandwidth suggest that this can be used as an a priori inclusion criterion. A large group of such subjects should be tested to confirm these results.

Psychophysical forward masking is an increase in threshold of detection of a sound (probe) when it is preceded by another sound (masker). This is reminiscent of the reduction in neuronal responses to a sound following prior stimulation. Studies in the auditory nerve and cochlear nucleus using signal detection theory techniques to derive neuronal thresholds showed that in centrally projecting neurons, increases in masked thresholds were significantly smaller than the changes measured psychophysically. Larger threshold shifts have been reported in the inferior colliculus of awake marmoset. The present study investigated the magnitude of forward masking in primary auditory cortical neurons of anaesthetised guinea-pigs. Responses of cortical neurons to unmasked and forward masked tones were measured and probe detection thresholds estimated using signal detection theory methods. Threshold shifts were larger than in the auditory nerve, cochlear nucleus and inferior colliculus. The larger threshold shifts suggest that central, and probably cortical, processes contribute to forward masking. However, although methodological differences make comparisons difficult, the threshold shifts in cortical neurons were, in contrast to subcortical nuclei, actually larger than those observed psychophysically. Masking was largely attributable to a reduction in the responses to the probe, rather than either a persistence of the masker responses or an increase in the variability of probe responses.

Psychophysical forward masking is an increase in threshold of detection of a sound (probe) when it is preceded by another sound (masker). This is reminiscent of the reduction in neuronal responses to a sound following prior stimulation. Studies in the auditory nerve and cochlear nucleus using signal detection theory techniques to derive neuronal thresholds showed that in centrally projecting neurons, increases in masked thresholds were significantly smaller than the changes measured psychophysically. Larger threshold shifts have been reported in the inferior colliculus of awake marmoset. The present study investigated the magnitude of forward masking in primary auditory cortical neurons of anaesthetised guinea-pigs. Responses of cortical neurons to unmasked and forward masked tones were measured and probe detection thresholds estimated using signal detection theory methods. Threshold shifts were larger than in the auditory nerve, cochlear nucleus and inferior colliculus. The larger threshold shifts suggest that central, and probably cortical, processes contribute to forward masking. However, although methodological differences make comparisons difficult, the threshold shifts in cortical neurons were, in contrast to subcortical nuclei, actually larger than those observed psychophysically. Masking was largely attributable to a reduction in the responses to the probe, rather than either a persistence of the masker responses or an increase in the variability of probe responses.

The directionality of hair cell stimulation combined with the vibration of the basilar membrane causes the auditory nerve fiber action potentials, in response to low-frequency stimuli, to occur at a particular phase of the stimulus waveform. Because direct mechanical measurements at the cochlear apex are difficult, such phase locking has often been used to indirectly infer the basilar membrane motion. Here, we confirm and extend earlier data from mammals using sine wave stimulation over a wide range of sound levels (up to 90 dB sound pressure level). We recorded phase-locked responses to pure tones over a wide range of frequencies and sound levels of a large population of auditory nerve fibers in the anesthetized guinea pig. The results indicate that, for a constant frequency of stimulation, the phase lag decreases with increases in the characteristic frequency (CF) of the nerve fiber. The phase lag decreases up to a CF above the stimulation frequency, beyond which it decreases at a much slower rate. Such phase changes are consistent with known basal cochlear mechanics. Measurements from individual fibers showed smaller but systematic variations in phase with sound level, confirming previous reports. We found a “null” stimulation frequency at which little variation in phase occurred with sound level. This null frequency was often not at the CF. At stimulation frequencies below the null, there was a progressive lag with sound level and a progressive lead for stimulation frequencies above the null. This was maximally 0.2 cycles.

Humans perceive a harmonic series as a single auditory object with a pitch equivalent to the fundamental frequency (F0) of the series. When harmonics are presented to alternate ears, the repetition rate of the waveform at each ear doubles. If the harmonics are resolved, then the pitch perceived is still equivalent to F0, suggesting the stimulus is binaurally integrated before pitch is processed. However, unresolved harmonics give rise to the doubling of pitch which would be expected from monaural processing (Bernstein and Oxenham, J. Acoust. Soc. Am., 113:3323–3334, 2003). We used similar stimuli to record responses of multi-unit clusters in the central nucleus of the inferior colliculus (IC) of anesthetized guinea pigs (urethane supplemented by fentanyl/fluanisone) to determine the nature of the representation of harmonic stimuli and to what extent there was binaural integration. We examined both the temporal and rate-tuning of IC clusters and found no evidence for binaural integration. Stimuli comprised all harmonics below 10 kHz with fundamental frequencies (F0) from 50 to 400 Hz in half-octave steps. In diotic conditions, all the harmonics were presented to both ears. In dichotic conditions, odd harmonics were presented to one ear and even harmonics to the other. Neural characteristic frequencies (CF, n = 85) were from 0.2 to 14.7 kHz; 29 had CFs below 1 kHz. The majority of clusters responded predominantly to the contralateral ear, with the dominance of the contralateral ear increasing with CF. With diotic stimuli, over half of the clusters (58%) had peaked firing rate vs. F0 functions. The most common peak F0 was 141 Hz. Almost all (98%) clusters phase locked diotically to an F0 of 50 Hz, and approximately 40% of clusters still phase locked significantly (Rayleigh coefficient >13.8) at the highest F0 tested (400 Hz). These results are consistent with the previous reports of responses to amplitude-modulated stimuli. Clusters phase locked significantly at a frequency equal to F0 for contralateral and diotic stimuli but at 2F0 for dichotic stimuli. We interpret these data as responses following the envelope periodicity in monaural channels rather than as a binaurally integrated representation.

The discrimination of a change in a stimulus is determined both by the magnitude of that change and by the variability in the neural response to the stimulus. When the stimulus is itself noisy, then the relative contributions of the neural (intrinsic) and stimulus induced variability becomes a critical question. We measured the contribution of intrinsic neural noise and interstimulus variability to the discrimination of interaural time differences (ITDs) and interaural correlation (IAC). We measured discharge rate versus characteristic frequency (CF) tone ITD functions, and CF-centered narrowband noise ITD and IAC functions in interleaved blocks in the same units in the inferior colliculus of urethane-anesthetized guinea pigs. Ten “frozen” tokens of noise were synthesized and the responses to each token were separately analyzed to allow the relative contributions of intrinsic and stimulus variability to be assessed. ITD and IAC discrimination thresholds were determined for a simulated two-interval forced-choice experiment, based on the firing rate distributions, using receiver operating characteristic analysis. On average, between stimulus variability contributed 19% (range, 1.5–30%) of the variance in noise ITD discrimination and 27% (range, 3–50%) in IAC discrimination. Noise ITD thresholds were slightly higher than tone ITD thresholds. Taking the mean of the thresholds for individual noise tokens gave a similar result to pooling across all noise tokens. This implies that although the stimulus induced variability is measurable, it is insignificant in relation to the intrinsic noise in ITD and IAC discrimination.

Sensitivity to changes in the interaural correlation of 50-ms bursts of narrowband or broadband noise was measured in single neurons in the inferior colliculus of urethane-anaesthetized guinea pigs. Rate vs. interaural correlation functions (rICFs) were measured using two methods. These methods compensated in different ways for the inherent variance in interaural correlation between tokens with the same expected correlation. The shape of all rICFs could be best described by power functions allowing them to be summarized by two parameters. Most rICFs were best fit by a power below 2, indicating that they were only slightly nonlinear. However, there were a few fitted functions that had a power of 3–6, indicating marked curvature. Modeling results indicate that the nonlinearity of the majority of rICFs was explicable in terms of the monaural transduction stages; however, some of the rICFs with power greater than 2 require either multiple inputs to the coincidence detector or additional nonlinearities to be included in the model. Discrimination thresholds were estimated at reference correlations of −1, 0, and +1 using receiver operating characteristic (ROC) analysis of the spike-count distribution at each correlation. Thresholds spanned the full possible range, from a minimum of 0.1 to the maximum possible of 2. Thresholds were generally highest with a reference correlation of −1, intermediate with a reference of 0, and lowest with a reference correlation of +1. Thresholds were lowest for the most steeply sloped rICFs, but thresholds were not strongly correlated to the spike rate variance. The lowest thresholds occurred using narrowband noise that was compensated for internal delays, but they were still about three times larger than human psychophysical thresholds measured using similar stimuli. The data suggest that, unlike pure tone interaural time difference, discrimination of a population measure is required to account for behavioral interaural correlation discrimination performance.

Considerable circumstantial evidence suggests that cells in the ventral cochlear nucleus, that respond predominantly to the onset of pure tone bursts, have a stellate morphology and project, among other places, to the dorsal cochlear nucleus. The characteristics of such cells make them leading candidates for providing the so-called “wideband inhibitory input” which is an essential part of the processing machinery of the dorsal cochlear nucleus. Here we use juxtacellular labeling with biocytin to demonstrate directly that large stellate cells, with onset responses, terminate profusely in the dorsal cochlear nucleus. They also provide widespread local innervation of the anteroventral cochlear nucleus and a small innervation of the posteroventral cochlear nucleus. In addition, some onset cells project to the contralateral dorsal cochlear nucleus.

We investigated how physiologically observed forward suppression interacts with stimulus frequency in neuronal responses in the guinea pig auditory cortex. The temporal order and frequency proximity of sounds influence both their perception and neuronal responses. Psychophysically, preceding sounds (conditioners) can make successive sounds (probes) harder to hear. These effects are larger when the two sounds are spectrally similar. Physiological forward suppression is usually maximal for conditioner tones near to a unit's characteristic frequency (CF), the frequency to which a neuron is most sensitive. However, in most physiological studies, the frequency of the probe tone and CF are identical, so the role of unit CF and probe frequency cannot be distinguished. Here, we systemically varied the frequency of the probe tone, and found that the tuning of suppression was often more closely related to the frequency of the probe tone than to the unit's CF, i.e. suppressed tuning was specific to probe frequency. This relationship was maintained for all measured gaps between the conditioner and the probe tones. However, when the probe frequency and CF were similar, CF tended to determine suppressed tuning. In addition, the bandwidth of suppression was slightly wider for off-CF probes. Changes in tuning were also reflected in the firing rate in response to probe tones, which was maximally reduced when probe and conditioner tones were matched in frequency. These data are consistent with the idea that cortical neurons receive convergent inputs with a wide range of tuning properties that can adapt independently.