Auditory Research publications

Significance A decline in organic brain health paralleled by degraded perceptual and cognitive abilities is a well-established aspect of human and animal aging. Here, in a rodent model, an elementary form of attention-demanding auditory training completed in young adulthood resulted in positive changes in modulation-rate discrimination abilities that carried forward into older age. These sustained behavioral impacts for processing speed were paralleled by enduring changes in physical and functional aspects of perceptual processing, documented at the level of the primary auditory cortex. Additionally, an important generalization of training impacts was recorded in the hippocampus, a brain structure for which physical and functional declines have been long associated with the progression to infirmity and dementia in aging.

Feedback signals from the primary auditory cortex (A1) can shape the receptive field properties of neurons in the ventral division of the medial geniculate body (MGBv). However, the behavioral significance of corticothalamic modulation is unknown. The aim of this study was to elucidate the role of this descending pathway in the perception of complex sounds. We tested the ability of adult female ferrets to detect the presence of a mistuned harmonic in a complex tone using a positive conditioned go/no-go behavioral paradigm before and after the input from layer VI in A1 to MGBv was bilaterally and selectively eliminated using chromophore-targeted laser photolysis. MGBv neurons were identified by their short latencies and sharp tuning curves. They responded robustly to harmonic complex tones and exhibited an increase in firing rate and temporal pattern changes when one frequency component in the complex tone was mistuned. Injections of fluorescent microbeads conjugated with a light-sensitive chromophore were made in MGBv, and, following retrograde transport to the cortical cell bodies, apoptosis was induced by infrared laser illumination of A1. This resulted in a selective loss of ∼60% of layer VI A1-MGBv neurons. After the lesion, mistuning detection was impaired, as indicated by decreased d' values, a shift of the psychometric curves toward higher mistuning values, and increased thresholds, whereas discrimination performance was unaffected when level cues were also available. Our results suggest that A1-MGBv corticothalamic feedback contributes to the detection of harmonicity, one of the most important grouping cues in the perception of complex sounds.SIGNIFICANCE STATEMENT Perception of a complex auditory scene is based on the ability of the brain to group those sound components that belong to the same source and to segregate them from those belonging to different sources. Because two people talking simultaneously may differ in their voice pitch, perceiving the harmonic structure of sounds is very important for auditory scene analysis. Here we demonstrate mistuning sensitivity in the thalamus and that feedback from the primary auditory cortex is required for the normal ability of ferrets to detect a mistuned harmonic within a complex sound. These results provide novel insight into the function of descending sensory pathways in the brain and suggest that this corticothalamic circuit plays an important role in scene analysis.

Liposome nanocarriers (LPNs) are potentially the future of inner ear therapy due to their high drug loading capacity and efficient uptake in the inner ear after a minimally invasive intratympanic administration. However, information on the biocompatibility of LPNs in the inner ear is lacking. The aim of the present study is to document the biocompatibility of LPNs in the inner ear after intratympanic delivery. LPNs with or without gadolinium-tetra-azacyclo-dodecane-tetra-acetic acid (Gd-DOTA) were delivered to the rats through transtympanic injection. The distribution of the Gd-DOTA-containing LPNs in the middle and inner ear was tracked in vivo using MRI. The function of the middle and inner ear barriers was evaluated using gadolinium-enhanced MRI. The auditory function was measured using auditory brainstem response (ABR). The potential inflammatory response was investigated by analyzing glycosaminoglycan and hyaluronic acid secretion and CD44 and TLR2 expression in the inner ear. The potential apoptosis was analyzed using terminal transferase (TdT) to label the free 3'OH breaks in the DNA strands of apoptotic cells with TMR-dUTP (TUNEL staining). As a result, LPNs entered the inner ear efficiently after transtympanic injection. The transtympanic injection of LPNs with or without Gd-DOTA neither disrupted the function of the middle and inner ear barriers nor caused hearing impairment in rats. The critical inflammatory biological markers in the inner ear, including glycosaminoglycan and hyaluronic acid secretion and CD44 and TLR2 expression, were not influenced by the administration of LPNs. There was no significant cell death associated with the administration of LPNs. The transtympanic injection of LPNs is safe for the inner ear, and LPNs may be applied as a drug delivery matrix in the clinical therapy of sensorineural hearing loss.

Human listeners place greater weight on the beginning of a sound compared to the middle or end when determining sound location, creating an auditory illusion known as the Franssen effect. Here, we exploited that effect to test whether human auditory cortex (AC) represents the physical vs. perceived spatial features of a sound. We used functional magnetic resonance imaging (fMRI) to measure AC responses to sounds that varied in perceived location due to interaural level differences (ILD) applied to sound onsets or to the full sound duration. Analysis of hemodynamic responses in AC revealed sensitivity to ILD in both full-cue (veridical) and onset-only (illusory) lateralized stimuli. Classification analysis revealed regional differences in the sensitivity to onset-only ILDs, where better classification was observed in posterior compared to primary AC. That is, restricting the ILD to sound onset-which alters the physical but not the perceptual nature of the spatial cue-did not eliminate cortical sensitivity to that cue. These results suggest that perceptual representations of auditory space emerge or are refined in higher-order AC regions, supporting the stable perception of auditory space in noisy or reverberant environments and forming the basis of illusions such as the Franssen effect.

Sound stimuli are characterized by their rich spectral and temporal dynamic properties. Individual neurons in auditory cortex (ACX) encode both spectral and temporal aspects of sounds e.g. sound onset and/or offset. While the different fields of the ACX show gradients of frequency selectivity the large-scale organization of sound dynamics is unknown. We used widefield imaging of GCaMP6s in awake mouse ACX combined with a novel unsupervised image segmentation technique to investigate the spatiotemporal representation of sound onset and offset. Using this technique, we identified known auditory fields but also detected novel ACX areas. Furthermore, we found that ACX areas differed in their responses to tone onset and offset. Multiple areas were preferentially activated by tone offset, and on-response areas were more spatially localized than off-response areas. We also found tonotopy in off-responses. Together our results demonstrate a different spatial distribution of neurons across ACX for processing sound onsets versus offsets.

Purpose Our study aimed to explore the feasibility of manganese‐enhanced magnetic resonance imaging ( MEMRI ) combined with visual evoked potentials ( VEP ) and auditory evoked visual cortex responses ( AVR ) in evaluating for the establishment of visual/auditory compensatory pathways after monocular blindness. Materials and Methods A total of 14 healthy neonatal male Sprague‐Dawley rats were randomly divided into two groups ( n = 7 for Groups A and B). Right optic nerve ( ON ) transection was performed on the 7 rats of Group A to obtain a monocularly blind model, and the 7 rats of Group B were chosen as the control group. Four months later, 400 mmol MnCl 2 was injected into the left eye in both groups via intravitreal injection. The changes in the visual pathways projected from the blind eye and the remaining eye in Group A and the normal eyes in Group B were compared to determine if new visual compensatory pathways were established. Additionally, VEP tests were performed to determine complete blindness, and AVR examinations were performed to help identify the generation of auditory compensatory function. Results The VEP test indicated complete visual loss after ON transection. In the monocularly blind rats, the contrast‐to‐noise ratio ( CNR ) of ON , optic tract ( OT ), lateral geniculate nucleus ( LGN ), superior colliculus ( SC ), optic radiation ( OR ) and visual cortex ( VC ) of visual pathway projected from the left eye was significantly higher than that of the right pathway ( p < .001). Moreover, the CNR of ON , OT , LGN , SC , OR and VC in the visual pathway projected from the left eye of monocularly blind rats was significantly lower than those of normal rats ( p < .05). The AVR results revealed that the corresponding bilateral visual cortex in monocularly blind rats did not respond to the auditory stimulus or showed dissimilation with the low frequency. Conclusion MEMRI combined with electrophysiology, including VEP and AVR , may be potentially helpful in the evaluation of the possible generation of new visual/auditory compensatory pathways after monocular blindness.

Hair cells of the cochlea are mechanosensors for the perception of sound. Mutations in the LRTOMT gene, which encodes a protein with homology to the catecholamine methyltransferase COMT that is linked to schizophrenia, cause deafness. Here, we show that Tomt/Comt2, the murine ortholog of LRTOMT, has an unexpected function in the regulation of mechanotransduction by hair cells. The role of mTOMT in hair cells is independent of mTOMT methyltransferase function and mCOMT cannot substitute for mTOMT function. Instead, mTOMT binds to putative components of the mechanotransduction channel in hair cells and is essential for the transport of some of these components into the mechanically sensitive stereocilia of hair cells. Our studies thus suggest functional diversification between mCOMT and mTOMT, where mTOMT is critical for the assembly of the mechanotransduction machinery of hair cells. Defects in this process are likely mechanistically linked to deafness caused by mutations in LRTOMT/Tomt.

Gamma‐aminobutyric acid (GABA) and glutamate are believed to have inhibitory and exhibitory neuromodulatory effects that regulate the brain's response to sensory perception. Furthermore, frequency‐specific synchronization of neuronal excitability within the gamma band (30–80 Hz) is attributable to a homeostatic balance between excitation and inhibition. However, our understanding of the physiological mechanism underlying gamma rhythms is based on animal models. Investigations of the relationship between GABA concentrations, glutamate concentrations, and gamma band activity in humans were mostly restricted to the visual cortex and are conflicting. Here, we performed a multimodal imaging study combining magnetic resonance spectroscopy (MRS) with electroencephalography (EEG) in the auditory cortex. In 14 healthy subjects, we investigated the impact of individual differences in GABA and glutamate concentration on gamma band response (GBR) following auditory stimulus presentation. We explored the effects of bulk GABA on the GBR across frequency (30–200 Hz) and time (−200 to 600 ms) and found no significant relationship. Furthermore, no correlations were found between gamma peak frequency or power measures and metabolite concentrations (GABA, glutamate, and GABA/glutamate ratio). These findings suggest that, according to MRS measurements, and given the auditory stimuli used in this study, GABA and glutamate concentrations are unlikely to play a significant role in the inhibitory and excitatory drive in the generation of gamma band activity in the auditory cortex. Hum Brain Mapp 38:3975–3987, 2017 . © 2017 Wiley Periodicals, Inc.

Glucose-6-phosphate dehydrogenase (G6PD) is the first and rate-limiting enzyme of the pentose phosphate pathway; it catalyzes the conversion of glucose-6-phosphate to 6-phosphogluconate and NADP+ to NADPH and is thought to be the principal source of NADPH for the cytosolic glutathione and thioredoxin antioxidant defense systems. We investigated the roles of G6PD in the cytosolic antioxidant defense in the cochlea of G6pd hypomorphic mice that were backcrossed onto normal-hearing CBA/CaJ mice. Young G6pd-deficient mice displayed a significant decrease in cytosolic G6PD protein levels and activities in the inner ears. However, G6pd deficiency did not affect the cytosolic NADPH redox state, or glutathione or thioredoxin antioxidant defense in the inner ears. No histological abnormalities or oxidative damage was observed in the cochlea of G6pd hemizygous males or homozygous females. Furthermore, G6pd deficiency did not affect auditory brainstem response hearing thresholds, wave I amplitudes or wave I latencies in young males or females. In contrast, G6pd deficiency resulted in increased activities and protein levels of cytosolic isocitrate dehydrogenase 1, an enzyme that catalyzes the conversion of isocitrate to α-ketoglutarate and NADP+ to NADPH, in the inner ear. In a mouse inner ear cell line, knockdown of Idh1, but not G6pd, decreased cell growth rates, cytosolic NADPH levels, and thioredoxin reductase activities. Therefore, under normal physiological conditions, G6pd deficiency does not affect the cytosolic glutathione or thioredoxin antioxidant defense in mouse cochlea. Under G6pd deficiency conditions, isocitrate dehydrogenase 1 likely functions as the principal source of NADPH for cytosolic antioxidant defense in the cochlea.SIGNIFICANCE STATEMENT Glucose-6-phosphate dehydrogenase (G6PD) is the first and rate-limiting enzyme of the pentose phosphate pathway; it catalyzes the conversion of glucose-6-phosphate to 6-phosphogluconate and NADP+ to NADPH and is thought to be the principal source of NADPH for the cytosolic glutathione and thioredoxin antioxidant defense systems. In the current study, we show that, under normal physiological conditions, G6pd deficiency does not affect the cytosolic glutathione or thioredoxin antioxidant defense in the mouse cochlea. However, under G6pd deficiency conditions, isocitrate dehydrogenase 1 likely functions as the principal source of NADPH for cytosolic antioxidant defense in the cochlea.

Rat ultrasonic vocalizations have been suggested to be either a byproduct of physical movement or, in the case of 50-kHz calls, a means to communicate positive affect. Yet there are up to 14 distinct types of 50-kHz calls, raising issues for both explanations. To discriminate between these theories and address the purpose for the numerous 50-kHz call types, we studied single juvenile rats that were waiting to play with a partner, a situation associated with a high number of 50-kHz calls. We used a Monte-Carlo shuffling procedure to identify vocalization-behavior correlations that were statistically different from chance. We found that certain call types ("split", "composite" and "multi-step") were strongly associated with running and jumping while other call types (those involving "trills") were more common during slower movements. Further, non-locomotor states such as resting and rearing were strongly predictive of a lack of vocalizations. We also found that the various sub-types of USVs can be clustered into 3-4 categories based on similarities in the way they are used. We did not find a one-to-one relationship between any movements and specific vocalizations, casting doubt on the motion byproduct theory. On the other hand, the use of specific calls during specific behaviors is problematic for the affect communication hypothesis. Based on our results, we suggest that ultrasonic calls may serve to coordinate moment-to-moment social interactions.

This study was guided by the hypothesis that the aging central nervous system progressively loses its ability to process rapid acoustic changes that are important for speech recognition. Specifically, we hypothesized that age-related deficits in neural synchrony and neuronal oscillatory activity occur independently in older adults and disrupt auditory temporal processing. Neural synchrony is largely dependent on phase locking within the central auditory pathway, beginning at the auditory nerve. In contrast, the resonance characteristics of oscillatory activity are dependent on the integrity and structure of long range cortical connections. We tested our hypotheses by assessing age-related differences in electrophysiologic correlates of neural synchrony and peak oscillatory frequency in younger and older adults with normal hearing and determining their associations with a behavioral measure of gap detection. Phase-locking values were smaller (poorer neural synchrony) and peak alpha frequency was lower for older than younger adults and decreased as gap detection thresholds increased; variations in phase-locking values and peak alpha frequency uniquely predicted gap detection thresholds. These effects were driven, in large part, by associations in older adults. These results reveal dissociable neural mechanisms associated with distinct underlying pathology that may differentially be present in older adults and contribute to auditory processing declines.

Nicotine enhances sensory and cognitive processing via actions at nicotinic acetylcholine receptors (nAChRs), yet the precise circuit- and systems-level mechanisms remain unclear. In sensory cortex, nicotinic modulation of receptive fields (RFs) provides a model to probe mechanisms by which nAChRs regulate cortical circuits. Here, we examine RF modulation in mouse primary auditory cortex (A1) using a novel electrophysiological approach: current-source density (CSD) analysis of responses to tone-in-notched-noise (TINN) acoustic stimuli. TINN stimuli consist of a tone at the characteristic frequency (CF) of the recording site embedded within a white noise stimulus filtered to create a spectral “notch” of variable width centered on CF. Systemic nicotine (2.1 mg/kg) enhanced responses to the CF tone and to narrow-notch stimuli, yet reduced the response to wider-notch stimuli, indicating increased response gain within a narrowed RF. Subsequent manipulations showed that modulation of cortical RFs by systemic nicotine reflected effects at several levels in the auditory pathway: nicotine suppressed responses in the auditory midbrain and thalamus, with suppression increasing with spectral distance from CF so that RFs became narrower, and facilitated responses in the thalamocortical pathway, while nicotinic actions within A1 further contributed to both suppression and facilitation. Thus, multiple effects of systemic nicotine integrate along the ascending auditory pathway. These actions at nAChRs in cortical and subcortical circuits, which mimic effects of auditory attention, likely contribute to nicotinic enhancement of sensory and cognitive processing.

While a topographic map of auditory space exists in the vertebrate midbrain, it is absent in the forebrain. Yet, both brain regions are implicated in sound localization. The heterogeneous spatial tuning of adjacent sites in the forebrain compared to the midbrain reflects different underlying circuitries, which is expected to affect the correlation structure, i.e., signal (similarity of tuning) and noise (trial-by-trial variability) correlations. Recent studies have drawn attention to the impact of response correlations on the information readout from a neural population. We thus analyzed the correlation structure in midbrain and forebrain regions of the barn owl’s auditory system. Tetrodes were used to record in the midbrain and two forebrain regions, Field L and the downstream auditory arcopallium (AAr), in anesthetized owls. Nearby neurons in the midbrain showed high signal and noise correlations (R NC s), consistent with shared inputs. As previously reported, Field L was arranged in random clusters of similarly tuned neurons. Interestingly, AAr neurons displayed homogeneous monotonic azimuth tuning, while response variability of nearby neurons was significantly less correlated than the midbrain. Using a decoding approach, we demonstrate that low R NC in AAr restricts the potentially detrimental effect it can have on information, assuming a rate code proposed for mammalian sound localization. This study harnesses the power of correlation structure analysis to investigate the coding of auditory space. Our findings demonstrate distinct correlation structures in the auditory midbrain and forebrain, which would be beneficial for a rate-code framework for sound localization in the nontopographic forebrain representation of auditory space.

The functional organization of human auditory cortex remains incompletely characterized. While the posteromedial two thirds of Heschl's gyrus (HG) is generally considered to be part of core auditory cortex, additional subdivisions of HG remain speculative. To further delineate the hierarchical organization of human auditory cortex, we investigated regional heterogeneity in the modulation of auditory cortical responses under varying depths of anesthesia induced by propofol. Non-invasive studies have shown that propofol differentially affects auditory cortical activity, with a greater impact on non-core areas. Subjects were neurosurgical patients undergoing removal of intracranial electrodes placed to identify epileptic foci. Stimuli were 50Hz click trains, presented continuously during an awake baseline period, and subsequently, while propofol infusion was incrementally titrated to induce general anesthesia. Electrocorticographic recordings were made with depth electrodes implanted in HG and subdural grid electrodes implanted over superior temporal gyrus (STG). Depth of anesthesia was monitored using spectral entropy. Averaged evoked potentials (AEPs), frequency-following responses (FFRs) and high gamma (70-150Hz) event-related band power were used to characterize auditory cortical activity. Based on the changes in AEPs and FFRs during the induction of anesthesia, posteromedial HG could be divided into two subdivisions. In the most posteromedial aspect of the gyrus, the earliest AEP deflections were preserved and FFRs increased during induction. In contrast, the remainder of the posteromedial HG exhibited attenuation of both the AEP and the FFR. The anterolateral HG exhibited weaker activation characterized by broad, low-voltage AEPs and the absence of FFRs. Lateral STG exhibited limited activation by click trains, and FFRs there diminished during induction. Sustained high gamma activity was attenuated in the most posteromedial portion of HG, and was absent in all other regions. These differential patterns of auditory cortical activity during the induction of anesthesia may serve as useful physiological markers for field delineation. In this study, the posteromedial HG could be parcellated into at least two subdivisions. Preservation of the earliest AEP deflections and FFRs in the posteromedial HG likely reflects the persistence of feedforward synaptic activity generated by inputs from subcortical auditory pathways, including the medial geniculate nucleus.

The manner in which the brain integrates different sensory inputs to facilitate perception and behavior has been the subject of numerous speculations. By examining multisensory neurons in cat superior colliculus, the present study demonstrated that two operational principles are sufficient to understand how this remarkable result is achieved: (1) unisensory signals are integrated continuously and in real time as soon as they arrive at their common target neuron and (2) the resultant multisensory computation is modified in shape and timing by a delayed, calibrating inhibition. These principles were tested for descriptive sufficiency by embedding them in a neurocomputational model and using it to predict a neuron's moment-by-moment multisensory response given only knowledge of its responses to the individual modality-specific component cues. The predictions proved to be highly accurate, reliable, and unbiased and were, in most cases, not statistically distinguishable from the neuron's actual instantaneous multisensory response at any phase throughout its entire duration. The model was also able to explain why different multisensory products are often observed in different neurons at different time points, as well as the higher-order properties of multisensory integration, such as the dependency of multisensory products on the temporal alignment of crossmodal cues. These observations not only reveal this fundamental integrative operation, but also identify quantitatively the multisensory transform used by each neuron. As a result, they provide a means of comparing the integrative profiles among neurons and evaluating how they are affected by changes in intrinsic or extrinsic factors. SIGNIFICANCE STATEMENT Multisensory integration is the process by which the brain combines information from multiple sensory sources (e.g., vision and audition) to maximize an organism's ability to identify and respond to environmental stimuli. The actual transformative process by which the neural products of multisensory integration are achieved is poorly understood. By focusing on the millisecond-by-millisecond differences between a neuron's unisensory component responses and its integrated multisensory response, it was found that this multisensory transform can be described by two basic principles: unisensory information is integrated in real time and the multisensory response is shaped by calibrating inhibition. It is now possible to use these principles to predict a neuron's multisensory response accurately armed only with knowledge of its unisensory responses.

Cortical responses to repeated stimuli are highly dynamic and rapidly adaptive. Such rapid changes are prominent in all sensory cortices, across which many aspects of circuitry are conserved. As an example, in the auditory cortex, preceding sounds can powerfully suppress responses to later, spectrally similar sounds – a phenomenon called forward suppression. Whether cortical inhibitory networks shape such suppression, or whether it is wholly regulated by common mechanisms such as synaptic depression or spike-frequency adaptation, is controversial. Here, we show that optogenetically suppressing somatostatin-positive interneurons reveals facilitation in neurons that are normally forward-suppressed. This is accompanied by a weakening of forward suppression, suggesting that these interneurons regulate the strength of forward interactions. In contrast, inactivating parvalbumin-positive interneurons does not change suppression strength, but does alter its frequency-dependence. These results establish a role of cortical inhibition in forward suppression and link specific aspects of rapid sensory adaptation to genetically distinct interneuron types.

The organization in the primary auditory cortex (Au1) is critical to the basic function of auditory information processing and integration. However, recent mapping experiments using in vivo two-photon imaging with different Ca2+ indicators have reached controversial conclusions on this topic, possibly because of the different sensitivities and properties of the indicators used. Therefore, it is essential to identify a reliable Ca2+ indicator for use in in vivo functional imaging of the Au1, to understand its functional organization. Here, we demonstrate that a previously reported indicator, Cal-520, performs well in both anesthetized and awake conditions. Cal-520 shows a sufficient sensitivity for the detection of single action potentials, and a high signal-to-noise ratio. Cal-520 reliably reported on both spontaneous and sound-evoked neuronal activity in anesthetized and awake mice. After testing with pure tones at a range of frequencies, we confirmed the local heterogeneity of the functional organization of the mouse Au1. Therefore, Cal-520 is a reliable and useful Ca2+ indicator for in vivo functional imaging of the Au1.

The Mongolian gerbil (Meriones unguiculatus) has become a key species in investigations of the neural processing of sound localization cues in mammals. While its sound localization has been tested extensively under free-field stimulation, many neurophysiological studies use headphones to present signals with binaural localization cues. The gerbil's behavioral sensitivity to binaural cues, however, is unknown for the lack of appropriate stimulation paradigms in awake behaving gerbils. We close this gap in knowledge by mimicking a headphone stimulation; we use free-field loudspeakers and apply cross-talk cancellation techniques to present pure tones with binaural cues via "virtual headphones" to gerbils trained in a sound localization task. All gerbils were able to lateralize sounds depending on the interaural time or level difference (ITD and ILD, respectively). For ITD stimuli, reliable responses were seen for frequencies ≤2.9 kHz, the highest frequency tested with ITD stimuli. ITD sensitivity was frequency-dependent with the highest sensitivity observed at 1 kHz. For stimuli with ITD outside the gerbil's physiological range, responses were cyclic indicating the use of phase information when lateralizing narrow-band sounds. For ILD stimuli, reliable responses were obtained for frequencies ≥2 kHz. The comparison of ITD and ILD thresholds with ITD and ILD thresholds derived from gerbils' free-field performance suggests that ongoing ITD information is the main cue for sound localization at frequencies <2 kHz. At 2 kHz, ITD and ILD cues are likely used in a complementary way. Verification of the use of the virtual headphones suggests that they can serve as a suitable substitute for conventional headphones particularly at frequencies ≤2 kHz.

Hair cells in the mature cochlea cannot spontaneously regenerate. One potential approach for restoring hair cells is stem cell therapy. However, when cells are transplanted into scala media (SM) of the cochlea, they promptly die due to the high potassium concentration. We previously described a method for conditioning the SM to make it more hospitable to implanted cells and showed that HeLa cells could survive for up to a week using this method. Here, we evaluated the survival of human embryonic stem cells (hESC) constitutively expressing GFP (H9 Cre-LoxP) in deaf guinea pig cochleae that were pre-conditioned to reduce potassium levels. GFP-positive cells could be detected in the cochlea for at least 7 days after the injection. The cells appeared spherical or irregularly shaped, and some were aggregated. Flushing SM with sodium caprate prior to transplantation resulted in a lower proportion of stem cells expressing the pluripotency marker Oct3/4 and increased cell survival. The data demonstrate that conditioning procedures aimed at transiently reducing the concentration of potassium in the SM facilitate survival of hESCs for at least one week. During this time window, additional procedures can be applied to initiate the differentiation of the implanted hESCs into new hair cells.

2-Hydroxypropyl-Beta-Cyclodextrin (HPβCD) can be used to treat Niemann-Pick type C disease, Alzheimer's disease, and atherosclerosis. But, a consequence is that HPβCD can cause hearing loss. HPβCD was recently found to be toxic to outer hair cells (OHCs) in the organ of Corti. Previous studies on the chronic effects of in vivo HPβCD toxicity did not know the intra-cochlear concentration of HPβCD and attributed variable effects on OHCs to indirect drug delivery to the cochlea. We studied the acute effects of known HPβCD concentrations administered directly into intact guinea pig cochleae. Our novel approach injected solutions through pipette sealed into scala tympani in the cochlear apex. Solutions were driven along the length of the cochlear spiral toward the cochlear aqueduct in the base. This method ensured that therapeutic levels were achieved throughout the cochlea, including those regions tuned to mid to low frequencies and code speech vowels and background noise. A wide variety of measurements were made. Results were compared to measurements from ears treated with the HPβCD analog methyl-β-cyclodextrin (MβCD), salicylate that is well known to attenuate the gain of the cochlear amplifier, and injection of artificial perilymph alone (controls). Histological data showed that OHCs appeared normal after treatment with a low dose of HPβCD, and physiological data was consistent with attenuation of cochlear amplifier gain and disruption of non-linearity associated with transferring acoustic sound into neural excitation, an origin of distortion products that are commonly used to objectively assess hearing and hearing loss. A high dose of HPβCD caused sporadic OHC losses and markedly affected all physiologic measurements. MβCD caused virulent destruction of OHCs and physiologic responses. Toxicity of HPβCD to OHC along the cochlear length is variable even when a known intra-cochlear concentration is administered, at least for the duration of our acute studies.