Experimental and Computational Studies Reveal Novel Interaction of Lymphocytes Antigen 6K to TGF-ß Receptor Complex.

TGF-ß signaling promotes migration, invasion, and distant colonization of cancer cells in advanced metastatic cancers. TGF-ß signaling suppresses the anti-tumor immune response in a tumor microenvironment, allowing sustained tumor growth. TGF-ß plays an important role in normal physiology; thus it is no surprise that the clinical development of effective and safe TGF-ß inhibitors has been hampered due to their high toxicity. We discovered that increased expression of LY6K in cancer cells led to increased TGF-ß signaling and that inhibition of LY6K could lead to reduced TGF-ß signaling and reduced in vivo tumor growth. LY6K is a highly cancer-specific protein, and it is not expressed in normal organs except in the testes. Thus, LY6K is a valid target for developing therapeutic strategies to inhibit TGF-ß signaling in cancer cells. We employed in vitro pull-down assays and molecular dynamics simulations to understand the structural determinants of the TGF-ß receptor complex with LY6K. This combined approach allowed us to identify the critical residues and dynamics of the LY6K interaction with the TGF-ß receptor complex. These data are critical in designing novel drugs for the inhibition of TGF-ß in LY6K expressing cancer, induction of anti-tumor immune response, and inhibition of tumor growth and metastatic spread.

Neurotransmitter phenotype and axonal projection patterns of VIP-expressing neurons in the inferior colliculus.

Neurons in the inferior colliculus (IC), the midbrain hub of the central auditory pathway, send ascending and descending projections to other auditory brain regions, as well as projections to other sensory and non-sensory brain regions. However, the axonal projection patterns of individual classes of IC neurons remain largely unknown. Vasoactive intestinal polypeptide (VIP) is a neuropeptide expressed by subsets of neurons in many brain regions. We recently identified a class of IC stellate neurons that we called VIP neurons because they are labeled by tdTomato (tdT) expression in VIP-IRES-Cre x Ai14 mice. Here, using fluorescence in situ hybridization, we found that tdT+ neurons in VIP-IRES-Cre x Ai14 mice express Vglut2, a marker of glutamatergic neurons, and VIP, suggesting that VIP neurons use both glutamatergic and VIPergic signaling to influence their postsynaptic targets. Next, using viral transfections with a Cre-dependent eGFP construct, we labeled the axonal projections of VIP neurons. As a group, VIP neurons project intrinsically, within the ipsilateral and contralateral IC, and extrinsically to all the major targets of the IC. Within the auditory system, VIP neurons sent axons and formed axonal boutons in higher centers, including the medial geniculate nucleus and the nucleus of the brachium of the IC. Less dense projections terminated in lower centers, including the nuclei of the lateral lemniscus, superior olivary complex, and dorsal cochlear nucleus. VIP neurons also project to several non-auditory brain regions, including the superior colliculus, periaqueductal gray, and cuneiform nucleus. The diversity of VIP projections compared to the homogeneity of VIP neuron intrinsic properties suggests that VIP neurons play a conserved role at the microcircuit level, likely involving neuromodulation through glutamatergic and VIPergic signaling, but support diverse functions at the systems level through their participation in different projection pathways.

Downregulated miRNAs associated with auditory deafferentation and compensatory neural plastic changes following single-sided deafness in the inferior colliculi of rats.

BACKGROUND AND AIMS: Deafferentation and compensatory neural plastic changes in the inferior colliculus (IC) have been suggested following single-sided deafness (SSD). We explored related miRNA changes in the IC of SSD rats using miRNA microarray analyses. METHODS: Eight-week-old rats were divided into control and SSD rats (n = 8 for each group). SSD rats underwent right-side cochlear ablation surgery, with the IC harvested two weeks post-surgery. miRNA microarray analysis was performed using GeneChip miRNA 4.0, microarray (Affymetrix Inc.). miRNAs whose expression levels differed between SSD and control rats with a fold-change ≥ 1.5 and P < 0.05 were examined using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Target genes of differentially expressed miRNAs were predicted using TargetScan software. The pathways related to predicted target genes were analyzed. mRNA levels of predicted target genes were estimated using qRT-PCR. RESULTS: The expression of miR-15b-5p, miR-202-5p, and miR-212-3p was lower in the contralateral (left) IC of SSD rats than that of control rats. In SSD rats, miRNA expression levels in the contralateral IC were 0.45-, 0.25-, and 0.50-fold lower for miR-15b-5p, miR-202-5p, and miR-212-3p, respectively (P < 0.05). The expression of predicted target genes (Spred1, Rasa1, Lsm11, and Srsf1) was higher in the contralateral IC of SSD rats than in control rats. The targets were predicted to be related with cleavage of growing transcripts in the termination region, mitogen-activated protein kinase family signaling cascades, RAF/AMP kinase cascade, regulation of RAS by GTPase activating proteins (GAPs), and RNA polymerase II transcription termination. For ipsilateral ICs, miR-425-3p, miR-199a-5p, and miR-134-3p showed lower expressions in SSD rats than in control rats, which were 0.55-, 0.61-, and 0.69-fold lower, respectively (P < 0.05). The expression of predicted target genes (Atp2b2, Grin2b, Foxp1, Ztbt20, Zfp91, and Strn) was higher in the ipsilateral IC of SSD rats; the regulation of synaptic plasticity, cAMP signaling pathway, metal ion binding, and calcium ion transport can be associated with these target genes. CONCLUSION: Adult rats with unilateral auditory deprivation showed miRNA changes in the IC. The contralateral IC showed decreased miRNA expression predicted to be related to MAPK and RAS signaling, whereas the ipsilateral IC revealed decreased miRNA expression predicted to be associated with synaptic plasticity and calcium ion transport.

Tumoración en el colículo inferior como causa de acúfeno unilateral / Tumor at inferior colliculus causing unilateral tinnitus

Resumen La patología del sistema nervioso central, habitualmente, no provoca síntomas auditivos unilaterales, ya que la vía auditiva central está formada por una red de conexiones cruzadas entre los diferentes núcleos que la forman. Además, hay que considerar que una lesión pequeña puede extenderse a más de una estructura provocando varios déficits neurológicos debido a la proximidad de los tractos y núcleos nerviosos. Las lesiones unilaterales circunscritas en el colículo inferior son infrecuentes. No obstante, se han descrito casos en los que lesiones unilaterales de diversas etiologías en esta localización causaban síntomas auditivos. Ya que la vía auditiva central es cruzada, síntomas auditivos detectados con más frecuencia afectaban concretamente a la capacidad de localización del sonido o la comprensión verbal. Presentamos el caso de un hombre de 44 años con acúfeno unilateral derecho de larga evolución, sin otra clínica asociada quien fue diagnosticado de un tumor en el colículo inferior derecho mediante resonancia magnética cerebral. Se exponen los hallazgos clínicos y radiológicos del caso.

Abstract Central nervous system diseases usually do not cause auditory symptoms because the central auditory pathway consists on a network of crossed connections between the different nuclei that form it. In addition, we must consider that a small lesion might extend to more than one structure producing many neurologic symptoms due to the proximity of tracts and nuclei in the midbrain. Unilateral circumscribed lesions at inferior colliculus are rare. Nevertheless, lesions at this location causing auditory symptoms have been described. Because of the crossed central auditory pathway, the most commonly detected auditory symptoms specifically affected the ability to locate sound or verbal comprehension. We present the case of a 44-year-old man with a long-term monoaural right-sided tinnitus without other complaints who was diagnosed of a tumour at right inferior colliculus by neuroimaging. Clinical and radiological findings of this case are discussed.

α3ß4∗ Nicotinic Acetylcholine Receptors Strongly Modulate the Excitability of VIP Neurons in the Mouse Inferior Colliculus.

The inferior colliculus (IC), the midbrain hub of the central auditory system, receives extensive cholinergic input from the pontomesencephalic tegmentum. Activation of nicotinic acetylcholine receptors (nAChRs) in the IC can alter acoustic processing and enhance auditory task performance. However, how nAChRs affect the excitability of specific classes of IC neurons remains unknown. Recently, we identified vasoactive intestinal peptide (VIP) neurons as a distinct class of glutamatergic principal neurons in the IC. Here, in experiments using male and female mice, we show that cholinergic terminals are routinely located adjacent to the somas and dendrites of VIP neurons. Using whole-cell electrophysiology in brain slices, we found that acetylcholine drives surprisingly strong and long-lasting excitation and inward currents in VIP neurons. This excitation was unaffected by the muscarinic receptor antagonist atropine. Application of nAChR antagonists revealed that acetylcholine excites VIP neurons mainly via activation of α3ß4∗ nAChRs, a nAChR subtype that is rare in the brain. Furthermore, we show that acetylcholine excites VIP neurons directly and does not require intermediate activation of presynaptic inputs that might express nAChRs. Lastly, we found that low frequency trains of acetylcholine puffs elicited temporal summation in VIP neurons, suggesting that in vivo-like patterns of cholinergic input can reshape activity for prolonged periods. These results reveal the first cellular mechanisms of nAChR regulation in the IC, identify a functional role for α3ß4∗ nAChRs in the auditory system, and suggest that cholinergic input can potently influence auditory processing by increasing excitability in VIP neurons and their postsynaptic targets.

Sensitivity to interaural time differences in the inferior colliculus of cochlear implanted rats with or without hearing experience.

For deaf patients cochlear implants (CIs) can restore substantial amounts of functional hearing. However, binaural hearing, and in particular, the perception of interaural time differences (ITDs) with current CIs has been found to be notoriously poor, especially in the event of early hearing loss. One popular hypothesis for these deficits posits that a lack of early binaural experience may be a principal cause of poor ITD perception in pre-lingually deaf CI patients. This is supported by previous electrophysiological studies done in neonatally deafened, bilateral CI-stimulated animals showing reduced ITD sensitivity. However, we have recently demonstrated that neonatally deafened CI rats can quickly learn to discriminate microsecond ITDs under optimized stimulation conditions which suggests that the inability of human CI users to make use of ITDs is not due to lack of binaural hearing experience during development. In the study presented here, we characterized ITD sensitivity and tuning of inferior colliculus neurons under bilateral CI stimulation of neonatally deafened and hearing experienced rats. The hearing experienced rats were not deafened prior to implantation. Both cohorts were implanted bilaterally between postnatal days 64-77 and recorded immediately following surgery. Both groups showed comparably large proportions of ITD sensitive multi-units in the inferior colliculus (Deaf: 84.8%, Hearing: 82.5%), and the strength of ITD tuning, quantified as mutual information between response and stimulus ITD, was independent of hearing experience. However, the shapes of tuning curves differed substantially between both groups. We observed four main clusters of tuning curves - trough, contralateral, central, and ipsilateral tuning. Interestingly, over 90% of multi-units for hearing experienced rats showed predominantly contralateral tuning, whereas as many as 50% of multi-units in neonatally deafened rats were centrally tuned. However, when we computed neural d' scores to predict likely limits on performance in sound lateralization tasks, we did not find that these differences in tuning shapes predicted worse psychoacoustic performance for the neonatally deafened animals. We conclude that, at least in rats, substantial amounts of highly precise, "innate" ITD sensitivity can be found even after profound hearing loss throughout infancy. However, ITD tuning curve shapes appear to be strongly influenced by auditory experience although substantial lateralization encoding is present even in its absence.

Restoration of Deafferentation Reduces Tinnitus, Anxiety, and Depression: A Retrospective Study on Cochlear Implant Patients.

Patients with profound bilateral deafness (BD) are prone to suffering from tinnitus, which further leads to psychological comorbidities and makes it more difficult for patients to communicate with people. This study was aimed at investigating the effect of cochlear implants (CIs) on tinnitus distress and psychological comorbidities in patients with profound BD. This multicenter retrospective study reviewed 51 patients with severe postlingual BD who underwent cochlear implantation; 49 patients underwent unilateral cochlear implantation, and 2 patients underwent bilateral cochlear implantation. The patients were asked to complete all the questionnaires, including the tinnitus handicap inventory (THI), the visual analog scale (VAS) score, the Hospital Anxiety and Depression Scale Questionnaire (HADS), the Categories of Auditory Performance (CAP), and the Speech Intelligibility Rating (SIR), at least 4 months after implantation when the CI was on or off, in approximately May-June 2019. In our study, 94% (48/51) of BD patients suffered from tinnitus before CI, and 77% (37/48) of them suffered from bilateral tinnitus. In addition, 50.9% (26/51) of the CI patients were suffering from anxiety, 52.9% (27/51) of them were suffering from depression (score ≥ 8), and 66.7% (34/51) (27/51) of them were suffering from anxiety or depression. Cochlear implantation could reduce tinnitus more obviously when the CI was on than when the CI was off. Cochlear implantation also reduced anxiety/depression severity. There were significantly positive correlations between tinnitus severity and anxiety/depression severity before and after surgery. Moreover, hearing improvement is positively correlated with reduction level of tinnitus, the better hearing, and the lesser severity of tinnitus. Thus, along with effective restoration of deafferentation, cochlear implantation shows positive therapeutic effects on tinnitus and psychological comorbidities, providing a reference for future clinical and research work.

Purinergic Signaling Controls Spontaneous Activity in the Auditory System throughout Early Development.

Spontaneous bursts of electrical activity in the developing auditory system arise within the cochlea before hearing onset and propagate through future sound-processing circuits of the brain to promote maturation of auditory neurons. Studies in isolated cochleae revealed that this intrinsically generated activity is initiated by ATP release from inner supporting cells (ISCs), resulting in activation of purinergic autoreceptors, K+ efflux, and subsequent depolarization of inner hair cells. However, it is unknown when this activity emerges or whether different mechanisms induce activity during distinct stages of development. Here we show that spontaneous electrical activity in mouse cochlea from both sexes emerges within ISCs during the late embryonic period, preceding the onset of spontaneous correlated activity in inner hair cells and spiral ganglion neurons, which begins at birth and follows a base to apex developmental gradient. At all developmental ages, pharmacological inhibition of P2Y1 purinergic receptors dramatically reduced spontaneous activity in these three cell types. Moreover, in vivo imaging within the inferior colliculus revealed that auditory neurons within future isofrequency zones exhibit coordinated neural activity at birth. The frequency of these discrete bursts increased progressively during the postnatal prehearing period yet remained dependent on P2RY1. Analysis of mice with disrupted cholinergic signaling in the cochlea indicate that this efferent input modulates, rather than initiates, spontaneous activity before hearing onset. Thus, the auditory system uses a consistent mechanism involving ATP release from ISCs and activation of P2RY1 autoreceptors to elicit coordinated excitation of neurons that will process similar frequencies of sound.SIGNIFICANCE STATEMENT In developing sensory systems, groups of neurons that will process information from similar sensory space exhibit highly correlated electrical activity that is critical for proper maturation and circuit refinement. Defining the period when this activity is present, the mechanisms responsible and the features of this activity are crucial for understanding how spontaneous activity influences circuit development. We show that, from birth to hearing onset, the auditory system relies on a consistent mechanism to elicit correlate firing of neurons that will process similar frequencies of sound. Targeted disruption of this activity will increase our understanding of how these early circuits mature and may provide insight into processes responsible for developmental disorders of the auditory system.

Long-range Channelrhodopsin-assisted Circuit Mapping of Inferior Colliculus Neurons with Blue and Red-shifted Channelrhodopsins.

When investigating neural circuits, a standard limitation of the in vitro patch clamp approach is that axons from multiple sources are often intermixed, making it difficult to isolate inputs from individual sources with electrical stimulation. However, by using channelrhodopsin assisted circuit mapping (CRACM), this limitation can now be overcome. Here, we report a method to use CRACM to map ascending inputs from lower auditory brainstem nuclei and commissural inputs to an identified class of neurons in the inferior colliculus (IC), the midbrain nucleus of the auditory system. In the IC, local, commissural, ascending, and descending axons are heavily intertwined and therefore indistinguishable with electrical stimulation. By injecting a viral construct to drive expression of a channelrhodopsin in a presynaptic nucleus, followed by patch clamp recording to characterize the presence and physiology of channelrhodopsin-expressing synaptic inputs, projections from a specific source to a specific population of IC neurons can be mapped with cell type-specific accuracy. We show that this approach works with both Chronos, a blue light-activated channelrhodopsin, and ChrimsonR, a red-shifted channelrhodopsin. In contrast to previous reports from the forebrain, we find that ChrimsonR is robustly trafficked down the axons of dorsal cochlear nucleus principal neurons, indicating that ChrimsonR may be a useful tool for CRACM experiments in the brainstem. The protocol presented here includes detailed descriptions of the intracranial virus injection surgery, including stereotaxic coordinates for targeting injections to the dorsal cochlear nucleus and IC of mice, and how to combine whole cell patch clamp recording with channelrhodopsin activation to investigate long-range projections to IC neurons. Although this protocol is tailored to characterizing auditory inputs to the IC, it can be easily adapted to investigate other long-range projections in the auditory brainstem and beyond.

Hemangioblastoma of Cerebral Aqueduct Removed via Sitting, Supracerebellar Intracollicular Approach.

BACKGROUND: Tumors protruding into the cerebral aqueduct are rare, and tumors arising from within the cerebral aqueduct are rarer still. CASE DESCRIPTION: In this report, we discuss the presentation and clinical outcome of a 65-year-old man who presented to us with symptoms of hydrocephalus. Prior imaging had revealed a small enhancing nodule within the cerebral aqueduct. In the 6 months between initial imaging and our seeing the patient, the tumor demonstrated substantial interval growth, so the patient was offered resection. The tumor was accessed using a sitting, supracerebellar, intracollicular approach, which allowed for gross total resection of the mass without complication. Histopathology later revealed the lesion to be a hemangioblastoma. Two years after surgery, the patient was doing well with no neurologic deficits. CONCLUSIONS: We report the first case of an aqueductal hemangioblastoma and describe our use of a sitting, supracerebellar, intracollicular approach to access tumors occupying this cerebrospinal fluid space.

A novel class of inferior colliculus principal neurons labeled in vasoactive intestinal peptide-Cre mice.

Located in the midbrain, the inferior colliculus (IC) is the hub of the central auditory system. Although the IC plays important roles in speech processing, sound localization, and other auditory computations, the organization of the IC microcircuitry remains largely unknown. Using a multifaceted approach in mice, we have identified vasoactive intestinal peptide (VIP) neurons as a novel class of IC principal neurons. VIP neurons are glutamatergic stellate cells with sustained firing patterns. Their extensive axons project to long-range targets including the auditory thalamus, auditory brainstem, superior colliculus, and periaqueductal gray. Using optogenetic circuit mapping, we found that VIP neurons integrate input from the contralateral IC and the dorsal cochlear nucleus. The dorsal cochlear nucleus also drove feedforward inhibition to VIP neurons, indicating that inhibitory circuits within the IC shape the temporal integration of ascending inputs. Thus, VIP neurons are well-positioned to influence auditory computations in a number of brain regions.

Relationship between auditory pathway fractional anisotropy and audibility in patients with cerebellopontine angle tumors on diffusion tensor imaging.

OBJECTIVE: Recent reports demonstrated that acoustic nerve disorders affect the auditory pathway on diffusion tensor imaging (DTI). The aim was to investigate whether auditory pathway fractional anisotropy (FA) values are associated with audibility in patients with cerebellopontine angle tumors. PATIENTS AND METHODS: Patients with cerebellopontine angle tumors were included in this retrospective study. Preoperatively, all patients underwent magnetic resonance imaging (MRI) including DTI. Two regions of interest on the lateral lemniscus (LL) and inferior colliculus (IC) were set bilaterally on DTI. FA values were calculated using software. Correlations between FA values and audibility were evaluated using Spearman's rank correlation coefficient. Statistical significance was defined as p < 0.05. RESULTS: Seventeen patients with cerebellopontine angle tumors were included in this study. FA values in the bilateral LL showed a significant negative correlation with hearing impairment severity (r = -0.758, -0.600, p < 0.05). FA values on the ipsilateral side of the IC showed a significant negative correlation with hearing impairment severity (r = -0.477, p < 0.05). FA values on the contralateral side of the IC did not correlate with hearing impairment severity (r = -0.201, p > 0.05). One patient with a low FA value on the contralateral side of the IC had postoperative hearing impairment despite good preoperative hearing ability. CONCLUSIONS: FA values in the bilateral LL and on the ipsilateral side of the IC reflected hearing impairment severity. Decreased FA values on the contralateral side of the IC might predict hearing impairment postoperatively.

A characterization of laminar architecture in mouse primary auditory cortex.

Laminar architecture of primary auditory cortex (A1) has long been investigated by traditional histochemical techniques such as Nissl staining, retrograde and anterograde tracings. Uncertainty still remains, however, about laminar boundaries in mice. Here we investigated the cortical lamina structure by combining neuronal tracing and immunofluorochemistry for laminar specific markers. Most retrogradely labeled corticothalamic neurons expressed Forkhead box protein P2 (Foxp2) and distributed within the laminar band of Foxp2-expressing cells, identifying layer 6. Cut-like homeobox 1 (Cux1) expression in layer 2-4 neurons divided the upper layers into low expression layers 2/3 and high expression layers 3/4, which overlapped with the dense terminals of vesicular glutamate transporter 2 (vGluT2) and anterogradely labeled lemniscal thalamocortical axons. In layer 5, between Cux1-expressing layers 2-4 and Foxp2-defined layer 6, retrogradely labeled corticocollicular projection neurons mostly expressed COUP-TF interacting protein 2 (Ctip2). Ctip2-expressing neurons formed a laminar band in the middle of layer 5 distant from layer 6, creating a laminar gap between the two laminas. This gap contained a high population of commissural neurons projecting to contralateral A1 compared to other layers and received vGluT2-immunopositive, presumptive thalamocortical axon collateral inputs. Our study shows that layer 5 is much wider than layer 6, and layer 5 can be divided into at least three sublayers. The thalamorecipient layers 3/4 may be separated from layers 2/3 using Cux1 and can be also divided into layer 4 and layer 3 based on the neuronal soma size. These data provide a new insight for the laminar structure of mouse A1.

Changes to Neural Activation Patterns (c-fos Labeling) in Chinchilla Auditory Midbrain following Neonatal Exposure to an Enhanced Sound Environment.

Sensory brain regions show neuroplastic changes following deficits or experimental augmentation of peripheral input during a neonatal period. We have previously shown reorganization of cortical tonotopic maps after neonatal cochlear lesions or exposure to an enhanced acoustic environment. Such experiments probe the cortex and show reorganization, but it is unclear if such changes are intrinsically cortical or reflect projections from modified subcortical regions. Here, we ask whether an enhanced neonatal acoustic environment can induce midbrain (inferior colliculus (IC)) changes. Neonatal chinchillas were chronically exposed to a 70 dB SPL narrowband (2 ± 0.25 kHz) sound stimulus for 4 weeks. In line with previous studies, we hypothesized that such exposure would induce widening of the 2 kHz tonotopic map region in IC. To probe c-fos expression in IC (central nucleus), sound-exposed and nonexposed animals were stimulated with a 2 kHz stimulus for 90 minutes. In sound-exposed subjects, we find no change in the width of the 2 kHz tonotopic region; thus, our hypothesis is not supported. However, we observed a significant increase in the number of c-fos-labeled neurons over a broad region of best frequencies. These data suggest that neonatal sound exposure can modify midbrain regions and thus change the way neurons in IC respond to sound stimulation.

Sensory overamplification in layer 5 auditory corticofugal projection neurons following cochlear nerve synaptic damage.

Layer 5 (L5) cortical projection neurons innervate far-ranging brain areas to coordinate integrative sensory processing and adaptive behaviors. Here, we characterize a plasticity in L5 auditory cortex (ACtx) neurons that innervate the inferior colliculus (IC), thalamus, lateral amygdala and striatum. We track daily changes in sound processing using chronic widefield calcium imaging of L5 axon terminals on the dorsal cap of the IC in awake, adult mice. Sound level growth functions at the level of the auditory nerve and corticocollicular axon terminals are both strongly depressed hours after noise-induced damage of cochlear afferent synapses. Corticocollicular response gain rebounded above baseline levels by the following day and remained elevated for several weeks despite a persistent reduction in auditory nerve input. Sustained potentiation of excitatory ACtx projection neurons that innervate multiple limbic and subcortical auditory centers may underlie hyperexcitability and aberrant functional coupling of distributed brain networks in tinnitus and hyperacusis.

Increased spontaneous firing rates in auditory midbrain following noise exposure are specifically abolished by a Kv3 channel modulator.

Noise exposure has been shown to produce long-lasting increases in spontaneous activity in central auditory structures in animal models, and similar pathologies are thought to contribute to clinical phenomena such as hyperacusis or tinnitus in humans. Here we demonstrate that multi-unit spontaneous neuronal activity in the inferior colliculus (IC) of mice is significantly elevated four weeks following noise exposure at recording sites with frequency tuning within or near the noise exposure band, and this selective central auditory pathology can be normalised through administration of a novel compound that modulates activity of Kv3 voltage-gated ion channels. The compound had no statistically significant effect on IC spontaneous activity without noise exposure, nor on thresholds or frequency tuning of tone-evoked responses either with or without noise exposure. Administration of the compound produced some reduction in the magnitude of evoked responses to a broadband noise, but unlike effects on spontaneous rates, these effects on evoked responses were not specific to recording sites with frequency tuning within the noise exposure band. Thus, the results suggest that modulators of Kv3 channels can selectively counteract increases in spontaneous activity in the auditory midbrain associated with noise exposure.

Subcortical sources dominate the neuroelectric auditory frequency-following response to speech.

Frequency-following responses (FFRs) are neurophonic potentials that provide a window into the encoding of complex sounds (e.g., speech/music), auditory disorders, and neuroplasticity. While the neural origins of the FFR remain debated, renewed controversy has reemerged after demonstration that FFRs recorded via magnetoencephalography (MEG) are dominated by cortical rather than brainstem structures as previously assumed. Here, we recorded high-density (64 ch) FFRs via EEG and applied state-of-the art source imaging techniques to multichannel data (discrete dipole modeling, distributed imaging, independent component analysis, computational simulations). Our data confirm a mixture of generators localized to bilateral auditory nerve (AN), brainstem inferior colliculus (BS), and bilateral primary auditory cortex (PAC). However, frequency-specific scrutiny of source waveforms showed the relative contribution of these nuclei to the aggregate FFR varied across stimulus frequencies. Whereas AN and BS sources produced robust FFRs up to ∼700 Hz, PAC showed weak phase-locking with little FFR energy above the speech fundamental (100 Hz). Notably, CLARA imaging further showed PAC activation was eradicated for FFRs >150 Hz, above which only subcortical sources remained active. Our results show (i) the site of FFR generation varies critically with stimulus frequency; and (ii) opposite the pattern observed in MEG, subcortical structures make the largest contribution to electrically recorded FFRs (AN ≥ BS > PAC). We infer that cortical dominance observed in previous neuromagnetic data is likely due to the bias of MEG to superficial brain tissue, underestimating subcortical structures that drive most of the speech-FFR. Cleanly separating subcortical from cortical FFRs can be achieved by ensuring stimulus frequencies are >150-200 Hz, above the phase-locking limit of cortical neurons.

Specific immediate early gene expression induced by high doses of salicylate in the cochlear nucleus and inferior colliculus of the rat / Expressão específica de genes precoces imediatos induzida por doses elevadas de salicilato no núcleo coclear e colículo inferior de rato

Abstract Introduction: Salicylate at high doses induces tinnitus in humans and experimental animals. However, the mechanisms and loci of action of salicylate in inducing tinnitus are still not well known. The expression of Immediate Early Genes (IEG) is traditionally associated with long-term neuronal modifications but it is still not clear how and where IEGs are activated in animal models of tinnitus. Objectives: Here we investigated the expression of c-fos and Egr-1, two IEGs, in the Dorsal Cochlear Nucleus (DCN), the Inferior Colliculus (IC), and the Posterior Ventral Cochlear Nucleus (pVCN) of rats. Methods: Rats were treated with doses known to induce tinnitus in rats (300 mg/kg i.p. daily, for 3 days), and c-fos and Egr-1 protein expressions were analyzed using western blot and immunocytochemistry. Results: After administration of salicylate, c-fos protein expression increased significantly in the DCN, pVCN and IC when assayed by western blot. Immunohistochemistry staining showed a more intense labeling of c-fos in the DCN, pVCN and IC and a significant increase in c-fos positive nuclei in the pVCN and IC. We did not detect increased Egr-1 expression in any of these areas. Conclusion: Our data show that a high dose of salicylate activates neurons in the DCN, pVCN and IC. The expression of these genes by high doses of salicylate strongly suggests that plastic changes in these areas are involved in the genesis of tinnitus.

Resumo Introdução: Salicilato em doses elevadas induz zumbido nos seres humanos e em animais experimentais. No entanto, os mecanismos e loci de ação do salicilato na indução de zumbido ainda não são bem conhecidos. A expressão dos genes precoces imediatos (GPIs) está tradicionalmente associada a alterações neuronais em longo prazo, mas ainda não está claro como e onde os GPIs são ativados em modelos animais de zumbido. Objetivos: No presente estudo investigamos a expressão de c-fos e Egr-1, dois GPIs, no núcleo coclear dorsal (NCD), colículo inferior (CI) e núcleo coclear ventral posterior (NCVp) de ratos. Métodos: Os ratos foram tratados com doses que, conhecidamente, induzem zumbido em ratos (300 mg/kg IP/dia, por três dias) e as expressões das proteínas c-fos e Egr-1 foram analisadas por meio de Western blot e imunoistoquímica. Resultados: Após a administração de salicilato, a expressão da proteína c-fos aumentou significativamente no NCD, NCVp e CI, quando analisados por Western blot. A coloração imunoistoquímica mostrou uma marcação mais intensa de c-fos no NCD, NCVp e CI e um aumento significativo de núcleos positivos de c-fos no NCVp e CI. Não detectamos aumento da expressão de Egr-1 em qualquer dessas áreas. Conclusão: Nossos dados mostram que uma dose alta de salicilato ativa neurônios no NCD, NCVp e CI. A expressão desses genes por doses altas de salicilato sugere que as alterações plásticas nessas áreas estão envolvidas na gênese do zumbido.

Auditory processing assessment suggests that Wistar audiogenic rat neural networks are prone to entrainment.

Epilepsy is a neurological disease related to the occurrence of pathological oscillatory activity, but the basic physiological mechanisms of seizure remain to be understood. Our working hypothesis is that specific sensory processing circuits may present abnormally enhanced predisposition for coordinated firing in the dysfunctional brain. Such facilitated entrainment could share a similar mechanistic process as those expediting the propagation of epileptiform activity throughout the brain. To test this hypothesis, we employed the Wistar audiogenic rat (WAR) reflex animal model, which is characterized by having seizures triggered reliably by sound. Sound stimulation was modulated in amplitude to produce an auditory steady-state-evoked response (ASSR; -53.71Hz) that covers bottom-up and top-down processing in a time scale compatible with the dynamics of the epileptic condition. Data from inferior colliculus (IC) c-Fos immunohistochemistry and electrographic recordings were gathered for both the control Wistar group and WARs. Under 85-dB SLP auditory stimulation, compared to controls, the WARs presented higher number of Fos-positive cells (at IC and auditory temporal lobe) and a significant increase in ASSR-normalized energy. Similarly, the 110-dB SLP sound stimulation also statistically increased ASSR-normalized energy during ictal and post-ictal periods. However, at the transition from the physiological to pathological state (pre-ictal period), the WAR ASSR analysis demonstrated a decline in normalized energy and a significant increase in circular variance values compared to that of controls. These results indicate an enhanced coordinated firing state for WARs, except immediately before seizure onset (suggesting pre-ictal neuronal desynchronization with external sensory drive). These results suggest a competing myriad of interferences among different networks that after seizure onset converge to a massive oscillatory circuit.

A novel BK channel-targeted peptide suppresses sound evoked activity in the mouse inferior colliculus.

Large conductance calcium-activated (BK) channels are broadly expressed in neurons and muscle where they modulate cellular activity. Decades of research support an interest in pharmaceutical applications for modulating BK channel function. Here we report a novel BK channel-targeted peptide with functional activity in vitro and in vivo. This 9-amino acid peptide, LS3, has a unique action, suppressing channel gating rather than blocking the pore of heterologously expressed human BK channels. With an IC50 in the high picomolar range, the apparent affinity is higher than known high affinity BK channel toxins. LS3 suppresses locomotor activity via a BK channel-specific mechanism in wild-type or BK channel-humanized Caenorhabditis elegans. Topical application on the dural surface of the auditory midbrain in mouse suppresses sound evoked neural activity, similar to a well-characterized pore blocker of the BK channel. Moreover, this novel ion channel-targeted peptide rapidly crosses the BBB after systemic delivery to modulate auditory processing. Thus, a potent BK channel peptide modulator is open to neurological applications, such as preventing audiogenic seizures that originate in the auditory midbrain.