Neurotrophin-3 regulates ribbon synapse density in the cochlea and induces synapse regeneration after acoustic trauma

Neurotrophin-3 (Ntf3) and brain derived neurotrophic factor (Bdnf) are critical for sensory neuron survival and establishment of neuronal projections to sensory epithelia in the embryonic inner ear, but their postnatal functions remain poorly understood. Using cell-specific inducible gene recombination in mice we found that, in the postnatal inner ear, Bbnf and Ntf3 are required for the formation and maintenance of hair cell ribbon synapses in the vestibular and cochlear epithelia, respectively. We also show that supporting cells in these epithelia are the key endogenous source of the neurotrophins. Using a new hair cell CreERT line with mosaic expression, we also found that Ntf3's effect on cochlear synaptogenesis is highly localized. Moreover, supporting cell-derived Ntf3, but not Bbnf, promoted recovery of cochlear function and ribbon synapse regeneration after acoustic trauma. These results indicate that glial-derived neurotrophins play critical roles in inner ear synapse density and synaptic regeneration after injury. DOI: http://dx.doi.org/10.7554/eLife.03564.00

Concomitant differentiation of a population of mouse embryonic stem cells into neuron‐like cells and schwann cell–like cells in a slow‐flow microfluidic device

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/135245/1/dvdy24466_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/135245/2/dvdy24466.pd

A Model of Electrically Stimulated Auditory Nerve Fiber Responses with Peripheral and Central Sites of Spike Generation

A computational model of cat auditory nerve fiber (ANF) responses to electrical stimulation is presented. The model assumes that (1) there exist at least two sites of spike generation along the ANF and (2) both an anodic (positive) and a cathodic (negative) charge in isolation can evoke a spike. A single ANF is modeled as a network of two exponential integrateand-fire point-neuron models, referred to as peripheral and central axons of the ANF. The peripheral axon is excited by the cathodic charge, inhibited by the anodic charge, and exhibits longer spike latencies than the central axon; the central axon is excited by the anodic charge, inhibited by the cathodic charge, and exhibits shorter spike latencies than the peripheral axon. The model also includes subthreshold and suprathreshold adaptive feedback loops which continuously modify the membrane potential and can account for effects of facilitation, accommodation, refractoriness, and spike-rate adaptation in ANF. Although the model is parameterized using data for either single or paired pulse stimulation with monophasic rectangular pulses, it correctly predicts effects of various stimulus pulse shapes, stimulation pulse rates, and level on the neural response statistics. The model may serve as a framework to explore the effects of different stimulus parameters on psychophysical performance measured in cochlear implant listeners

Assessment and Preservation of Auditory Nerve Integrity in the Deafened Guinea Pig

Profound hearing loss is often caused by cochlear hair cell loss. Cochlear implants (CIs) essentially replace hair cells by encoding sound and conveying the signal by means of pulsatile electrical stimulation to the spiral ganglion cells (SGCs) which form the auditory nerve. SGCs progressively degenerate following hair cell loss, as a result of lost neurotrophic signaling from the hair cells. Degeneration of the auditory nerve may compromise the ability to hear with a CI. Therefore, the first goal of this thesis was to extend the current knowledge on this degeneration process, with emphasis on changes in functionality. To this end, normal hearing and deafened guinea pigs received a CI, and the responses of the auditory nerve to electrical stimulation (electrically evoked compound action potentials; eCAPs) were recorded. Several characteristics of the eCAP correlated well with SGC survival, suggesting that eCAP recordings in human CI users, too, may be indicative of the extent of degeneration of the auditory nerve. The ability to estimate the extent of neural loss in CI users may greatly advance current research into the effect of auditory nerve degeneration on sound perception with a CI. In animal models of severe hair cell loss, treatment with exogenous neurotrophic factors such as brain-derived neurotrophic factor (BDNF) has been shown to prevent secondary SGC degeneration. However, the functionality of the SGCs rescued by the treatment has not been properly evaluated. In order to provide a comprehensive assessment of SGC functionality after neurotrophic treatment, deafened guinea pigs were chronically implanted with an intracochlear electrode array combined with a cannula, through which BDNF was slowly infused into the cochlea over a period of four weeks. Histological analysis at the end of the treatment showed almost complete survival of SGCs; eCAP recordings indicated that SGC responsiveness to electrical stimulation was often similar to that in normal-hearing controls. In order to assess clinical practicability of BDNF treatment, the long-term effects of the four-week treatment were evaluated both at the histological and at the functional level. Sustained survival of SGCs was observed both four and eight weeks after cessation of the BDNF treatment. Moreover, near-normal SGC functionality was preserved during this eight week period as well. These findings are important for translation to clinical application, since continuous administration is impractical, and may even bring risk of tumorigenesis on the long term. In summary, the experimental work described in this thesis has led to the identification of useful electrophysiological measures with which the extent of auditory nerve degeneration can be estimated. It has additionally demonstrated that, in case of auditory nerve degeneration, treatment with neurotrophic factors prevent further degeneration, preserves the nerve’s functionality, and is clinically feasible

Efficacy of a new charge-balanced biphasic electrical stimulus in the isolated sciatic nerve and the hippocampal slice

Most deep brain stimulators apply rectangular monophasic voltage pulses. By modifying the stimulus shape, it is possible to optimize stimulus efficacy and find the best compromise between clinical effect, minimal side effects and power consumption of the stimulus generator. In this study, we compared the efficacy of three types of charge-balanced biphasic pulses (CBBPs, nominal duration 100 μs) in isolated sciatic nerves and in in vitro hippocampal brain slices of the rat. Using these two models, we tested the efficacy of several stimulus shapes exclusively on axons (in the sciatic nerve) and compared the effect with that of stimuli in the more complex neuronal network of the hippocampal slice by considering the stimulus-response relation. We showed that (i) adding an interphase gap (IPG, range 100-500 μs) to the CBBP enhances stimulus efficacy in the rat sciatic nerve and (ii) that this type of stimuli (CBBP with IPG) is also more effective in hippocampal slices. This benefit was similar for both models of voltage and current stimulation. In our two models, asymmetric CBBPs were less beneficial. Therefore, CBBPs with IPG appear to be well suited for application to DBS, since they enhance efficacy, extend battery life and potentially reduce harmful side effects

mTOR Signaling in BDNF-Treated Guinea Pigs after Ototoxic Deafening

The mammalian target of rapamycin (mTOR) signaling plays a critical role in cell homeostasis, growth and survival. Here, we investigated the localization of the main mTOR signaling proteins in the organ of Corti of normal-hearing and deafened guinea pigs, as well as their possible modulation by exogenously administered brain-derived neurotrophic factor (BDNF) in deafened guinea pigs. Animals were ototoxically deafened by systemic administration of kanamycin and furosemide, and one week later, the right cochleas were treated with gelatin sponge soaked in rhBDNF, while the left cochleas were used as negative controls. Twenty-four hours after treatment, animals were euthanized, and the cochleas were processed for subsequent analysis. Through immunofluorescence, we demonstrated the localization of AKT, pAKT, mTOR, pmTOR and PTEN proteins throughout the cochlea of guinea pigs for the first time, with a higher expression in supporting cells. Moreover, an increase in mTOR immunostaining was observed in BDNF-treated cochleas by means of fluorescence intensity compared to the other groups. Conversely, Western blot analysis showed no significant differences in the protein levels between groups, probably due to dilution of proteins in the neighboring tissues of the organ of Corti. Altogether, our data indicate that mTOR signaling proteins are expressed by the organ of Corti (with a major role for supporting cells) and that the modulation of mTOR may be a protective mechanism triggered by BDNF in the degenerating organ of Corti

No Protective Effects of Hair Cells or Supporting Cells in Ototoxically Deafened Guinea Pigs upon Administration of BDNF

We investigated whether treatment with brain-derived neurotrophic factor (BDNF), which is known to protect spiral ganglion cells (SGCs), could also protect hair cells (HCs) and supporting cells (SCs) in the organ of Corti of a guinea pig model of sensorineural hearing loss. Hearing loss was induced by administration of kanamycin/furosemide and two BDNF treatments were performed: (1) by gelatin sponge (BDNF-GS) with acute cochlear implantation (CI), and (2) through a mini-osmotic pump (BDNF-OP) with chronic CI. Outer HCs (OHCs), inner HCs (IHCs), Border, Phalangeal, Pillar, Deiters’, and Hensen’s cells were counted. The BDNF-GS cochleas had significantly fewer OHCs compared to the untreated ones, while the IHC and SC numbers did not differ between treated and untreated cochleas. The BDNF-OP group showed similar cell numbers to the untreated group. SGC packing density was not correlated with the total number of SCs for either BDNF group. Our data suggest that: (1) BDNF does not prevent cell death in the organ of Corti, and that the protection of SGCs could result from a direct targeting by BDNF; (2) BDNF might induce a different function/activity of the remaining cells in the organ of Corti (independently from cell number)