Primary auditory neurons (PANs) play a critical role in hearing by transmitting sound information from the inner ear to the brain. Their progressive degeneration is associated with excessive noise, disease and aging. The loss of PANs leads to permanent hearing impairment since they are incapable of regenerating. Spiral ganglion non-neuronal cells (SGNNCs), comprised mainly of glia, are resident within the modiolus and continue to survive after PAN loss. These attributes make SGNNCs an excellent target for replacing damaged PANs through cellular reprogramming. We used the neurogenic pioneer transcription factor Ascl1 and the auditory neuron differentiation factor NeuroD1 to reprogram SGNNCs into induced neurons (iNs). The overexpression of both Ascl1 and NeuroD1 in vitro generated iNs at high efficiency. Transcriptome analyses revealed that iNs displayed a transcriptome profile resembling that of endogenous PANs, including expression of several key markers of neuronal identity: Tubb3, Map2, Prph, Snap25, and Prox1. Pathway analyses indicated that essential pathways in neuronal growth and maturation were activated in cells upon neuronal induction. Furthermore, iNs extended projections toward cochlear hair cells and cochlear nucleus neurons when cultured with each respective tissue. Taken together, our study demonstrates that PAN-like neurons can be generated from endogenous SGNNCs. This work suggests that gene therapy can be a viable strategy to treat sensorineural hearing loss caused by degeneration of PANs.

Edited by Roger J. ColbranSynaptic inhibition depends on a transmembrane gradient of chloride, which is set by the neuron-specific K ؉ -Cl ؊ co-transporter KCC2. Reduced KCC2 levels in the neuronal membrane contribute to the generation of epilepsy, neuropathic pain, and autism spectrum disorders; thus, it is important to characterize the mechanisms regulating KCC2 expression. In the present study, we determined the role of KCC2-protein interactions in regulating total and surface membrane KCC2 expression. Using quantitative immunofluorescence in cultured mouse hippocampal neurons, we discovered that the kainate receptor subunit GluK2 and the auxiliary subunit Neto2 significantly increase the total KCC2 abundance in neurons but that GluK2 exclusively increases the abundance of KCC2 in the surface membrane. Using a live cell imaging assay, we further determined that KCC2 recycling primarily occurs within 1-2 h and that GluK2 produces an ϳ40% increase in the amount of KCC2 recycled to the membrane during this time period. This GluK2-mediated increase in surface recycling translated to a significant increase in KCC2 expression in the surface membrane. Moreover, we found that KCC2 recycling is enhanced by protein kinase C-mediated phosphorylation of the GluK2 C-terminal residues Ser-846 and Ser-868. Lastly, using gramicidin-perforated patch clamp recordings, we found that the GluK2-mediated increase in KCC2 recycling to the surface membrane translates to a hyperpolarization of the reversal potential for GABA (E GABA ). In conclusion, our results have revealed a mechanism by which kainate receptors regulate KCC2 expression in the hippocampus.The classic fast hyperpolarizing inhibition of the mature brain results primarily from the activation of GABA A receptors.These receptors are Cl Ϫ -permeable ion channels, and inhibition results from the influx of Cl Ϫ into the neuron (1). This inward gradient for Cl Ϫ is set by the neuron-specific K ϩ /Cl Ϫ co-transporter KCC2 (2, 3). Despite the requirement of KCC2 for hyperpolarizing inhibition, the mechanisms that regulate KCC2 expression and function are still under intense investigation. In addition to the critical role KCC2 plays in synaptic inhibition, KCC2 is highly localized to excitatory synapses (4, 5), where it plays important roles in the development (6) and the function of glutamatergic synapses (7,8). In fact, single-particle tracking revealed that KCC2 is tightly confined to excitatory synapses (5), which may result from local protein interactions. Thus, understanding how proteins associated with excitatory synapses regulate KCC2 function may provide critical insight to the function of KCC2.KCC2 exists in a multiprotein complex and is regulated by components of excitatory synaptic transmission (9 -11). Specifically, KCC2 interacts with both the kainate-type ionotropic glutamate receptor subunit GluK2 (9) and its auxiliary subunit Neto2 (10). Neto2 regulates KCC2-mediated Cl Ϫ extrusion by binding to the active oligomeric form of KCC2 (10), whereas the GluK2-KCC2 i...

Disabling hearing loss affects over 5% of the world’s population and impacts the lives of individuals from all age groups. Within the next three decades, the worldwide incidence of hearing impairment is expected to double. Since a leading cause of hearing loss is the degeneration of primary auditory neurons (PANs), the sensory neurons of the auditory system that receive input from mechanosensory hair cells in the cochlea, it may be possible to restore hearing by regenerating PANs. A direct reprogramming approach can be used to convert the resident spiral ganglion glial cells into induced neurons to restore hearing. This review summarizes recent advances in reprogramming glia in the CNS to suggest future steps for regenerating the peripheral auditory system. In the coming years, direct reprogramming of spiral ganglion glial cells has the potential to become one of the leading biological strategies to treat hearing impairment.