Inner ear stem cells can be isolated by neurosphere formation from the vestibular organs and the cochlea. The cells are pluripotent, with the potential to become hair cells and neurons, the cochlear cell types whose loss causes deafness. Here we describe the control of cell fate decisions that determine the phenotype adopted by these progenitors, and we determine whether differentiation to sensory neurons is preferred over other types of neurons. Differentiation of progenitor cells recapitulated developmental pathways of embryonic sensory neurons. Based on marker expression, retinoic acid increased the yield of neurons and the percentage of sensory neurons obtained and caused a sharp increase in Pax2, a key transcription factor of cranial placodes. Markers of embryonic auditory and other sensory neurons, GATA3, Brn3a, and islet1, could be detected after 3 days of differentiation of the cells, and markers of the sensory phenotype, peripherin, calretinin, TrkC, and TrkB were expressed after 10 days. The differentiated cells had tetrodotoxin-sensitive sodium currents and fired action potentials, and recordings revealed functional AMPA type-glutamate receptors, further indicating that these cells had developed neuronal features. Neurons differentiated from these stem cells grew processes to hair cells in vitro. Development of functional activity in cells with the markers of sensory neurons suggested that the inner ear stem cells might have the capacity to replace cells lost due to neural degeneration.

Hearing loss can be caused by primary degeneration of spiral ganglion neurons or by secondary degeneration of these neurons after hair cell loss. The replacement of auditory neurons would be an important step in any attempt to restore auditory function in patients with damaged inner ear neurons or hair cells. Application of beta-bungarotoxin, a toxin derived from snake venom, to an explant of the cochlea eradicates spiral ganglion neurons while sparing the other cochlear cell types. The toxin was found to bind to the neurons and to cause apoptotic cell death without affecting hair cells or other inner ear cell types as indicated by TUNEL staining, and, thus, the toxin provides a highly specific means of deafferentation of hair cells. We therefore used the denervated organ of Corti for the study of neuronal regeneration and synaptogenesis with hair cells and found that spiral ganglion neurons obtained from the cochlea of an untreated newborn mouse reinnervated hair cells in the toxin-treated organ of Corti and expressed synaptic vesicle markers at points of contact with hair cells. These findings suggest that it may be possible to replace degenerated neurons by grafting new cells into the organ of Corti.

We observed that perspective 3D-volume rendering of the cochlea enables the determination of key features of cochlear morphology and orientation that may escape detection with routine computed tomographic scanning. Infants and young toddler candidates demonstrate greater variability in the dimensions of the cochlear base and in the orientation of the cochlea within the cranium. As evolving surgical techniques and device design enhance the ability of the surgeon to avoid cochlear damage and optimize electrode location, refined morphometric information may assist the surgeon in tailoring strategies of scala tympani implantation.