(648d) Combined Physical and Biochemical Cues Direct the Growth of Inner Ear Neurites

Cochlear implant (CI) devices help to restore hearing to individuals who suffer from sensorineural hearing loss. While CIs provide basic auditory perception, implant patients only experience limited voice and tonal perception. Due to spatial separation between the electrode array and the neural receptors, significant current spread and nonspecific stimulation of the spiral ganglion neurons (SGNs), or inner ear neurons, occurs. Thus, devices with advanced electrical design are unable to recapitulate the tonotopic stimulation of neurons because of spatial resolution. Strategies to decrease the distance between CI electrodes and primary neural receptors have focused on regenerating the SGN neurites. To be effective, the spatial order of the neurites must remain intact and thus cellular cues must orient and guide the pathfinding of SGN neurites. In this work, physical and biochemical cues are used to direct the regrowth of SGN neurites. Utilizing the inherent temporal and spatial control of photopolymerization, physical microgrooves are fabricated using a photomask in a single step process. The spatial control of the photon-induced reaction forms ridges under transparent band and depressions under reflective regions with channel amplitudes ranging from 250 nm to 10 µm. Biochemical patterns are generated by adsorbing laminin, a cell adhesion protein, to acrylate polymer surfaces followed by irradiation through a photomask with UV light to deactivate protein in exposed areas and generate parallel biochemical patterns. Laminin showed deactivation with increased exposure time as measured by immunofluorescence. Adding photoinitiator decreased the time required to deactivate the adsorbed laminin. To evaluate neural response to the patterns, SGNs were first cultured on physically patterned substrates and showed increased alignment with higher amplitude and lower periodicity polymers. SGNs were also cultured on biochemical patterns and showed excellent alignment to the parallel bands confirming the photodeactivation process was effective in creating active laminin patterns. To further evaluate the ability of biochemical and physical features to be used in overcoming conflicting cellular cues, competing patterns were tested. Physical microgroove channels were first fabricated followed by a perpendicular laminin protein pattern. The strength of the physical cue was independently varied by changing the amplitude or the periodicity. Lower periodicity, or more frequent, surfaces induced a greater degree of alignment along the channels. Higher amplitudes, or deeper channel depths, also increased the guidance of the SGNs. When SGN neurites were seeded onto substrates with perpendicular biochemical and physical patterns, alignment was strongly dependent on both amplitude and periodicity. At low amplitudes neurites tended to grow along the laminin stripes. As the amplitude was decreased, alignment was disrupted and neurite segments followed neither the biochemical or physical pattern. Similarly, at lower periodicity, SGN neurites were not strongly aligned to either pattern. As the periodicity was increased, alignment shifted to follow the biochemical laminin pattern. These results demonstrate the ability of photopolymerized micro-features to modulate alignment of inner ear neurites even in the presence of conflicting physical and biochemical cues laying the groundwork for next generation cochlear implants and neural prosthetic devices.