The manipulation of gene expression by RNA interference could play a key role in future neurotherapies, for example in the development of biohydrid implants to bridge nerve and spinal cord lesion gaps. Such resorbable biomaterial prostheses could serve as growth substrates together with specific siRNA to foster neuronal regeneration. To the best of our knowledge, we are the first to biofunctionalize neuronal prostheses with siRNA. We analyzed neuronal and Schwann cell responses to scrambled siRNA coated polydioxanone polymer filaments designed to imitate pro-regenerative bands of Bungner for oriented axonal regrowth. With a view to future clinical applications we were especially interested in potentially detrimental side effects. We employed a variety of in vitro methods, including a novel impedance electrode microchamber assay, fluorescence and scanning electron microscopy, metabolic labeling and RT-PCR. We found that the application of chitosan/siRNA nanoparticles (1) did not affect glial cell motility or (2) axonal growth in contrast to other formulations, (3) only slightly reduced proliferation, and (4) did not induce inflammatory responses that might hamper axonal regeneration. The data suggest that chitosan/siRNA nanoparticle-coated polymer filaments are suitable for use in biohybrid implants with no significant side effects on neuronal and glial cells.