Scale of Carbon Nanomaterials Affects Neural Outgrowth and Adhesion

Carbon nanomaterials have become increasingly popular microelectrode materials for neuroscience applications. Here we study how the scale of carbon nanotubes and carbon nanofibers affect neural viability, outgrowth, and adhesion. Carbon nanotubes were deposited on glass coverslips via a layer-by-layer method with polyethylenimine (PEI). Carbonized nanofibers were fabricated by electrospinning SU-8 and pyrolyzing the nanofiber depositions. Additional substrates tested were carbonized and SU-8 thin films and SU-8 nanofibers. Surfaces were O2-plasma treated, coated with varying concentrations of PEI, seeded with E18 rat cortical cells, and examined at 3, 4, and 7 days in vitro (DIV). Neural adhesion was examined at 4 DIV utilizing a parallel plate flow chamber. At 3 DIV, neural viability was lower on the nanofiber and thin film depositions treated with higher PEI concentrations which corresponded with significantly higher zeta potentials (surface charge); this significance was drastically higher on the nanofibers suggesting that the nanostructure may collect more PEI molecules, causing increased toxicity. At 7 DIV, significantly higher neurite outgrowth was observed on SU-8 nanofiber substrates with nanofibers a significant fraction of a neuron's size. No differences were detected for carbonized nanofibers or carbon nanotubes. Both carbonized and SU-8 nanofibers had significantly higher cellular adhesion post-flow in comparison to controls whereas the carbon nanotubes were statistically similar to control substrates. These data suggest a neural cell preference for larger-scale nanomaterials with specific surface treatments. These characteristics could be taken advantage of in the future design and fabrication of neural microelectrodes.