Electrospun hyaluronic acid-carbon nanotube nanofibers for neural engineering

Abstract Electroactive biomaterials enable the delivery of an electrical stimulus to modulate neuronal activity in neural engineering treatments spanning neurological disorders, pain management, and regenerative medicine. Our research focused on developing a biocompatible conductive material composite consisting of an ultra-low concentration of carbon nanotubes (CNT) within hyaluronic acid (HA) nanofibers. We hypothesized that electrical stimulation through composite nanofibers would improve neurite outgrowth by providing a substrate that is both permissive to neuron growth and to electronic to ionic conduction that could activate voltage sensitive regenerative pathways. We evaluated the electrical properties of the composite material through impedance spectroscopy and cyclic voltammetry. Mechanics and surface characteristics were evaluated through atomic force microscopy using quantitative nanomechanical mapping. A custom stimulation chamber was fabricated and electrical stimulus parameters, using a charge-balanced biphasic square wave, required to elicit increased neurite outgrowth were determined. Neurons cultured on stimulated substrates of HA-CNT nanofibers exhibited significantly longer growth than unstimulated HA-CNT and HA control substrates. The HA-CNT material could be translated to other regenerative medicine applications including skin and cardiac tissue. Statement of significance This article describes a unique conductive biomaterial consisting of hyaluronic acid, combined with multi-walled carbon nanotubes, electrospun into a nanofiber composite. We have shown that the biomaterial's electrical properties can be improved (decreased impedance and increased charge capacity) by adding less than 0.01% carbon nanotubes. By using this ultra-low concentration, our material is biocompatible and mechanically appropriate for nerve regeneration. While electrical stimulation has been previously shown to increase neurite outgrowth, electrically conductive materials are typically non-degradable, brittle and/or mechanically stiff. We have optimized stimulation voltage and duration to activate voltage sensitive ion channels and increase nerve growth and provided robust mechanical characterization to show the synergistic effect of material properties and electrical stimulation on neuron behavior.