Investigation of neurotrophic factor concentrations with a novel in vitro concept for peripheral nerve regeneration.

Abstract

The regeneration of nerves of the peripheral nervous system after injuries is a complex process. This study presents a novel in vitro neurite regeneration concept to investigate the regeneration of neurons and their processes with different concentrations of neurotrophic factors. The core part of the concept is a transparent microfluidic neurite isolation (NI) device affixed on top of a microelectrode array (MEA), providing a fast and easy way to assess both the growth and the electrical activity of neurites. The NI-MEA isolates neurites from the culture with microchannels that serve as guidance tubes, equipped with microelectrodes. Thus, the NI-MEA allows neurite growth, as observed by microscopy, to be correlated with neurite electrical activity, as measured by electrophysiological recordings. To demonstrate proof of concept of neurite regeneration, we cultured cells from the superior cervical ganglion of postnatal mice under different concentrations of nerve growth factor (NGF). During the regeneration process, we observed an increase in the number of neurites entering the microchannels along with an increase in spike activity recorded by the microelectrodes in the microchannels. We also observed a concentration-dependent effect of neurotrophic factor on the excitability of the growing neurites, with neurites bathed in 20 ng/ml NGF exhibiting enhanced early growth. Thus, our neurite regeneration concept with the NI-MEA device allows further study of neurotrophic factors and reduces the requirement for in vivo experiments on the regeneration of peripheral nerves after injury.