Traction Force Microscopy Experiments to Measure Axonal Dynamics During Neuronal Growth

To perform neuronal activities and form functional networks, neurons have to extend to their target destination through axonal outgrowth and guidance. A quantitative description of axonal extension is crucial for a deep understanding of the development of the nervous system. This understanding also empowers new techniques to regenerate and stimulate the outgrowth of neurons after injuries and other neural diseases. The axonal outgrowing process is directed by the growth cone, which generates traction forces by adhering to the extracellular matrix (ECM). These traction forces are generated inside the cell by the collective work of stress fibers, actin filaments, myosin, and motor proteins that help to anchor the neuron to the surrounding ECM. Traction forces are then transmitted to the ECM through focal adhesions. In this work, we use traction force microscopy (TFM) to measure traction forces exerted by the growth cones of cortical neurons cultured on poly-acrylamide hydrogels. By analyzing the displacements of the fluorescence beads embedded in the substrate we are able to measure the traction stress fields exerted by growing axons. Our results show that when axons exert forces on the substrate, neurons get stiffened and that neuronal growth is directed by a contact guidance mechanism, in which axons are guided by external mechanical signals. This works highlights the importance of biomechanical interactions in neuronal growth and opens new insights into medical applications and nerve regeneration.