Substrate Composition and Mechanical Engagement in Neuron Regeneration

Development and regeneration of the nervous system relies on the interaction between growing neurites and the extracellular environment. Growth cones, the motile sensory tips of elongating neurites, interpret chemical cues and exert forces against external structures to form an intricate web of connections. We have used regenerating Aplysia californica bag cell neurons to investigate the mechanical interaction between growth cone and substrate and the effect of substrate composition on regeneration. We found that laminin and substrate-adsorbed hemolymph proteins from Aplysia have a synergistic effect on bag cell regeneration. This novel interaction promotes both outgrowth and branching, demonstrating that substrate composition can have a profound effect on regeneration. Forces exerted by the growth cone against their substrate can also influence regeneration. We used traction force microscopy to measure the force generated by migrating bag cell growth cones. We found that traction stresses are primarily transmitted to the substrate through the peripheral actin-rich domain. Calculation of net growth cone traction force revealed that neurites are under tension, although a substantial proportion of the force is balanced within the growth cone and does not contribute to tension. Measuring traction force over time revealed that stress patterns are extremely dynamic but tension levels are stable, suggesting that the neurite imposes a constant tension that is continuously redistributed among dynamic adhesions. Our observations suggest that peripheral traction may initiate directional change by bending the end of the neurite towards regions of high force. We also present evidence that extracellular proteins, in addition to serving as a growth-promoting adhesion platform, may be capable storing and generating force. The results presented here reveal many complexities in the interaction between neurites and their extracellular environment, which will provide the basis for future studies.