Embryonic Xenopus neurites integrate and respond to simultaneous electrical and adhesive guidance cues

Nerve cells detect and respond to multiple extrinsic guidance cues during development and regeneration using a motile growth cone. Navigational decisions may be required of the growth cone when different guidance cues are encountered simultaneously. We have tested the relative potencies of two opposing cues by presenting Xenopus spinal cord nerve cells growing on a micropatterned laminin culture substratum with an orthogonal DC electric field. Substrata composed of repeating 25-micron laminin tracks and spaces failed to influence the position of neuritogenesis from nerve cell soma. Once established, however, growth cone movement was constrained by laminin tracks such that neurites of 65% of cells were aligned after 5 h in vitro. Two hours after the application of a 100-140 mV/mm DC field the majority of cells remained aligned with the laminin tracks. Around 70% of Xenopus neurites normally orient cathodally on homogenous laminin substrata; therefore the galvanotropic response was impeded by prior exposure to a patterned laminin substrate. However, a proportion of aligned neurites did orient cathodally and evidence of a response to both directional cues was even found within the same cell. Video-enhanced contrast, differential interference contrast (VEC-DIC) microscopy was used to examine the detailed behavior of growth cones on micropatterned laminin substrata. The present study has demonstrated that growth cones can detect and integrate at least two morphogenetic guidance cues simultaneously. The strength of the galvanotropic response in Xenopus growth cones, however, was often insufficient to override established adhesive guidance in this model system.