Alignment, zippering and splaying of microtubules during axogenesis

Microtubules are reconfigured from a scattered array to a paraxial bundle during axogenesis. One popular model posits that this reconfiguration results from the assembly dynamics of the microtubules together with structural proteins that stabilize and crosslink them. Here we show that the microtubules undergo behaviors that are not consistent with a model based on these principles alone. When cultured on polylysine, rat sympathetic neurons extend modest lamellae which contain a mass of nonaligned microtubules. Application of a mixture of growth factors called matrigel results in a rapid expansion of the lamellae followed by the outgrowth of axons. Fluorescence‐imaging of living neurons reveals sluggish movements of microtubules in neurons not exposed to matrigel. After addition of matrigel, the movements become rapid and result in dramatic changes in the microtubule array. Microtubules now extend to the periphery of the lamellae where they invade newly forming axons. The microtubules align with one another and relative to the cell cortex, and draw together into bundles. Microtubules within a bundle move apart as well, particularly in the growth cones that form at the tips of developing axons. These microtubule behaviors, which we refer to as alignment, zippering and splaying, are reminiscent of those observed in vitro when microtubules are subjected to forces generated by molecular motor proteins. High‐resolution images obtained with electron microscopy support this view, and also suggest that microtubule reconfiguration occurs in conjunction with alterations in actin, which may be a substrate for microtubule movements. We conclude that the reconfiguration of microtubules underlying axogenesis involves complex microtubule behaviors, many of which are consistent with movements generated by motor‐driven forces.