Microtubule stability decreases axon elongation but not axoplasm production.

Microtubules are a primary cytoskeletal constituent of axons and growth cones. In addition to serving as a scaffolding for axon assembly, they also provide a means of transport of organelles that are essential for outgrowth and maintenance of synaptic function. Pharmacological manipulations that disrupt net assembly of microtubules also interfere with growth cone advance and axon extension. Less is known after the effects of disrupting microtubule dynamics without affecting net assembly. To investigate this, we studied the effects of low doses of nocodazole on axon extension and microtubule organization in rat superior cervical ganglion neurons. We report that 165-330 nM nocodazole significantly reduces axon extension rate and increases axon diameter without affecting the rate of production of axoplasm or microtubule polymer, and without decreasing the average length or number of microtubules. Two observations suggested that microtubule dynamics were depressed by this dose of nocodazole. First, this treatment eliminated the highly divergent lengths and positions of microtubules characteristic of normal growth cones, inducing an array in which each microtubule terminated at roughly the same position in the proximal regions of the growth cone. Second, there was a decrease in the proportion of microtubule length containing mostly tyrosinated (newly assembled) alpha-tubulin and an increase in the proportion of microtubule length containing mostly acetylated (older, more stable) alpha-tubulin. Together, these data suggest that a decrease in dynamic instability of microtubules is sufficient to disrupt axon extension but does not interfere with axoplasm production.