Two Stages in Neurite Formation Distinguished by Differences in Tubulin Metabolism

Abstract

Changes in tubulin solubility during neurite formation were studied biochemically using rat dorsal root ganglion neurons in culture. When fractionated with Ca" -containing buffer at low temperature, a considerable proportion of total cellular tubulin was recovered in the insoluble fraction . We designated this cold/Ca 2+insoluble tubulin (InsT) and distinguished it from cold/ Ca" -soluble tubulin (SoIT) . From the relative amount of InsT, neurite formation was found to proceed through two distinct stages . The first 6 days after plating (stage 1) in which the proportion of InsT increased dramatically (from 5 to 60%) coincided with neurite outgrowth. In the following period (stage 2), a constant level of InsT was maintained, whereas neurite maturation took place. Pulse-labeling experiments further revealed that the two stages differed significantly in terms of tubulin metabolism . High rates of synthesis as well as conversion from SoIT to InsT were observed in stage 1, whereas stage 2 was characterized by a decrease in both of these rates and an increase in the rate of degradation . The results show for the first time the coordinated changes in tubulin metabolism that underlie the process of neurite formation.

Selective accumulation of the high molecular weight neurofilament subunit within the distal region of growing axonal neurites

“…Axons of DRG elongated rapidly within the first 3 days of culture under these conditions, attaining a length of 240 Ϯ 10 m, and continued to elongate at a slower rate (i.e., to 250 Ϯ 10 m) by day 7 in cultures. As shown previously [e.g., Dahl, 1988;Sekimoto et al, 1995], this period (days 3-7) is accompanied by a transition of the axonal cytoskeleton from a form consistent with rapid elongation to one that imparts stability to axons, accomplished in part by the accumulation of phospho-NFs (Fig. 1).…”

Developmental and activity‐dependent regulation of ARMS/Kidins220 in cultured rat hippocampal neurons

“…The time frame of these changes in expression (high upon initial plating, and reaching a lower steady-state level after 12 days in culture) also correlates closely with a study of process maturation, which identified two stages of neurite growth on the basis of microtubule metabolism. The first stage, of active growth, occurred within the first week of plating in cultured dorsal root ganglion neurons, and the second stage, of maturation, occurred during the second week in culture (Sekimoto et al, 1995). Our data provide more evidence to support the role of ARMS/ Kidins220 in process outgrowth.…”