Cytoskeletal and Morphological Alterations Underlying Axonal Sprouting after Localized Transection of Cortical Neuron AxonsIn Vitro

Abstract

We examined the cytoskeletal dynamics that characterize neurite sprouting after axonal injury to cortical neurons maintained in culture for several weeks and compared these with initial neurite development. Cultured neocortical neurons, derived from embryonic day 18 rats, were examined at 3 d in vitro (DIV) and at various time points after axotomy at 21 DIV. The postinjury neuritic response was highly dynamic, progressing through an initial phase of retraction, followed by substantial axonal sprouting within 4-6 hr. Postinjury sprouts were motile and slender with expanded growth cone-like end structures. Microtubule markers were localized to sprout shafts and the proximal regions of putative growth cones and filamentous actin was distributed throughout growth cones, whereas neurofilament proteins were restricted to sprout shafts. A similar distribution of cytoskeletal proteins was present in developing neurites at 3 DIV. Exposure of developing and mature, injured cultures to the microtubule stabilizing agent taxol (10 microg/ml) caused growth inhibition, process distension, the transformation of growth cones into bulbous structures, and abnormal neurite directionality. Microtubule and neurofilament segregation occurred after taxol exposure in developing neurites and postinjury sprouts. Exposure to the microtubule destabilizing agent nocodazole (100 microg/ml) resulted in substantial morphological alteration of developing neurons and inhibited neurite growth and postinjury axonal sprouting. Our results indicate that the axons of cortical neurons have an intrinsic ability to sprout after transection, and similar cytoskeletal dynamics underlie neurite development and postinjury axonal sprouting.

Taxol Facilitates Axon Regeneration in the Mature CNS

“…1 B). These findings are in accordance with a previous report demonstrating that, at high concentrations, Taxol blocks neurite extension of cortical neurons in vitro (Chuckowree and Vickers, 2003). This depen- dence on a specific and low concentration range can be explained by the dynamic instability of microtubules and the influence of Taxol on the growing and shrinking rates of microtubules.…”

“…Taxol stabilizes microtubules and as a result interferes with the normal breakdown of microtubules during cell division (Vyas and Kadow, 1995). At low concentrations, Taxol alters microtubule dynamics and promotes polymerization at the plus end (Chuckowree and Vickers, 2003). Taxol can prevent the formation of axon retraction bulbs after spinal cord injury, and, in culture, it can overcome myelin inhibition of neurite outgrowth (Ertürk et al, 2007).…”

Microtubule stability and MAP1B upregulation control neuritogenesis in CAD cells

“…It was the first natural product shown to stabilize microtubules by formation of non-functional microtubule bundles. Drugs that disturb microtubule stability or satalilize microtubules, represented by nocodazole and taxol respectively, abrogate neurite outgrowth and axonal extension [3] . Therefore, the balance between microtubule stability and destabilization is essential for neurite development.…”

Phrenicotomy alters phrenic long-term facilitation following intermittent hypoxia in anesthetized rats

“…69). Studies of acute axotomy reveal relatively rapid changes in the morphology of mammalian neurons (13) and increases in intracellular Ca 2ϩ concentration in invertebrate (59) and vertebrate (45) axons. Accordingly, phrenic motoneuron excitability may be increased after PhrX.…”