Live Imaging of Axon Stretch Growth in Embryonic and Adult Neurons

Abstract

Strategies for nervous system repair arise from knowledge of growth mechanisms via a growth cone. The distinctive process of axon stretch growth is a robust, long-term growth that may reveal new pathways to accelerate nerve repair. Here, a live imaging bioreactor was engineered to closely explore cellular events initiated by applied tension. The stretch growth potential between adult and embryonic dorsal root ganglion (DRG) neurons was investigated, an important difference in nerve repair. Embryonic axons were capable of unidirectional stretch growth rates of 4?mm/d and reliably reached 4?cm in length within 2 weeks. Adult axons could only reach 2?mm/d and took over 3 weeks to reach 4?cm. Utilizing time-lapse imaging, we observed growth cone motility in coordination with stretch growth. Upon initiation of stretching, growth cones retracted. However, within 10?h of continuous stretching, growth cones extended at a rate of 0.2?mm/d opposite the direction of applied tension, contributing to overall axon elongation. We analyzed fast mitochondrial transport under increasing levels of strain to determine the effect of stretch on axonal transport. Transport began to diminish at 24% strain, and was almost completely absent at 39% strain. Surprisingly, axons recovered and were capable of subsequent stretch growth. When tension was completely released (?5% strain), stretch grown axons retracted at rates up to 6.1??m/sec and slowed as resting tension was restored. This ability to assess the process of axon stretch growth in real time will allow detailed study of how tension can be used to drive axonal growth and retraction.