Cytoskeletal dynamics in response to tensile loading of mammalian axons

Abstract

In response to an applied tensile load, axons of cultured neurons exhibit a number of morphological responses. We designed and implemented a cell stretching device to study the cellular mechanisms governing these responses. Rat sensory neurons were seeded onto a flexible silicone substrate and imaged during substrate stretch. The positions of stationary mitochondria, docked to the axonal cytoskeleton, were determined before and after 10% stretch, and used to calculate the resulting "instantaneous" strain in regions of the axon. There was dramatic heterogeneity in strain along the length of the stretched axons, particularly in regions shorter than 20 μm. The substrate was then held at 10% strain and the axons imaged for 20 min during "relaxation." Both strain magnitude and variability were larger at small lengths in stretched axons during the initial phase of relaxation, but after 14 min, decreased to levels smaller than those seen in unstretched axons. Mitochondrial pairs in stretched axons showed uncoordinated movement with each other at all lengths, suggesting that cytoskeletal cohesion is reduced after stretch. Collectively, these data present the axonal cytoskeleton as a dynamic structure, which responds to stretch rapidly and locally. Globally, the axon behaves as a viscoelastic continuum. Below a characteristic length, though, it appears to behave as a series of independent linked elements, each with unique mechanical properties which suggests a length scale within which cytoskeletal structural elements may be altered to modulate the biomechanical response of the axon. Finally, testable hypotheses of strain accomodation in the axon are suggested.