Axon Stretch Growth: The Mechanotransduction of Neuronal Growth

Abstract

During pre-synaptic embryonic development, neuronal processes traverse short distances to reach their targets via growth cone. Over time, neuronal somata are separated from their axon terminals due to skeletal growth of the enlarging organism (Weiss 1941;Gray, Hukkanen et al. 1992). This mechanotransduction induces a secondary mode of neuronal growth capable of accommodating continual elongation of the axon (Bray 1984;Heidemann and Buxbaum 1994;Heidemann, Lamoureux et al. 1995;Pfister, Iwata et al. 2004).Axon Stretch Growth (ASG) is conceivably a central factor in the maturation of short embryonic processes into the long nerves and white matter tracts characteristic of the adult nervous system. To study ASG in vitro, we engineered bioreactors to apply tension to the short axonal processes of neuronal cultures (Loverde, Ozoka et al. 2011). Here, we detail the methods we use to prepare bioreactors and conduct ASG. First, within each stretching lane of the bioreactor, neurons are plated upon a micro-manipulated towing substrate. Next, neurons regenerate their axonal processes, via growth cone extension, onto a stationary substrate. Finally, stretch growth is performed by towing the plated cell bodies away from the axon terminals adhered to the stationary substrate; recapitulating skeletal growth after growth cone extension.Previous work has shown that ASG of embryonic rat dorsal root ganglia neurons are capable of unprecedented growth rates up to 10mm/day, reaching lengths of up to 10cm; while concurrently resulting in increased axonal diameters (Smith, Wolf et al. 2001;Pfister, Iwata et al. 2004;Pfister, Bonislawski et al. 2006;Pfister, Iwata et al. 2006;Smith 2009). This is in dramatic contrast to regenerative growth cone extension (in absence of mechanical stimuli) where growth rates average 1mm/day with successful regeneration limited to lengths of less than 3cm (Fu and Gordon 1997;Pfister, Gordon et al. 2011). Accordingly, further study of ASG may help to reveal dysregulated growth mechanisms that limit regeneration in the absence of mechanical stimuli.

The mechanical control of nervous system development

“…Development 140 (15) growth (Weiss, 1941;Bray, 1984;Loverde et al, 2011). Pfister et al showed that mechanical tension induces extreme 'stretch growth' of integrated axon tracts at remarkable rates and extents (8 mm/day) (Pfister et al, 2004), indicating that axonal lengthening is mainly limited by tension, or rather its relative absence.…”

Role of cytoskeleton in axonal regeneration after neurodegenerative diseases and CNS injury

“…Most mechanical signals are transmitted through the cytoskeleton (Aguilar et al, 2013), which may involve the change of the entire cellular structure, the activation of enzymes or gene expression, and induce morphological and biochemical changes in the cytoskeleton subsequently. In addition, some results hinted that the absence of mechanical stimuli could lead to growth limitation of the growth cone in regeneration (Loverde et al, 2011). Thus, it may become the main challenge in the future to identify the role of mechanical signals in cytoskeleton regulation during axonal regeneration in the CNS.…”

Engineering Approaches for Creating Skeletal Muscle

“…It has been demonstrated that when neurons are cultured in such a way that the nerve body and the axon are attached to different, sliding surfaces, the axons can be lengthened by slowly moving the surfaces apart. 69 This phenomenon is known as axon stretch growth. The neurons could be stretched at 42 mm per hour.…”

“…The neurons could be stretched at 42 mm per hour. 69 Therefore, neurons are definitely capable of compensating for applied strains through axonal elongation. Other researchers have found that neurite outgrowth is increased by cyclic equibiaxial stretching.…”