Wallerian degeneration of zebrafish trigeminal axons in the skin is required for regeneration and developmental pruning

Abstract

SUMMARYFragments of injured axons that detach from their cell body break down by the molecularly regulated process of Wallerian degeneration (WD). Although WD resembles local axon degeneration, a common mechanism for refining neuronal structure, several previously examined instances of developmental pruning were unaffected by WD pathways. We used laser axotomy and time-lapse confocal imaging to characterize and compare peripheral sensory axon WD and developmental pruning in live zebrafish larvae. Detached fragments of single injured axon arbors underwent three stereotyped phases of WD: a lag phase, a fragmentation phase and clearance. The lag phase was developmentally regulated, becoming shorter as embryos aged, while the length of the clearance phase increased with the amount of axon debris. Both cell-specific inhibition of ubiquitylation and overexpression of the Wallerian degeneration slow protein (Wld S ) lengthened the lag phase dramatically, but neither affected fragmentation. Persistent Wld S -expressing axon fragments directly repelled regenerating axon branches of their parent arbor, similar to self-repulsion among sister branches of intact arbors. Expression of Wld S also disrupted naturally occurring local axon pruning and axon degeneration in spontaneously dying trigeminal neurons: although pieces of Wld S -expressing axons were pruned, and some Wld S -expressing cells still died during development, in both cases detached axon fragments failed to degenerate. We propose that spontaneously pruned fragments of peripheral sensory axons must be removed by a WD-like mechanism to permit efficient innervation of the epidermis.

Live Imaging of Calcium Dynamics during Axon Degeneration Reveals Two Functionally Distinct Phases of Calcium Influx

“…Calcium dysregulation contributes to axon and dendrite degeneration during developmental pruning (Kanamori et al, 2013;Yang et al, 2013), as a result of injury (Schlaepfer and Bunge, 1973;Ziv and Spira, 1993;Ouardouz et al, 2003;Mishra et al, 2013;Villegas et al, 2014), and in neurodegenerative diseases (Mattson, 2007;Berridge, 2011). Wallerian degeneration (WD) occurs in axon fragments that have been separated from their cell body by injury (Waller, 1850) and is a Figure 1.…”

“…For example, chelating extracellular calcium (Schlaepfer and Bunge, 1973;Mishra et al, 2013), blocking voltage-gated calcium channels (George et al, 1995;Knöferle et al, 2010), or inhibiting calcium release from intracellular stores (Ouardouz et al, 2003;Villegas et al, 2014) all delay axon fragmentation. Conversely, increasing calcium promotes axon degeneration (Schlaepfer, 1977;Glass et al, 1994;George et al, 1995;Knöferle et al, 2010;Mishra et al, 2013). Despite the key role that calcium plays in WD, when and where it functions to promote fragmentation is unclear.…”

No simpler than mammals: axon and dendrite regeneration in Drosophila: Figure 1.

“…Albeit infrequently, lower vertebrates and invertebrates have been used for some time in neuronal injury studies, and these studies demonstrate the potential lower organisms have for revealing insights into nerve regeneration (Spira et al 2003;Martin et al 2010;Chen et al 2011). Because of their large size and reliable growth ability, neurons from Aplysia californica and squid have been a powerful model in studying how neurites respond to injury in vivo (Krause et al 1994;Spira et al 2003;Erez et al 2007).…”

“…Furthermore, now that laser-assisted axotomy can be facilitated with customized microfluidic devices, C. elegans is poised for high-throughput genetic and pharmacological studies (Guo et al 2008;Ghannad-Rezaie et al 2012). Such axotomy procedures have also been used in other model organisms, such as zebrafish and Drosophila (Sugimura et al 2003;O'Brien et al 2009;Martin et al 2010). In general, these studies are consistent with the dogma that axons in these lower organisms are all capable of regeneration.…”

Differential protection of neuromuscular sensory and motor axons and their endings in Wld mutant mice

“…The fusion boundaries bridge the com-plete sequence of one native gene (Nmnat-1) and the partial, N-terminal sequence of another (Ufd2, now re-ferred to as its mammalian homologue Ube4b) (Lyon et al, 1993;Coleman et al, 1998;Conforti et al, 2000). Trans genic expression of this chimaeric candidate gene con-firmed its strong neuroprotective phenotype (Mack et al,2001;Adalbert et al, 2005;Hoopfer et al, 2006;MacDonald et al, 2006;Martin et al, 2010). Both principal com-ponents of the Wld S protein contain short peptide se-quences that are necessary for axon protection, including the catalytic site for Nmnat enzymic activity and a valosin-containing peptide-binding motif in the N-terminus of Ube4b .…”