N‐cadherin regulates primary motor axon growth and branching during zebrafish embryonic development

Abstract

N-cadherin is a classical type I cadherin that contributes to the formation of neural circuits by regulating growth cone migration and the formation of synaptic contacts. This study analyzed the role of N-cadherin in primary motor axons growth during development of the zebrafish (Danio rerio) embryo. After exiting the spinal cord, primary motor axons migrate ventrally through a common pathway and form the first neuromuscular junction with the muscle pioneer cells located at the horizontal myoseptum, which serves as a choice point for cell-type specific pathway selection. Analysis of N-cadherin mutants (cdh2hi3644Tg) and embryos injected with N-cadherin antisense morpholinos showed primary motor axons extending aberrant axonal branches at the choice point in ~40% of the somitic hemisegments, and an ~150% increase in the number of branches per axon length within the ventral myotome. Analysis of individual axons trajectories showed that the caudal (CaP) and rostral (RoP) motor neurons axons formed aberrant branches at the choice point which abnormally extended in the rostrocaudal axis and ventrally to the horizontal myoseptum. Expression of a dominant-interfering N-cadherin cytoplasmic domain in primary motor neurons caused some axons to abnormally stall at the horizontal myoseptum and to impair their migration into the ventral myotome. However, in N-cadherin depleted embryos the majority of primary motor axons innervated their appropriate myotomal territories indicating that N-cadherin regulates motor axon growth and branching without severely affecting the mechanisms that control axonal target selection.

Presynaptic architecture of the larval zebrafish neuromuscular junction

“…This technique has been established also for larval stages of the zebrafish D. rerio (Nixon et al, 2009;Schieber et al, 2010). We decided first to compare HPF/FS with classically fixed preparations of larval zebrafish NMJs because previous studies of these structures were based solely on chemical fixation (Waterman, 1969;Westerfield et al, 1990;Drapeau et al, 2001;Brus es, 2011;Denker et al, 2011). Additionally, electron tomography can be applied to both of these preparations to obtain a high resolution in 3D.…”

“…Furthermore, the zebrafish is an emerging model for understanding motoneuron disorders such as amyotrophic lateral sclerosis and spinal muscular atrophy (Kabashi et al, 2011;Babin et al, 2014;Patten et al, 2014). NMJs of the zebrafish have been analyzed by classical electron microscopy techniques (Waterman, 1969;Westerfield et al, 1990;Drapeau et al, 2001;Brus es, 2011;Denker et al, 2011). However, a systematic analysis of their 3D architecture with high resolution as provided by electron tomography is missing up to now.…”

β-Catenin gain of function in muscles impairs neuromuscular junction formation

“…It is possible that this mechanism is involved in early development of the NMJ. In fact, N-cadherin was recently shown to regulate primary motor axon growth and branching during zebrafish embryonic development (Bruses, 2011). It is worth pointing out that NMJ formation or function is apparently not altered by both -catenin LOF and GOF in motoneurons (supplementary material Fig.…”

Cadherin-Based Transsynaptic Networks in Establishing and Modifying Neural Connectivity

“…Early studies showed that many different types of cultured neurons exhibit robust axonal outgrowth when grown on a purified N-cadherin substrate, on astrocytes, or on various cell lines expressing N-cadherin on their surface (Bixby & Zhang, 1990; Doherty, Skaper, Moore, Leon, & Walsh, 1992; Matsunaga, Hatta, Nagafuchi, & Takeichi, 1988; Tomaselli et al, 1988), effects that are blocked in the presence of N-cadherin-neutralizing antibodies (Doherty, Rowett, Moore, Mann, & Walsh, 1991; Drazba & Lemmon, 1990; Neugebauer, Tomaselli, Lilien, & Reichardt, 1988) or interfering peptides targeting the conserved HAV sequence in Type I cadherins (Blaschuk, Sullivan, David, & Pouliot, 1990). More recent genetic analyses in Drosophila and zebrafish also provide strong evidence for N-cadherin and R-cadherin in axon extension, growth cone guidance, and targeting (Babb et al, 2005; Bruses, 2011; Lee, Herman, Clandinin, Lee, & Zipursky, 2001; Nern, Zhu, & Zipursky, 2008; Schwabe, Neuert, & Clandinin, 2013). Cadherin dominant-negative reagents have also been used to investigate cadherin-based outgrowth and targeting.…”