Netrin signal is produced in leech embryos by segmentally iterated sets of central neurons and longitudinal muscle cells

Abstract

Abstract. Netrins are secreted molecules capable of attracting or repelling growing axons. They and their receptors, along with other netrin-interacting proteins, are widely conserved among animals from a broad range of phyla. We have raised and purified an antibody against a recently cloned leech netrin, which has allowed us to characterize embryonic netrin expression by cells in peripheral tissues and in the central nervous system. During early gangliogenesis, netrin expression was detected at particularly high levels in five bilateral pairs of central neurons. Towards the end of the period of axonal outgrowth, netrin expression was observed to be restricted to only six central neurons, comprising two bilateral pairs and two unpaired cells. A pair of netrin-producing central neurons, the bipolar cells, was identified by their expression of the antigen recognized by the monoclonal antibody Laz1-1. Double staining of sensory afferents from segmental sensilla with the monoclonal antibody Lan3-2 and the bipolar cells with the netrin antibody revealed that the terminals of these afferents grow up to the bipolar cells and turn anteriorly or posteriorly, without extending any further medially. Peripheral netrin expression was found to be restricted to longitudinal muscle cells in the ventral half of the body wall. Extracellular, secreted netrin was detected in a broad longitudinal stripe located symmetrically with respect to the ventral midline. The pattern of expression of netrin in leech embryos is consistent with observed expression patterns in other animals, suggesting that developmental netrin functions are conserved among all bilateral animals.

Heterochronically expressed midline netrin was recruited to guide mesoderm migration in epibolic gastrulation of the leech

“…The expression pattern of a netrin gene in the developing CNS was previously characterized in the medicinal leech Hirudo and is in agreement with the conventional idea that 6 it serves as the midline landmark that guides the morphogenesis of CNS (Aisemberg et al, 2001;Gan et al, 1999). Nonetheless, netrin expression in earlier developmental stages remained unknown for the leech.…”

Molecular evidence for deep evolutionary roots of bilaterality in animal development

“…Netrins are a family of secreted guidance factors, which can attract or repulse migrating neurons and neuronal growth cones depending on the context of the signal and the particular receptor binding the netrin ligand (36) in a diverse array of bilaterians (36)(37)(38)(39)(40)(41). In protostomes, netrin is expressed at the ventral midline, and in the chordates, netrin is expressed in the ventral region of the dorsal nerve cord (the embryonic dorsal pole) (36)(37)(38)(39)(40)(41), which has led some authors to argue for D-V inversion of body plan organization in these groups (42). The N. vectensis ortholog of bilaterian netrin, NvNetrin (Fig.…”

Multiple Changes in Peptide and Lipid Expression Associated with Regeneration in the Nervous System of the Medicinal Leech

“…Thus, the detection of EMPAII [11], IL16 [12], C1q [13], [20] and the antimicrobial peptides [14] is expected. The second wave of molecules seems to be related to physical reconstruction and tissue repairing, with the up-regulation of cytoskeleton proteins and modulatory components like myohemerytin, filarin, gliarin, macrolin, leech CAM, and tractin [7], [23]–[27]. All of these proteins support the neuronal outgrowth that will be responsible for reconnecting neurons.…”

“…All of these proteins support the neuronal outgrowth that will be responsible for reconnecting neurons. Then neuronal sprouting and axon guidances proteins, like liprin, hillarin, leech Ena, netrin and syntaxin [23], [28]–[30] can be detected. This is then followed by a phase in which, during the course of tissue repair, signaling pathways are reestablished through the heightened expression of novel hemi-channels related to gap junctions formation (innexins), synapsin, ReN3, voltage-gated Na channels isoform [3], [21], [31], [32].…”