Abstract
The cell neural adhesion molecule contactin-2 plays a key role in axon extension and guidance, fasciculation, and myelination during development. We thus asked, whether contactin-2 is also important in nervous system regeneration after trauma. In this study, we used an adult zebrafish spinal cord transection model to test the functions of contactin-2 in spinal cord regeneration. The expression patterns of contactin-2 at different time points after spinal cord injury were studied at the mRNA level by qPCR and in situ hybridization, and contactin-2 protein levels and immunohistological localization were detected by Western blot and immunofluorescence analyses, respectively. Contactin-2 mRNA and protein levels were increased along the central canal at 6 days and 11 days after spinal cord injury, suggesting a requirement for contactin-2 in spinal cord regeneration. Co-localization of contactin-2 and islet-1 (a motoneuron marker) was observed in spinal cords before and after injury. To further explore the functions of contactin-2 in regeneration, an anti-sense morpholino was used to knock down the expression of contactin-2 protein by application at the time of injury. Motion analysis showed that inhibition of contactin-2 retarded the recovery of swimming functions when compared to standard control morpholino. Anterograde and retrograde tracing at 6 weeks after injury showed that knock down of contactin-2 inhibited axonal regrowth from NMLF neurons beyond lesion site. The combined observations indicate that contactin-2 contributes to locomotor recovery and successful regrowth of axons after spinal cord injury in adult zebrafish.
Highlights
Contactin-2, called transient axonal glycoprotein-1 (TAG1) or axonin-1, belongs to the contactin, L1 and immunoglobulin superfamilies, with six immunoglobulin-like domains and four fibronectin type III homologous repeats [1]
To investigate the involvement of contactin-2 in spinal cord regeneration in adult zebrafish, the expression patterns of contactin-2 in the caudal part of spinal cord after injury were examined by real time Quantitative RT-PCR (qPCR) and in situ hybridization at different time points after complete spinal cord lesion
The mRNA levels of contactin-2 were determined by real time qPCR and normalized to the levels of GAPDH
Summary
Contactin-2, called transient axonal glycoprotein-1 (TAG1) or axonin-1, belongs to the contactin, L1 and immunoglobulin superfamilies, with six immunoglobulin-like domains and four fibronectin type III homologous repeats [1]. At formative stages of nervous system development, contactin-2 is involved mainly in neurogenesis. After the stage of abundant neurogenesis, contactin-2 becomes expressed by migrating neurons and outgrowing axons. In the rat GABA-expressing interneurons, originating in the medial ganglionic eminence of the ventral telencephalon, migrate along contactin-2-expressing axons of the developing corticofugal system to reach the dorsal telencephalon [5]. In the adult rodent brain, expression of contactin-2 is restricted to certain areas exhibiting persistent neural plasticity, e.g. olfactory bulb and hippocampus, and possibly cerebellar granule cells [10]. These features implicate contactin-2 in plasticity of the adult central nervous system of mammals
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