Semaphorin4D promotes axon regrowth and swimming ability during recovery following zebrafish spinal cord injury

Semaphorins comprise a family of proteins involved in axon guidance during development. Semaphorin4D (Sema4D) has both neuroregenerative and neurorepressive functions, being able to stimulate both axonal outgrowth and growth cone collapse during development, and therefore could play an important role in neurological recovery from traumatic injury. Here, we used a zebrafish spinal cord transection model to study the role of Sema4D in a system capable of neuroregeneration. Real-time qPCR and in situ hybridization showed upregulated Sema4D expression in the acute response phase (within 3days post SCI), and downregulated levels in the chronic response phase (11-21days after SCI). Double-immunostaining for Sema4D and either Islet-1 (motoneuron marker) or Iba-1 (microglial marker) showed that microglia surrounded Sema4D-positive motoneurons along the central canal at 4h post injury (hpi) and 12hpi. Following administration of Sema4D morpholino (MO) to transected zebrafish, double-immunostaining showed that Sema4D-positive motoneurons surrounded by microglia decreased at 7days and 11days compared with standard control MO. Anterograde and retrograde tracing indicate that Sema4D participates in axon regeneration in the spinal cord following spinal cord injury (SCI) in the zebrafish. Swim tracking shows that MO-mediated inhibition of Sema4D retarded the recovery of swimming function when compared to standard control MO. The combined results indicate that Sema4D expression in motoneurons enhances locomotor recovery and axon regeneration, possibly by regulating microglia function, after SCI in adult zebrafish.