Background : Every year there are more than 15,000 new spinal cord injuries in the United States.1 Of those, a complete spinal cord transection, characterized by a lesion through both the motor and sensory tracts, is one of the most devastating injuries to patients. During the subacute and chronic phases of injury, glial cells in vertebrae species accumulate at the lesion site to provide a protective barrier through scar formation. Although the glial scar is imperative in protecting against additional tissue loss, it blocks axon regeneration, causing additional loss of function. A powerful model that lacks this glial scar formation is the zebrafish (Danio rerio). It’s genetic commonalities to humans and larval translucency make it one of the more resourceful models when studying new pharmacological therapeutics for spinal cord injury.2 4-Aminopyridine (4-AP) is a therapeutic that is inhibitory to voltage-gated potassium channels (Kv) allowing for increasing action potentials across demyelinated axons.3 It is essential to understand 4-AP’s therapeutic effect on glial cells spinal cord injury patients in the future and the zebrafish model allows a window to discover its role in glial cell proliferation, migration, and remodeling.

Methods: A complete spinal cord injury was performed on an Tg(gfap:EGFP) x Tg(elavl3:mcherry) transgenic zebrafish line five days post fertilization. On one day post injury, continuous dosing of E2 recovery buffer supplemented with either DMSO or 4-AP was given to an equal number of transected zebrafish and uninjured zebrafish. Utilizing the Leica Thunder System, Z stack images at 20x were taken from day of injury until five days post injury to analyze pixel intensity of the glial cell bridging to ensure a complete transection and progress of bridging. Zebrafish were fed one day during the recovery period and multi-well plates were replenished with new recovery buffer each day after imaging.

Results: Continuous dosing of 4-AP one-day post injury increased glial cell proliferation at both the rostral and caudal ends of the lesion site. The 4-AP zebrafish also had narrower injury sites by the fifth day post injury. The DMSO zebrafish showed higher proliferation at the rostral lesion site.

Conclusion: In closing, it was found that continuous dosing of 4-AP one day post injury enhances glial cell bridging, promoting regeneration of the spinal cord in the zebrafish model. Varying the dosing schedules and monitoring locomotion behaviors are needed to advance the knowledge of 4-AP's effects on regeneration after spinal cord injury.

References:

(Spinal Cord Injury Prevalence In The U.S. | Reeve Foundation (christopherreeve.org)).

Effectiveness of zebrafish models in understanding human diseases—A review of models - ScienceDirect)

Neuroprotective Properties of 4-Aminopyridine - PMC (nih.gov) Neuroprotective Properties of 4-Aminopyridine - PMC (nih.gov)

Building bridges, not walls: spinal cord regeneration in zebrafish - PMC (nih.gov)