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Targeted viral vector transduction of peripheral nerves with fluorescent reporters for in vivo visualization of nerve-to-nerve interactions
Suresh Mohan, MD, Mass. Eye and Ear/ Harvard Medical School, Boston, MA, Iván Coto Hernández, PhD, Mass Eye and Ear, Boston, MA, Luk H Vandenberghe, PhD, Schepens Eye Research Institute/Mass Eye and Ear, Boston, MA and Nate Jowett, MD, Facial Nerve Center - Dept. of Otolaryngology, Harvard Medical School / Massachusetts Eye and Ear, Boston, MA

INTRODUCTION

Though end-to-side techniques are commonly employed in peripheral nerve surgery, the topography of regenerating axons at such junctures has not been well characterized. Herein, we employ viral vectors encoding distinct fluorescent reporter proteins to visualize axonal behavior at nerve-to-nerve coaptation sites in a rodent model.

MATERIALS & METHODS

Whisker pad musculature of Lewis rats was injected with recombinant adeno-associated virus carrying GFP (right-side) or TdTomato (left-side) transgenes for retrograde transduction of facial nerve branches. One week later, end-to-side cross-facial coaptation of the proximal right buccal branch to the intact left buccal branch was performed using sciatic nerve interposition isografts. End-to-side neurorrhaphy sites were serially imaged in vivo over six weeks using two-photon excitation microscopy.

RESULTS

Targeted transduction of right and left facial nerves with distinct fluorescent reporter proteins is described. Three-dimensional axonal topography at end-to-side coaptation sites of rodent motor nerves is characterized.

CONCLUSIONS

We report novel means for in vivo visualization of distinct peripheral nerve branches in an animal model. This approach is useful for characterizing axonal regeneration at nerve junctures and may prove advantageous for study of axon guidance.


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