27. A Novel Rodent Model for Neurotrophic Keratopathy: Retro-orbital Nasociliary Neurotomy

PURPOSE: Corneal nerves are essential for epithelial healing. Neurotrophic Keratopathy (NK) results from the loss of corneal innervation, leading to ulceration, scarring, and blindness. Current NK (Neurotrophic Keratopathy) animal models use highly invasive methods for ophthalmic neurolysis (i.e., intracranial electrocautery) that result in incomplete corneal denervation and frequent mortality. Therefore, we created a novel rodent NK model that is minimally invasive, reliable, straightforward, and allows the potential for more precise control over the degree of corneal denervation over previous models. METHODS: In wild-type rats, a supra-tarsal incision provided retro-orbital access to isolate the nasociliary branch of the ophthalmic nerve supplying the cornea. Using this approach, the nasociliary nerve was transected at its divergence from the ophthalmic nerve. The same procedure was replicated in transgenic mice expressing Green Fluorescent Protein (GFP) in corneal nerves and tdTomato (red fluorescent protein) in corneal Schwann cells. After the surgery, the corneal blink reflex was used as an indicator of sensory loss. β-III immunostaining of the corneal nerves was employed to confirm denervation. In-vivo fluorescence imaging was employed in these mice to monitor corneal innervation and Schwann cell survival before and after denervation. Daily fluorescein imaging was used to assess corneal healing after wounding the denervated corneas. RESULTS: All rats (n=10) and mice (n=10) fully recovered from surgery with no mortality and showed an absence of the blink reflex. Both β-III immunostaining and in vivo fluorescence imaging revealed absence of corneal nerves by day 3 post-denervation. In-vivo fluorescence imaging in Sox10-tdT mice showed reduction in corneal Schwann cells on day 3 post-denervation. Fluorescein imaging identified a delay in corneal epithelial healing consistent with corneal denervation. CONCLUSION: The retro-orbital and peripheral approach of nasociliary nerve neurotomy is a simple, minimally invasive technique for corneal denervation in mice and rats. It results in complete corneal denervation, allows the possibility of precise control over the degree of denervation, and avoids mortality.