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Adeno-associated virus gene delivery targeting corneal-projecting trigeminal ganglion neurons

Year: 2026
Type: article
Abstract: Corneal innervation is essential for ocular surface homeostasis and protection, yet methods for selectively labeling and manipulating cornea-projecting neurons remain limited. Conventional retrograde tracers can identify cornea-projecting cell bodies in the trigeminal ganglion (TG), but they do not enable the expression of genetically encoded effector proteins, sensors, or recombinases. We describe and validate intrastromal adeno-associated virus (AAV) delivery as an approach for labeling cornea-projecting TG neurons in mice. A 1-2-μL viral injection was delivered into the corneal stroma and labeling was assessed 2-12 weeks later in cleared whole TG, TG sections, corneal whole mounts, and thick brainstem sections. Multiple AAV vectors, including tdTomato-, eGFP-, or mCherry-expressing vectors and Cre-expressing vectors in Ai14 tdTomato reporter mice, produced consistent neuronal labeling concentrated in the dorsomedial ophthalmic division of the TG. Native tdTomato fluorescence was readily detected, and eGFP labeling could be enhanced using immunolabeling with fluorophore-enhancing antibodies, increasing detection of labeled somata and axonal fibers. AAV-mediated labeling in the TG substantially overlapped with WGA and Fast Blue tracing from the cornea, although AAV labeled fewer neurons overall. Cre-expressing AAV vectors activated reporter expression in Ai14 mice, demonstrating compatibility with transgenic reporter strategies. Labeled axons were visualized at both peripheral and central projection sites. At the ocular surface, reporter expression was correlated to the extent of neuronal transduction. Central projections were observed in subnucleus principalis, the interpolaris/caudalis transition zone, and caudal subnucleus caudalis extending into upper cervical spinal cord. AAV transduction resulted in no gross immune activation. These findings indicate that intrastromal AAV delivery provides a versatile approach for anatomical tracing and genetic access to cornea-projecting neurons, enabling expression of genetically encoded tools for optogenetic and chemogenetic control, activity monitoring, and targeted genetic manipulation.
DOI: 10.3389/fncel.2026.1925806
Authors: Samuel S. Reynolds, Md Zeeshan Rasheed, Ken Niyonkuru, Sachin Parikh, Kyle Whyland, Brooklynn M. Merrill, William Perez, Sreya Mitra, Tomasz Zal, Hongmin Yun, Brian Davis, Anthony J. St. Leger, Anna Matynia, Ian D. Meng
Institutions: University of New England, University of Maine, University of New England, University of Houston, University of California, Los Angeles, University of Pittsburgh
Language: English
Subfield: Genetics
Open Access status: gold
Estimated APC paid: $3,801
Awards: Center Core Grant for Vision Research, Core Grant for Vision Research, Central and Peripheral Mechanisms of Corneal Pain, Vision Research Core at UCLA, Contribution of sympathetic nerves to herpes stromal keratitis, Assessing how ocular surface nerves, immune cells, and epithelial cells communicate to encourage neuro-immune homeostasis
OpenAlex ID: W7223188652
FWCI:
Cites: 81
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Related to: 10