Transgenic corneal neurofluorescence in mice: a new model for in vivo investigation of nerve structure and regeneration.
OBJECTIVE
To quantify the level of neuron-specific fluorescence in the corneas of transgenic mice expressing yellow fluorescent protein (YFP) driven by the thy1 promoter and examine the viability of using thy1-YFP mice as a model for studying nerve regeneration in vivo.


METHODS
The structure of corneal innervation in thy1-YFP mice visible with reporter gene fluorescence was compared with that visible with traditional immunofluorescence techniques. The percentage of corneal nerves with YFP fluorescence in wholemounted corneas and trigeminal neuron cultures was determined. Regeneration of fluorescent corneal neuronal processes after wounding was monitored in vivo.


RESULTS
In the mouse cornea, neuron-specific immunostaining determined that nerves enter the stroma in several bundles that then extend throughout the entire cornea. These stromal nerve bundles form a subbasal plexus beneath the corneal epithelium. Fine nerves from this plexus travel superficially to the ocular surface. Neuron-specific expression of YFP allowed visualization of nearly all large nerve bundles of the stroma but only some of the many finer nerves of the subbasal plexus and surface. In the subbasal nerve plexus, 46% of total neuronal processes exhibited YFP neurofluorescence. In vitro, 22% of cultured trigeminal neurons exhibited YFP neurofluorescence. After corneal nerve transection, nerve processes distal to the site of injury degenerated, whereas those proximal to the site regenerated in a pattern different from original nerve architecture.


CONCLUSIONS
Thy1-YFP mice display neurofluorescence and provide a novel model for monitoring the patterning, injury, and growth of corneal nerves in vivo.
Yu
Charles Q
CQ
Department of Ophthalmology and Vision Science, University of California-Davis School of Medicine, 4860 Y Street, Sacramento, CA 95817, USA.
Rosenblatt
Mark I
MI
D000818:Animals; D003315:Cornea / Q000294:innervation*; D065306:Corneal Injuries; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation / Q000502:physiology; D008164:Luminescent Proteins / Q000235:genetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D023421:Models, Animal; D009412:Nerve Fibers / Q000502:physiology; D009416:Nerve Regeneration / Q000502:physiology*; D009474:Neurons / Q000166:cytology; D009882:Ophthalmic Nerve / Q000033:anatomy & histology* / Q000502:physiology*; D012668:Trigeminal Ganglion / Q000502:physiology; D014949:Wounds, Nonpenetrating / Q000503:physiopathology
D008164:Luminescent Proteins