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Jiucheng He, Haydee E. P. Bazan; Neuroanatomy and Neurochemistry of Mouse Corneal Innervation. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3527.
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© ARVO (1962-2015); The Authors (2016-present)
The mouse is a popular model used for the study of corneal neurobiology because it is easy to manipulate genetically. However, the detailed nerve architecture and neuropeptide distribution of the mouse cornea remain unclear. Here we study the entire nerve architecture and sensory neuropeptide content of adult mouse corneas using a modified technique of immunofluorescence and imaging.
Adult ICR mice (8-weeks old) of both genders were euthanized and the whole corneas were excised and fixed. The corneas were stained with a neuronal-class β-tubulin III, calcitonin gene-related peptide (CGRP), and substance P (SP) antibodies, and whole-mount images were acquired to build a whole view of the corneal nerve architecture. To obtain the relative contents of the neuropeptides, the specimens were double stained with β-tubulin III. Relative nerve fiber densities for each fiber population were assessed on the basis of a whole mount view of the entire corneal innervation by computer-assisted analysis.
There was an average of 21.7 ± 3.5 (n = 20 eyes, Mean ± SD) thick stromal nerves that entered the cornea in a radial pattern and ran towards the anterior stroma, subsequently dividing into smaller branches. Some of the branches connected with each other to constitute the stromal nerve network, but most of them penetrated upwards into the epithelium to give place to the subbasal nerve bundles. Those bundles ran centripetally and converged into the central area to form the whorl-like structure. The number and pattern of whorl-like structures differed among eyes. Subbasal nerve density was 28.08% ± 2.80% in the central area and 18.04% ± 3.43% (N=10 eyes, P<0.05) in the peripheral area. Nerve terminals /mm2 were also greater in the center (3,995±301) than in the periphery (1989±482). Double staining showed that in the central zone (Φ=1.5 mm), CGRP-positive nerve fibers constituted 70.04 ± 9.23% of the total subbasal nerve contents, while SP-positive nerves constituted 58.85 ± 6.82% (N=5 eyes). In the same zone, CGRP positive terminals were 62.04 ± 4.94% of the total terminals, while SP took up 59.32 ± 3.80%.
This is the first study to show a complete map of the entire nerve architecture and the total sensory neuropeptide distribution of the mouse cornea. The normal innervation pattern described in this study will provide baseline data for researchers who use the mouse model to assess corneal nerve alterations.
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