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Jiucheng He, Thang Luong PHAM, Azucena H Kakazu, Haydee E P Bazan; Comprehensive mapping of rat corneal nerve architecture by immunofluorescence. Invest. Ophthalmol. Vis. Sci. 2020;61(7):386.
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© ARVO (1962-2015); The Authors (2016-present)
Rat corneal innervations have been studied extensively, but the detailed nerve architecture and neuropeptide distribution remain unclear. We here characterize the entire corneal nerve architecture, its neuropeptide distribution, and the changes with aging.
Sprague–Dawley rats (aged 1 day to 2 years old) of both sexes were sacrificed, and the whole corneas were immunostained with PGP9.5. Whole-mount images were acquired to build a complete map of corneal nerve architecture. To define the neuropeptide contents, the specimens were double-labeled with calcitonin gene-related peptide (CGRP), substance P (SP), neuropeptide Y (NPY), and vasoactive intestinal peptide (VIP) antibodies. Relative nerve fiber density for each neuropeptide fiber population was assessed based on whole mount images of the entire nerve architecture by computer-assisted image analysis.
There was an average of 12 ± 3 (n=10 corneas) thick nerve trunks which enter cornea, running towards the anterior stroma, subsequently dividing into smaller branches. The branches penetrate upwards into the epithelium to produce the subbasal nerve bundles which run centripetally and converge to form the vortex. Along the path, the subbasal bundles splint into numerous divisions to form the subbasal nerve plexus. Free endings budding from the plexus innervate the epithelial cells. Subbasal nerve density, calculated as the percentage of total area, was 21.5% ± 2.7% in the central area and 14.4% ± 1.8% in the peripheral area with a significant difference (P<0.001, n=6 rats). No significant difference was found between genders. Quantitative analysis of double-labeled corneal subbasal nerve fibers within the central zone revealed that CGRP, SP, VIP, and NPY took up 47.62% ± 3.51%, 34.95% ± 5.06%, 18.37% ± 5.75%, and 6.92% ± 2.66%, respectively. Development results showed that epithelial nerve formed a whirl-like structure as early as two weeks after birth. Epithelial nerve density gradually increased with age, reached full development at 5 weeks, and maintained the highest value to 36 weeks, then decreased, with the lowest density at 120 weeks.
This is the first study to provide a complete map of nerve architecture and neuropeptide contents of rat cornea. The normal innervation pattern described here will provide an essential baseline for investigators who use the rat model for assessing corneal pathologies involved nerve alterations.
This is a 2020 ARVO Annual Meeting abstract.
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