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Bernadette O. Ibitokun, Kenneth E. Miller; Excitatory Amino Acid Receptor Localization In Rat Cornea. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4194.
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© ARVO (1962-2015); The Authors (2016-present)
Normal corneal physiology depends in part on trophic effects of neurotransmitters such as glutamate, substance P and calcitonin gene related peptide released from corneal afferent nerves. In peripheral tissues, glutamate is transported and released as part of neurogenic inflammation from nerves, via vesicular glutamate transporter 1 or 2. Likewise, in the cornea, it is released during corneal nerve injury, inflammation and wound healing. From previous experiments in our lab, we identified glutamatergic nerve fibers in the cornea as well as retrogradely labeled VGLUT 2 positive neuronal cell bodies in the trigeminal ganglion. Indeed our data supports the hypothesis that glutamate is released into the cornea. If glutamate is released on corneal afferent nerves then we would expect that excitatory amino acid receptors will be present in the cornea. Our aim in this study, therefore, was the immunohistochemical localization of excitatory amino acid receptors in the rat cornea.
Adult Sprague Dawley rats were anesthetized and transcardially perfused with fixative, the cornea was excised and tissue was processed for immunohistochemistry using primary antisera: Rabbit anti pNR1 (1/500 dilution, Upstate) and Rabbit anti GluR2/3 (1/250 dilution, Chemicon). The secondary antisera was Alexafluor 488 conjugated goat anti-rabbit IgG (1/2000 dilution). Tissue was mounted using Prolong gold mounting medium and images were taken using a SPOT camera with an Olympus BX5 epifluorescence microscope.
The corneal afferent nerves were immunoreactive for excitatory amino acid receptors: NMDA R1 and AMPA GluR2/3 subunits. Scattered punctae were present within the sub epithelial nerve plexus and the stromal nerve bundles also had intense immunoreactivity for the NR1 subunit and AMPA GluR2/3 subunit. On whole mounts, the NR1 subunit had a rimmed distribution on the epithelial cells and immunoreactive punctae were observed within the epithelial cell nuclei.
Excitatory amino acid receptors, NMDA R1 and AMPA GluR2/3 subunits, were demonstrated in rat corneal afferents. These results further support the hypothesis that glutamate is released from corneal afferents and may play a role in corneal nociception. In other peripheral tissues, glutamate release from primary afferents causes a sensitizing effect on surrounding afferents and resident cells. The current results suggest that a similar effect may occur in the cornea via NMDA and AMPA receptors.
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