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Kenneth E. Miller, Bernadette O. Ibitokun; Localization Of Glutamate, Glutaminase, And Aspartate Aminotransferase In Rat Corneal Afferents. Invest. Ophthalmol. Vis. Sci. 2011;52(14):315.
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The mammalian cornea is densely innervated and contains neurotransmitters, such as substance P (SP) and calcitonin gene related peptide (CGRP), that are released peripherally in the cornea. Since all primary afferent neurons are glutamatergic (Miller et al., Pharmacology and Therapeutics, 2011), it was of interest to determine if afferents in the cornea could be identified with glutamatergic markers. A previous study from our laboratory identified vesicular glutamate transporter 2 in corneal afferents. The aim of this study, therefore, was the further exploration of glutamatergic markers in corneal afferents using immunohistochemistry.
Adult Sprague Dawley rats were transcardially perfused with fixative. The cornea was excised and whole mount tissue was processed for immunohistochemistry using primary antisera: rabbit anti glutaminase (GLS; 1:10,000; Curthoys, Colo. St.), mouse anti-glutamate (1:20,000; Madl, Colo. St.), sheep anti-aspartate aminotransferase (AATase; 1:1,000; Rockland). The secondary antisera were Alexafluor 488 conjugated anti-IgG’s (1/2000 dilution). Tissue was mounted with Prolong Gold mounting medium and images obtained with a SPOT camera attached to an Olympus BX5 epifluorescence microscope.
Stromal nerve bundles were observed with GLS, glutamate, and AATase immunoreactivity. Immunoreactive varicose fibers for GLS, glutamate, and AATase were observed in the subepithelial neve plexus and among corneal epithelial cells.
The current results confirm the presence of glutamatergic primary afferent nerve fibers in the rat cornea. 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 and a similar mechanism may occur in the cornea.
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