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Mortimer Lorber, Eva C. Permaul, Supti Sen, Deborah L. Berry; The Rat Harderian Gland Contains Immunoreactive Cholecystokinin. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3734.
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© ARVO (1962-2015); The Authors (2016-present)
Because the human exorbital lacrimal gland contains immuoreactive cholecystokinin (CCK-8) (Abstract # 4165 ARVO 2010), might the rat Harderian gland, a major lacrimal organ in non-primate terrestrial vertebrates, contain enteric peptides, specifically CCK?
Four Harderian glands were excised from four adult female rats, formalin-fixed and embedded. Immunohistochemical staining was by a rabbit polyclonal antibody marketed as Anti-Cholecystokinin (26-33) (sulfated CCK-8) by Sigma-Aldrich. It is essentially specific, having cross-reactivities with human Gastrin I of 0.2% and with Big Gastrin of 1%. It cross-reacts with rat tissues. Five micron sections were deparaffinized. Following epitope retrieval (98oC in pH 6 citrate buffer), endogenous peroxidase was blocked with 3% peroxide. Sections were exposed to 10% goat serum, the primary anti-CCK diluted 1:1400 for 1h at room temp, a biotin-conjugated secondary anti-rabbit antibody (Vector Labs), Vectastain ABC reagent, DAB chromogen (Dako), and hematoxylin counterstaining. Positive controls were rat antrum and duodenum. Negative ones omitted the primary antibody.
Great acinar diversity existed. Many acinar cell nuclei immunostained strongly with faint to impressive cytoplasmic granulation, but even within the same acinus other cells were negative. Golden cytoplasmic granules varied from absent, to faintly occupying the apical part of the cell, to very often filling the entire cytoplasm of large groups of acini. Myoepithelial nuclei immunostained as did some cytoplasmic granules. In tubules only a few nuclei immunostained but their lumens were often full of golden granules. In the negative controls porphyrin bodies were unchanged. All other gland components did not immunostain.
CCK-8 in Harderian acini and tubular lumens enters tears. After traversing the nasolacrimal ducts tears enter the nasopharynx and are swallowed. CCK-8’s entry into the alimentary tract is a scant supplement to the large amounts made in the antrum and small intestine. Porphyrin bodies are unassociated with this enteric peptide.
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