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S Narayan, G Prasanna, R Krishnamoorthy, C Hulet, T Yorio; Constitutive and Regulated Endothelin-1 (ET-1) Production in Human Retinal Pigment Epithelial Cells-Implication in Optic Neuropathies? . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4561.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Endothelin-1 (ET-1), a potent vasoactive peptide has been implicated in glaucoma-like optic neuropathy. Cholinergics have been shown to influence ET-1 production. The present study examines the role of the retinal pigment epithelium (ARPE-19) as a site for local ET-1 production and its regulation following cholinergic stimulation. Methods: Cultured ARPE-19 cells (differentiated and undifferentiated) were treated with carbachol (CCh), a cholinergic agonist, at different concentrations (1, 10, 100 µM) for 24 hours. Atropine (1 µM), pirenzepine (1 µM) and U73122 (2 µM) were used to delineate muscarnic receptor involvement. ET-1 secreted in the media was quantified by radioimmunoassay. Muscarinic receptor expression was determined by RT-PCR and immunoblotting. Intracellular calcium ([Ca2+]i) mobilization was determined following carbachol stimulation. Differentiated and undifferentiated ARPE-19 cells were characterized by the presence of tight junctions (ZO-1 expression) and melanin. Results: All five muscarinic receptor subtypes and preproET-1, endothelin converting enzyme-1 (ECE-1), ETA and ETB receptors were expressed at the transcript level in both forms of ARPE-19 cells. CCh 1 µM significantly enhanced the secretion of ET-1 in undifferentiated cells. Pirenzepine abolished CCh mediated rise in [Ca2+]i and subsequent ET-1 secretion, suggesting the predominance of the M1 mucarinic receptor subtype. Carbachol did not potentiate ET-1 release in differentiated ARPE-19 despite the expression of muscarinic receptors and robust CCh induced ([Ca2+]i) response. Conclusion: RPE can serve as a source for ET-1 and CCh may influence ET-1 release in these cells. Undifferentiated ARPE-19 cells appear to be more susceptible to CCh regulation than differentiated cells.
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