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purpose. To study the presence of muscarinic and α-and β-adrenergic receptors in a normal human conjunctival epithelial (IOBA-NHC) cell line.

methods. Neurotransmitter receptors were determined in IOBA-NHC cells by flow cytometry, immunofluorescence, and Western blot analysis. Antibodies to M 1-, M 2-, and M 3-muscarinic and to α 1A-, α 1B-, α 1D-, α 2A-, α 2B-, α 2C-, β 1-, β 2-, and β 3-adrenergic receptor subtypes were used. Different culture media were tested, including the addition of tumor necrosis factor (TNF)-α and/or interferon (IFN)-γ. Normal human conjunctiva biopsy specimens and rat tissues were used in control experiments.

results. By immunofluorescence microscopy, all receptor subtypes, except the α 2C-adrenergic receptor, were detected in control biopsy specimens. By flow cytometry, the M 2-and M 3-muscarinic receptors and α 1A-, α 1B-, α 1D-, α 2A-, α 2B-, α 2C-, β 1-, and β 3-adrenergic receptors were detected intracellularly and in cell membranes of the IOBA-NHC cells. M 1-muscarinic and β 2-adrenergic receptors were detected only intracellularly, but were mobilized to the cell membrane when cholera toxin and hydrocortisone were omitted from the culture medium. Confocal microscopy detected the M 2 and M 3-muscarinic and α 1A-, α 2A-, α 2B-, β 1-and β 2-adrenergic receptor subtypes. Western blot analyses showed bands for all receptors. M 2-muscarinic and α 1B-and α 2B-adrenergic receptors expression was upregulated when cells were treated with the proinflammatory cytokines IFNγ and/or TNFα.

conclusions. The IOBA-NHC cell line maintained expression of the neurotransmitter receptors expressed in …