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Joanna Vrouvlianis, Robin Hodges, Rachel Scott, Darlene Dartt; Identification of P2X Purinergic Receptors Activated by ATP in Rat Lacrimal Gland. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3705.
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To identify the type of purinergic receptors activated by ATP in rat lacrimal gland and determine their function.
Purinergic receptors were identified by RT-PCR, Western blot analysis, and immunofluorescence techniques. Acini from rat lacrimal gland were isolated by collagenase digestion. Acini were incubated with the fluorescent indicator fura2, and intracellular [Ca2+] ([Ca2+]i) was determined. Protein secretion was measured by fluorescent assay.
We previously showed that P2X7 receptors are present and functional in the lacrimal gland. In this study, we detected P2X1-4, and P2X6 receptors in the lacrimal by RT-PCR, western blot, and immunofluorescence analysis. P2X5 receptors were not detected. ATP increased [Ca2+]i and protein secretion in a concentration-dependent manner. Repeated applications of ATP decreased the [Ca2+]i. Incubation with the P2X1 receptor inhibitor NF023 did not alter the ATP-stimulated [Ca2+]i. Incubation with zinc that potentiates P2X2 and P2X4 receptor responses or lowering the pH from 7.4 to 6.8 that potentiates P2X2 receptor responses also did not alter the ATP-stimulated [Ca2+]i. P2X3 receptor inhibitors A-317491 and TNP-ATP, significantly decreased ATP-stimulated [Ca2+]i and protein secretion while the P2X3 receptor agonist α,β methylene ATP significantly increased [Ca2+]i and protein secretion. The P2X7 receptor inhibitor A438079 had no effect on ATP-stimulated [Ca2+]i at 10-6 M, but did at 10-4 M.
Purinergic receptors P2X1-4, and P2X6 are present in the lacrimal gland. ATP utilizes P2X3 and P2X7 receptors to stimulate an increase in [Ca2+]i and protein secretion.
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