Nucleotide‐evoked calcium signals and anion secretion in equine cultured epithelia that express apical P2Y₂ receptors and pyrimidine nucleotide receptors

Abstract

1 Experiments with a spontaneously transformed equine epithelial cell line showed that certain nucleotides increased intracellular free calcium ([Ca 2+ ] i ) in cells plated on glass coverslips. The rank order of potency was ATP=UTP45-Br-UTP, whilst UDP and ADP were ine ective. The response thus appears to be mediated by P2Y 2 receptors. 2 Nucleotides also increased short circuit current (I SC ) in cells grown into epithelial monolayers and the rank order of potency was UDP4UTP45-Br-UTP4ATP4ADP. The increase in [Ca 2+ ] i and the rise in I SC thus have di erent pharmacological properties. Cross-desensitization experiments indicated that, as well as P2Y 2 receptors, the monolayer cultures express at least one additional receptor population that allowed nucleotides to increase I SC .

Stimulation of Cl− Secretion via Membrane-restricted Ca2+ Signaling Mediated by P2Y Receptors in Polarized Epithelia

“…Previous studies have identified the functional expression of apical purinoceptors (P2Y2) and pyrimidinoceoptors (P2Y4 and/or P2Y6) when the cells are grown on permeable supports. P2X receptors are not involved in mediating the nucleotide responses (16). This cell line also represents the simplest calcium-dependent secretory system in which cAMP-dependent Cl Ϫ secretion (i.e.…”

“…cystic fibrosis transmembrane conductance regulator function) is absent (15). Transepithelial secretion can only be activated by calciummobilizing agonists, such as extracellular ATP, UTP, UDP, and bradykinin (16).…”

“…Cells reach confluence after 4 days with a resistance greater than 150 ohm cm 2 . Simultaneous Measurements of [Ca 2ϩ ] i and I SC -Nucleotide-evoked calcium signals and anion secretion were measured simultaneously as described previously (6,13,14,16). Cells grown on Transwell-col membrane were loaded with Fura-2 by incubation (45 min at 37°C) in bicarbonate-buffered saline containing 3 M Fura-2-acetoxymethylester and 1.6 M pluronic F127.…”

“…The Ca 2ϩ -mediated Cl Ϫ secretion activated by various nucleotides was explored in an equine sweat gland epithelial cell line with emphasis on the polarized Ca 2ϩ mobilization and influx in response to apical versus basolateral P2Y receptor stimulation. Previous studies have shown that the cultured cells will form polarized monolayer on permeable support (15) and express both apical purine and pyrimidine receptors (7,16) that are linked to internal Ca 2ϩ release and capacitative calcium entry (17).…”

Identification of Calcium‐Activated Potassium Channels in Cultured Equine Sweat Gland Epithelial Cells

“…ATP-induced activation of KCa channels on a cell-attached configuration was seen as an increase in Popen, but this response only lasted for around 2·5 min. Similarly, ATP transiently increased the Ca¥-dependent Cl¦ secretion in monolayers of cultured equine sweat gland epithelial cells (Ko et al 1997;Wilson et al 1998). These results suggest that the ATP-induced increase in [Ca¥]é may serve as a common promoter for Cl¦ and K¤ channels in the process of transepithelial ion transport.…”