Regulation of transepithelial ion transport by two different purinoceptors in the apical membrane of canine kidney (MDCK) cells

O. Zegarra-Moran, G. Romeo, L. J V Galietta

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The effect of extracellular nucleotides on the transepithelial ion transport of Madin Darby canine kidney cells (MDCK) was investigated. Cells were grown up to confluency on permeable supports and the short circuit current (I(sc)) was measured with an Ussing chamber-like mini-perfusion system. Apical ATP stimulated a biphasic I(sc) increase consisting of a first rapid and transient peak followed by a broader one. The first peak evoked by ATP was reversibly blocked by basilen blue (BB) in a concentration-dependent fashion, with an EC50 of 7.5 μM. The P(2Y) receptor agonist, 2-methylthioATP (2-MeSATP) caused a single transient I(sc) increase that was completely blocked by pretreatment with BB. On the contrary, the P(2x) agonist, α,β-methylene ATP (α,β-meATP) was almost completely ineffective on I(sc). UTP essentially induced a monaphasic response the time-course of which resembled that of the second peak stimulated by ATP. The agonist potency order was 2-MeSATP ≥ ATP >> UTP, α-β-meATP for the first peak and UTP ≥ ATP > 2-MeSATP > α,β-meATP for the second peak. Monolayer incubation with the membrane permeable calcium chelator [bis-o-aminophenoxy)-ethane-N,N,N',N',-tetraacetic acid, tetra(acetoximethyl)-ester] (BAPTA/AM) inhibited the ATP-evoked first peak. The non-hydrolyzable ATP analogue, adenosine-5'-O-(3-thio)-trisphosphate (ATP-γ-S) elicited a biphasic response similar to that of ATP. The P1 receptor agonist, 2-chloroadenosine and CGS-21680, were almost unable to induce an I(sc) increase. These results rule out the involvement of ATP hydrolysis and P1 receptor activation as responsible for I(sc) increase. Inhibition of prostaglandins synthesis by indomethacin abolished the second ATP-evoked peak. Chloride replacement with gluconate on both sides of the epithelium completely inhibited the second peak induced by ATP but only reduced the amplitude of the first spike. The results suggest that ATP stimulates I(sc) increase by two mechanisms. The first one is mediated by a P(2Y) receptor and by intracellular calcium increase. The second induces prostaglandin synthesis probably through a P(ZU) receptor activation.

Original languageEnglish
Pages (from-to)1052-1056
Number of pages5
JournalBritish Journal of Pharmacology
Issue number5
Publication statusPublished - 1995


  • ATP
  • MDCK cells
  • P purinoceptors
  • Short circuit current
  • Transepithelial ion transport
  • UTP

ASJC Scopus subject areas

  • Pharmacology


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