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Stimulation of chloride secretion by P1 purinoceptor agonists in cystic fibrosis phenotype airway epithelial cell line CFPEo-.

Abstract
1. P1 purinoceptor agonists like adenosine have been shown to stimulate Cl- transport in secretory epithelia. In the present study, we investigated whether P1 agonist-induced Cl- secretion is preserved in cystic fibrosis airway epithelium and which signalling mechanism is involved. The effects of purinoceptor agonists on Cl- secretion were examined in a transformed cystic fibrosis airway phenotype epithelial cell line, CFPEo-. 2. Addition of adenosine (ADO; 0.1-1 mM) markedly increased 125I efflux rate. The rank order of potency of purinoceptor agonists in stimulating 125I efflux was ADO > AMP > ADP approximately equal to ATP. A similar order of potency was seen in transformed cystic fibrosis nasal polyp cells, CFNPEo- (ADO > ATP > AMP > ADP). These results are consistent with the activation of Cl- secretion via a P1 purinoceptor. 3. The P1 agonists tested (at 0.01 and 0.1 mM) revealed a rank order of potency of 5'-N-ethylcarboxamine adenosine (NECA) > 2-chloro-adenosine (2-Cl-ADO) > R-phenylisopropyl adenosine (R-PIA). 4. The known potent A2 adenosine receptor (A2AR) agonist, 5'-(N-cyclopropyl) carboxamidoadenosine (CPCA, 2 microM) but not the A1 adenosine receptor agonist, N6-phenyl adenosine (N6-phenyl ADO, 10 microM) markedly increased 125I efflux rate (baseline, 5.9 +/- 2.0% min-1, + CPCA, 10.9 +/- 0.6% min-1; P < 0.01). The stimulant effect of CPCA (10 microM) was abolished by addition of the A2AR antagonist 3,7-dimethyl-1-propargylxanthine (DMPX) (100 microM; reported K(i) = 11 +/- 3 microM). These results favour the involvement of A2AR. 5. ADO (0.1-mM) and CPCA (2 microM) both induced a marked increase in intracellular [Ca2+] ([Ca2+]i); the effect of the latter was again abolished by pretreatment of the cells with DMPX. By contrast, N6-phenyl ADO did not affect [Ca2+]i. 6. In patch-clamp experiments, ADO (1 mM) induced an outwardly-rectified whole-cell Cl- current (baseline, 2.5 +/- 0.8 pA pF-1, + ADO, 78.4 +/- 23.8 pA pF-1; P < 0.02), which was largely inhibited in cells internally perfused with a selective inhibitory peptide of the multifunctional Ca2+/calmodulin-dependent protein kinase, CaMK [273-302] (20 microM), as compared to a control peptide, CaMK [284-302]. Addition of BAPTA (10 mM), a Ca2+ chelator, to the perfusion pipette also abolished the ADO-elicited Cl- current. 7. In conclusion, our results suggest that A2AR participates in regulation of airway C1 secretion via aCa2+-dependent signalling pathway, which involves CaMK and appears to be at least partially conserved in cystic fibrosis airway epithelial cells.
AuthorsA C Chao, J B Zifferblatt, J A Wagner, Y J Dong, D C Gruenert, P Gardner
JournalBritish journal of pharmacology (Br J Pharmacol) Vol. 112 Issue 1 Pg. 169-75 (May 1994) ISSN: 0007-1188 [Print] England
PMID8032638 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)
Chemical References
  • Adenine Nucleotides
  • Chloride Channels
  • Chlorides
  • Iodine Radioisotopes
  • Purinergic P1 Receptor Antagonists
  • Receptors, Purinergic P1
  • N-cyclopropyl adenosine-5'-carboxamide
  • 3,7-dimethyl-1-propargylxanthine
  • Iodine
  • Adenosine
  • Theobromine
  • Calcium
Topics
  • Adenine Nucleotides (pharmacology)
  • Adenosine (analogs & derivatives, pharmacology)
  • Calcium (metabolism)
  • Cell Line
  • Chloride Channels (drug effects, metabolism)
  • Chlorides (metabolism)
  • Cystic Fibrosis (metabolism)
  • Electrophysiology
  • Epithelium (drug effects, metabolism)
  • Humans
  • Iodine (metabolism)
  • Iodine Radioisotopes
  • Phenotype
  • Purinergic P1 Receptor Antagonists
  • Receptors, Purinergic P1 (drug effects, metabolism)
  • Signal Transduction (drug effects)
  • Spectrometry, Fluorescence
  • Theobromine (analogs & derivatives, antagonists & inhibitors, pharmacology)

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