ATP reduces macromolecule permeability of endothelial monolayers despite increasing [Ca2+]i.
We investigated the relationship between the ATP-evoked rise of cytosolic Ca2+ concentration ([Ca2+]i) and barrier function in porcine aortic endothelial monolayers. ATP (0.01-100 microM) induced a transient rise of [Ca2+]i and reduced permeability in a concentration-dependent manner. In contrast, the Ca2+ ionophore ionomycin (1 microM) elicited a rise in [Ca2+]i comparable to that induced by ATP (10 microM), but it increased permeability. For the reduction of permeability, nucleotides were found to be in the following order of potency: ATP = ATPgammaS > ADP = UTP. Blockade of adenosine receptors by 8-phenyltheophylline (10 microM) did not affect ATP (10 microM)-induced reduction of permeability. ATP reduced permeability even in endothelial monolayers that had been loaded with the Ca2+ chelator BAPTA to prevent the rise in [Ca2+]i. U-73122 (1 microM), an inhibitor of phospholipase C (PLC), completely abolished the effect of ATP (10 microM) on permeability. It also abolished the translocation of protein kinase C (PKC) in response to ATP, which could also be achieved by the PKC inhibitors Gö-6976 (100 nM) or bisindolylmaleimide I (1 microM). In the presence of PKC inhibitors, however, the permeability effect of ATP was not affected. The presence of inhibitors of adenylate or guanylate cyclase (50 microM SQ-22536 or 20 microM ODQ) prevented changes in cyclic nucleotides but did not affect the permeability effects of ATP. The study shows that ATP reduces macromolecule permeability via a PLC-mediated mechanism that is independent of the concomitant effects of ATP on cytosolic Ca2+, cyclic nucleotides, or PKC.
Noll
T
T
Physiologisches Institut und, Abteilung für Kardiologie, Justus-Liebig-Universität, D-35392 Giessen, Germany. thomas.noll@physiologie.med.uni-giessen.de
Hölschermann
H
H
Koprek
K
K
Gündüz
D
D
Haberbosch
W
W
Tillmanns
H
H
Piper
H M
HM
D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002199:Capillary Permeability / Q000187:drug effects / Q000502:physiology*; D002417:Cattle; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D046911:Macromolecular Substances; D009712:Nucleotides, Cyclic / Q000378:metabolism; D009994:Osmolar Concentration; D011493:Protein Kinase C / Q000378:metabolism; D013552:Swine; D010738:Type C Phospholipases / Q000378:metabolism
D046911:Macromolecular Substances; D009712:Nucleotides, Cyclic; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D010738:Type C Phospholipases; D002118:Calcium