Blackwell Publishing Journal Backfiles 1879-2005
Abstract: Adenosine transport in cultured chromaffin cells was inhibited by purinergic P2y-receptor agonists without significant changes in the affinity constant, the values being between 1 ± 0.4 and 1.6 ± 0.6 μM. The Vmax parameter was modified significantly, being 40 ± 1.0, 26 ± 5.0, 32 ± 3.0, and 22 ± 4.7 pmol/106 cells/min for control, adenosine-5′-O-(2-thiodiphosphate), 5′-adenylylimidodiphosphate, and P1,P4-di(adenosine-5′-) tetraphosphate (Ap4A) (100 μM for every effector), respectively. Ap4A, a physiological ligand for P2y receptors in chromaffin cells, showed the highest inhibitory effect (45%). This transport inhibition is explained by an increase in the cytosolic Ca2+ concentration ([Ca2+]i) and the activation of protein kinase C (PKC). Experiments of [Ca2+]i measurement with the fura-2 technique showed that P2y agonists, as well as bradykinin, were able to increase [Ca2+]i, this effect being independent of the presence of extracellular Ca2+. The peptide bradykinin, determined to be coupled to phosphatidylinositol hydrolysis and internal Ca2+ mobilization in chromaffin cells, exhibited a behavior similar to that of P2y agonists in adenosine transport inhibition (39%). P2y agonists and bradykinin increased PKC activity associated with the membrane fraction (about 50% increase in particulate PKC activity with respect to controls). The present studies suggest that adenosine transport is regulated by P2y-purinergic receptors mediated via Ca2+ mobilization and PKC activation.
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