P2Y11 receptors activate adenylyl cyclase and contribute to nucleotide-promoted cAMP formation in MDCK-D1 cells -: A mechanism for nucleotide-mediated autocrine-paracrine regulation

Extracellular nucleotides activate P2Y receptors, thereby increasing cAMP formation in Madin-Darby canine kidney (MDCK-D-1) cells, which express P2Y(1), P2Y(2), and P2Y(11) receptors (Post, S. R., Rump, L. C., Zambon, A., Hughes, R. J., Buda, M. D., Jacobson, J. P., Kao, C. C., and Insel, P. A. (1998) J. Biol. Chem. 273, 23093-23097). The cyclooxygenase inhibitor indomethacin (indo) eliminates UTP-promoted cAMP formation (i.e. via P2Y(2) receptors) but only partially blocks ATP-promoted cAMP formation. The latter response is completely blocked by the nonselective P2Y receptor antagonist suramin. We have sought to identify the mechanism for this P2Y receptor-mediated, indo-resistant cAMP formation. The agonist rank order potencies for cAMP formation were: ADPbetaS greater than or equal to MT-ADP > 2-MT-ATP > ADP, ATP, ATPgammaS > UTP, AMP, adenosine. We found a similar rank order in MDCK-D-1 cells overexpressing cloned green fluorescent protein-tagged P2Y(11) receptors, but the potency of the agonists was enhanced, consistent with a P2Y(11) receptor-mediated effect. cAMP generation by the P2Y(1) and P2Y(11) receptor agonist ADPbetaS was not inhibited by several P2Y(1)-selective antagonists (PPADS, A2P5P, and MRS 2179). Forskolin synergistically enhanced cAMP generation in response to ADPbetaS or PGE(2), implying that, like PGE(2), ADPbetaS activates adenylyl cyclase via G(8), a conclusion supported by results showing ADPbetaS and MT-ADP promoted activation of adenylyl cyclase activity in MDCK-D-1 membranes. We conclude that nucleotide-promoted, indo-resistant cAMP formation in MDCK-D-1 cells occurs via G(8)-linked P2Y(11) receptors. These data describing adenylyl cyclase activity via endogenous P2Y(11) receptors define a mechanism by which released nucleotides can increase cAMP in MDCK-D-1 and other P2Y(11)-containing cells.