Heterogeneity of P-2u- and P-2y-purinergic receptor regulation of phospholipases in MDCK cells

We have characterized the signaling pathways of purinergic receptors present on the renal epithelial cell line, Madin-Darby canine kidney (MDCK, D-1 subclone). Several lines of evidence are consistent with the conclusion that coexisting P-2u and P-2y receptors release arachidonic acid and metabolites (AA) from MDCK-D-1 cells: 1) relative potencies of nucleotide analogues, 2) blockade of P-2y agonist- but not P-2u agonist-mediated release by suramin, and 3) additivity by 2-methylthio-ATP and UTP. Differences exist between the signaling pathways of the two receptors: pertussis toxin treatment partially inhibits P-2u- but not P-2y-mediated AA release, and P-2y (but not P-2u) receptors appear to stimulate D-myo-inositol 1,4,5-trisphosphate production. P-2u-receptor occupancy results in both homologous and heterologous desensitization; P-2y-receptor occupancy elicits only homologous desensitization. Both receptors stimulate phosphatidylcholine hydrolysis via phospholipase C activation. However, AA release appears to result from phospholipid deacylation by phospholipase A(2) activation, rather than from alternate pathways that may include PLC activation. These results demonstrate for the first time that two subtypes of P-2-purinergic receptors, P-2u and P-2y receptors, coexist on a single renal epithelium cell type and that these two receptor subtypes can promote AA release, probably via activation of PLA(2).