P2Y6 receptor mediates colonic NaCl secretion via differential activation of cAMP-mediated transport

Abstract

Extracellular nucleotides are important regulators of epithelial ion transport. Here we investigated nucleotide-mediated effects on colonic NaCl secretion and the signal transduction mechanisms involved. Basolateral UDP induced a sustained activation of Cl -secretion, which was completely inhibited by 293B, a specific inhibitor of cAMP-stimulated basolateral KCNQ1/KCNE3 K + channels. We therefore speculated that a basolateral P2Y 6 receptor could increase cAMP. Indeed UDP elevated cAMP in isolated crypts. We identified an epithelial P2Y 6 receptor using crypt [Ca 2+ ] i measurements, RT-PCR, and immunohistochemistry. To investigate whether the rat P2Y 6 elevates cAMP, we coexpressed the P2Y 1 or P2Y 6 receptor together with the cAMP-regulated cystic fibrosis transmembrane conductance regulator (CFTR) Cl -channel in Xenopus oocytes. A two-electrode voltage clamp was used to monitor nucleotide-induced Cl -currents. In oocytes expressing the P2Y 1 receptor, ATP transiently activated the endogenous Ca 2+ -activated Cl -current, but not CFTR. In contrast, in oocytes expressing the P2Y 6 receptor, UDP transiently activated the Ca 2+ -activated Cl -current and subsequently CFTR. CFTR Cl -currents were identified by their halide conductance sequence. In summary we find a basolateral P2Y 6 receptor in colonic epithelial cells stimulating sustained NaCl secretion by way of a synergistic increase of [Ca 2+ ] i and cAMP. In support of these data P2Y 6 receptor stimulation differentially activates CFTR in Xenopus oocytes.