UTP-preferring P2 receptor mediates inhibition of sodium transport in porcine thyroid epithelial cells

1 The effects of adenosine 5'-triphosphate (ATP). uridine 5'-triphosphate (UTP) and analogues on forskolin-stimulated absorption of Na+ by porcine thyroid epithelial cells were analysed in cultures grown as confluent monolayers on permeable supports in Transwell Ussing chambers. 2 85% of the forskolin (10 mu M)-stimulated short-circuit current was inhibited by phenamil (1 mu M), which is a selective antagonist for epithelial type Na+ channels. 3 Phenamil-sensitive current was inhibited in a dose dependent manner by nucleotides added to the apical compartment of Ussing chambers. In contrast, the phenamil-resistant current, previously shown to represent anion secretion, was unaffected by nucleotides. 4 The order of potency (with EC50 values given in mu M) was UTP (0.08) > > ATP (6.3) = uridine 5'-diphosphate (UDP) (6.6) > 2methyl-thio-adenosine-5'-triphosphate (2MeSATP) (84.5) > adenosine 5'-diphosphate (ADP) (147.8) > alpha,beta-methylene ATP (> 150) > >adenosine (> 1000). 5 P-2 receptors mediating inhibition of sodium absorption were present on the apical membrane of the cells since addition of UTP (1 - 1000 mu M) to the basal compartment of the Ussing chambers had little effect while subsequent addition to the apical compartment produced a normal response. 6 Cibachron blue (Reactive blue 2) (1-100 mu M), an antagonist at some P2 receptor subtypes, inhibited phenamil sensitive current in a dose dependent manner with half maximal inhibition occurring at 14.25 mu M. 7 Suramin (100 mu M), pyridoxalphosphate-6-azophenyl-2'.4'-disulphonic acid (PPADS) (100 mu M) and pyridoxal 5'-phosphate (P5P) (100 mu M) showed only slight competitive antagonism against the response to UTP. 8 These results indicate that a UTP-preferring P-2 receptor located on the apical membrane of thyroid epithelial cells mediates inhibition of Na+ absorption.