DEPENDENCE OF P2-NUCLEOTIDE RECEPTOR AGONIST-MEDIATED ENDOTHELIUM-INDEPENDENT RELAXATION ON ECTONUCLEOTIDASE ACTIVITY AND A(2A)-RECEPTORS IN RAT PORTAL-VEIN

1 The mechanism of action of P2 nucleotide receptor agonists that prod uce endothelium-independent relaxation and the influence of ecto-ATPas e activity on this relaxing effect have been investigated in rat porta l vein smooth muscle. 2 At 25 degrees C, ATP, 2-methylthioATP (2-MeSAT P) and 2-chloroATP (2-ClATP), dose-dependently inhibited spontaneous c ontractile activity of endothelium-denuded muscular strips from rat po rtal vein. The rank order of agonist potency defined from the half-inh ibitory concentrations was 2-ClATP (2.7+/-0.5 mu M, n=7)>ATP (12.9+/-1 .1 mu M, n=9)greater than or equal to 2-MeSATP (21.9+/-4.8 mu M, n=4). In the presence of alpha beta-methylene ATP (alpha beta-MeATP, 200 mu M) which itself produced a transient contractile effect, the relaxing action of ATP and 2-MeSATP was completely abolished and that of 2-ClA TP strongly inhibited. 3 The non-selective P2-receptor antagonist pyri doxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS, 100 mu M) di d not affect the relaxation induced by ATP, 2-MeSATP, and 2-ClATP. 4 T he A(2A)-adenosine receptor antagonist ZM 241385 inhibited the ATP-ind uced relaxation in a concentration-dependent manner (1-100 nM). In the presence of 100 nM ZM 241385, the relaxing effects of 2-MeSATP and 2- ClATP were also inhibited. 5 ADP, AMP and adenosine also produced conc entration-dependent inhibition of spontaneous contractions. The relaxi ng effects of AMP and adenosine were insensitive to alpha beta-MeATP ( 200 mu M) but were inhibited by ZM 241385 (100 nM).6 Simultaneous meas urements of contraction and ecto-ATPase activity estimated by the degr adation of [gamma-P-32]-ATP showed that muscular strips rapidly (10-60 s) hydrolyzed ATP. This ecto-ATPase activity was abolished in the pre sence of EDTA and was inhibited by 57+/-11% (n=3) by 200 mu M alpha be ta-MeATP. 7 These results suggest that ATP and other P2-receptor agoni sts are relaxant in rat portal vein smooth muscle, because ectonucleot idase activity leads to the formation of adenosine which activates A(2 A)-receptors.