Hansmann, G; Bultmann, R; Tuluc, F; Starke, K
The receptors through which 2-methylthio ATP (MeSATP),
adenosine 5'-O-(2-thiodiphosphate) (ADP beta S), UTP and ATP elicit endothelium-dependent
relaxation of noradrenaline-precontracted rings of the rat aorta were characterized by means of a
series of antagonists. The acetylcholine-induced relaxation and the degradation of MeSATP, UTP
and ATP were also studied. The potency of the nucleotides at producing relaxation decreased in
the order MeSATP (EC50 0.24 mu M) > ADP beta S (0.43 mu M) > UTP (1.09 mu M) > ATP (3.53 mu M).
MeSATP, ADP beta S and UTP did not cause relaxation when the endothelium had been destroyed; high
concentrations of ATP still caused some relaxation. The relaxation by MeSATP, ADP beta S and UTP
became very small after treatment of the rings with N-G-nitro-L-arginine methyl ester; the
relaxation by ATP was less affected. Pre-exposure to MeSATP (100 mu M) abolished or almost
abolished the relaxation normally elicited by MeSATP and ADP beta S, did not change that elicited
by UTP and slightly enhanced the relaxation elicited by ATP Of nine compounds examined as
antagonists, six attenuated selectively the effect of some or all of the nucleotides (as compared
to acetylcholine): suramin, reactive blue 2,pyridoxalphosphate-6-azophenyl-2',5'-dissulphonate
(iso -PPAD S), pyridoxalphosphate-6-azophenyl-2',4'-disulphonate (PPADS), reactive red 2 and
5,5'-(1,1'-biphenyl-4,4'-diylbisazo)-bis-7-amino-6-hydroxy- naphthalene-1,4-disulphonate
(NH05). Decreases of maximal relaxations and slopes different from unity in Schild plots often
indicated non-competitive kinetics of the antagonism. For each of the six 'selective'
antagonist, the apparent Kd values against MeSATP and against ADP beta S were similar: none of
the six differentiated between MeSATP and ADP beta S. Also, for each of four 'selective'
antagonists, the apparent K-d values against UTP and against ATP were similar: none of the four
differentiated between these two nucleotides (two antagonists did not act against UTP and ATP in
the 'selective' concentration range). On the other hand, for five of the six 'selective'
antagonists (the exception being NH05), the apparent K-d values against MeSATP and ADP beta S
were considerably lower than those against UTP and ATP. At the highest concentrations 'tested
against agonist-evoked relaxations, the antagonists did not alter the removal from the incubation
medium, by pieces of rat aorta, of MeSATP, UTP and ATP. It is concluded that nucleotides cause
endothelium-dependent relaxation of the rat aorta through two sites: a P2Y-receptor and a P2U-
receptor. The receptors may be pharmacologically similar to a bovine endothelial P2Y (P2Y(1)) and
a cloned rat P2U (P2Y(2)) receptor, respectively. ATP acts mainly through the P2U-receptor.
Suramin, reactive blue 2, iso-PPADS, PPADS and reactive red 2 are more potent at the P2Y- than
the P2U-receptor. NH05 does not discriminate between the two receptors but is the most potent P2U
antagonist so far described.