Evidence for the existence of two different ADP-binding sites on rat platelets.
[3H]-2-Methylthio-ADP ([3H]-2-MeS-ADP), a stable analogue of ADP bound to one type of specific binding sites on rat platelets (KD = 0.77 +/- 0.07 nM, Bmax = 160 +/- 11 fmol/10(8) cells). 2-MeS-ADP and ADP antagonized [3H]-2-MeS-ADP binding, showing respective Ki values of 1.4 +/- 0.1 nM and 486 +/- 78 nM. Clopidogrel, a potent and specific inhibitor of ADP-induced platelet aggregation partially inhibited (approximately 70% inhibition) the binding of [3H]-2-MeS-ADP at the same time it abrogated 2-MeS-ADP- and ADP-induced adenylyl cyclase inhibition and aggregation. A population of clopidogrel-resistant [3H]-2-MeS-ADP binding sites was detected on platelets from treated animals. These receptor sites (KD = 0.9 +/- 0.2 nM, Bmax = 47 +/- 5 fmol/10(8) platelets) which showed high affinity for both ADP and 2-MeS-ADP (Ki values in the nanomolar range) might be involved in the ADP-induced shape change, a clopidogrel-resistant ADP-induced event. Using clopidogrel which acts via a direct and irreversible inhibition of ADP binding to its adenylyl cyclase-coupled receptor sites on platelets, we were able to discriminate between two types of ADP receptor sites. The former which was clopidogrel-sensitive represented about 70% of the total [3H]-2-MeS-ADP receptors and was responsible for ADP-induced platelet aggregation and adenylyl cyclase inhibition. The latter which was not affected by clopidogrel might be involved in ADP-induced shape-change.
Savi
P
P
Sanofi Recherche, Hemobiology Research Department, Toulouse, France.
Laplace
M C
MC
Herbert
J M
JM
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism* / Q000648:ultrastructure; D000077144:Clopidogrel; D005260:Female; D010974:Platelet Aggregation; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000378:metabolism
D000244:Adenosine Diphosphate; D000077144:Clopidogrel; D000262:Adenylyl Cyclases; D013988:Ticlopidine