The P2Y 1 receptor is present in the heart, in skeletal and various smooth muscles, and in platelets, where its activation is linked to aggregation. Adenosine 3',5'- and 2',5'-bisphosphates have been identified as selective antagonists at the P2Y 1 receptor (Boyer et al. Mol. Pharmacol. 1996, 50, 1323-1329) and have been modified structurally to increase receptor affinity (Camaioni et al. J. Med. Chem. 1998, 41, 183-190). We have extended the structure-activity relationships to a new series of deoxyadenosine bisphosphates with substitutions in the adenine base, ribose moiety, and phosphate groups. The activity of each analogue at P2Y 1 receptors was determined by measuring its capacity to stimulate phospholipase C in turkey erythrocyte membranes (agonist effect) and to inhibit phospholipase C stimulation elicited by 10 nM 2-(methylthio)adenosine 5'-diphosphate (antagonist effect). 2'-Deoxyadenosine bisphosphate analogues containing halo, amino, and thioether groups at the 2-position of the adenine ring were more potent P2Y 1 receptor antagonists than analogues containing various heteroatom substitutions at the 8-position. An N 6 -methyl-2-chloro analogue, 6, was a full antagonist and displayed an IC 50 of 206 nM. Similarly, N 6 - methyl-2-alkylthio derivatives 10, 14, and 15 were nearly full antagonists of IC 50 < 0.5 μM. On the ribose moiety, 2'-hydroxy, 4'-thio, carbocyclic, and six-membered anhydrohexitol ring modifications have been prepared and resulted in enhanced agonist properties. The 1,5-anhydrohexitol analogue 36 was a pure agonist with an EC 50 of 3 μM, i.e., similar in potency to ATP. 5'-Phosphate groups have been modified in the form of triphosphate, methyl phosphate, and cyclic 3',5'-diphosphate derivatives. The carbocyclic analogue had enhanced agonist efficacy, and the 5'-O-phosphonylmethyl modification was tolerated, suggesting that deviations from the nucleotide structure may result in improved utility as pharmacological probes. The N 6 -methoxy modification eliminated receptor affinity. Pyrimidine nucleoside 3',5'- bisphosphate derivatives were inactive as agonists or antagonists at P2Y receptor subtypes.