1. By use of a number of analogues of adenine nucleotides, the structure-activity relationships of the human platelet receptor for adenosine 5'-diphosphate (ADP) mediating increases in intracellular calcium were investigated, and compared with the known structure-activity relationships for induction by ADP of platelet aggregation. 2. ADP, 2-methylthioadenosine 5'-diphosphate (2-methylthio-ADP), adenosine 5'-O-(1-thiodiphosphate) (ADP-alpha-S) and adenosine 5'-O-(2-thiodiphosphate) (ADP-beta-S) each induced increases in intracellular calcium in a manner similar to their reported ability to induce human platelet aggregation. The effects of these agonists were antagonized by ATP, with a pA2 value in each case consistent with the inhibition by ATP of ADP-induced aggregation. In the case of ADP, the inhibition by ATP of increases in intracellular calcium was shown to be competitive by Schild analysis. 3. Of the analogues tested as inhibitors of the effect of ADP on intracellular calcium, 2-chloroadenosine 5'-triphosphate (2-chloro-ATP), adenosine 5'-O-(1-thiotriphosphate) (ATP-alpha-S), P1, P5-diadenosine pentaphosphate (Ap5A) and adenylyl 5'-(beta,gamma-methylene)diphosphonate (AMPPCP) were apparently competitive antagonists, although only one concentration of each antagonist was used. There was a good correlation between the pA2 values found here for these antagonists including ATP, and their pA2 values reported for inhibition of ADP-induced aggregation. Adenosine 5'-(alpha, beta-methylene)triphosphate (AMPCPP) and uridine 5'-triphosphate (UTP) (100 microM) were only very weak inhibitors of the effect of ADP on intracellular calcium, and this is consistent with their weak actions as inhibitors of aggregation. 2-Methylthioadenosine 5'-triphosphate (2-methylthio-ATP) (50 microM) non-competitively inhibited the effect of ADP on intracellular calcium, in a very similar way to its inhibition of ADP-induced aggregation.4. The good correspondence found for these analogues between their effect on intracellular calcium and on aggregation confirms that there is a causal relationship between these actions of ADP, and that they are mediated by the same receptor on platelets. These findings cast further doubt on the use of the affinity reagent 5'-fluorosulphonylbenzoyladenosine (FSBA) as an antagonist and label for the ADP receptor, as this compound has been reported to inhibit aggregation but not ADP-induced increases in intracellular calcium.