Recently (J Pharmacol Exp Ther 261:580, 1992), we have shown that K562 leukemia cells express a calcium-signaling purinoceptor with characteristics of the P2T receptor subtype for adenosine diphosphate (ADP) previously found only in platelets. Because these results suggested that the P2T receptor may be an early marker for megakaryocytic differentiation, we studied whether this calcium- signaling receptor is also expressed in Dami cells, a human megakaryocytic leukemia cell line. Here we report evidence that Dami cells express a P2T receptor for ADP. The calcium response EC50 values for ADP, 2-methylthioadenosine diphosphate (2-MeS-ADP), and adenosine 5′-O-(2-thiodiphosphate) (ADP beta S) in Dami cells are 0.4 mumol/L, 0.04 mumol/L, and 2 mumol/L, respectively, which approximate the potencies of these agonists in K562 cells and in platelets. The platelet P2T receptor antagonists 2-methylthioadenosine triphosphate (2- MeS-ATP), and 2-chloroadenosine triphosphate (2-Cl-ATP) were surprisingly potent agonists at the P2T receptor in both Dami and K562 cells. Dami cells, unlike K562 cells and platelets, also respond to adenosine triphosphate (ATP) and uridine triphosphate (UTP) with an increase in intracellular calcium. Adenosine monophosphate (AMP) is an effective antagonist of the response to ADP, 2-MeS-ADP, ADP beta S, 2- MeS-ATP, and 2-Cl-ATP, but not to ATP and UTP. The responses to maximal concentrations of UTP in combination with either ADP, 2-MeS-ADP, ADP beta S, or 2-MeS-ATP are additive. In contrast, ADP in combination with either 2-MeS-ADP, ADP beta S, 2-MeS-ATP, or 2-Cl-ATP are not additive. UTP desensitized Dami cells to ATP but not to ADP, 2-MeS-ADP, ADP beta S, or 2-MeS-ATP. Addition of ATP after UTP desensitization antagonized subsequent responsiveness to ADP. The data suggest that the receptor for ADP may be a unique P2T subtype, and the receptor for ATP and UTP is distinct from that of ADP and is most characteristic of the P2U (nucleotide) receptor subtype. Activation of either the P2T or P2U receptor causes a rapid generation of inositol trisphosphate in Dami cells.