The difficulty of conducting electrophysiologic recordings from the platelet has restricted investigations into the role of ion channels in thrombosis and hemostasis. We now demonstrate that the well-established synergy between P2Y₁ and P2Y₁₂ receptors during adenosine diphosphate (ADP)–dependent activation of the platelet α_IIbβ₃ integrin also exists in murine marrow megakaryocytes, further supporting the progenitor cell as a bona fide model of platelet P2 receptor signaling. In patch clamp recordings, ADP (30 μM) stimulated a transient inward current at –70 mV, which was carried by Na⁺ and Ca²⁺ and was amplified by phenylarsine oxide, a potentiator of certain transient receptor potential (TRP) ion channels by phosphatidylinositol 4,5-bisphosphate depletion. This initial current decayed to a sustained phase, upon which repetitive transient inward cation currents with pre-dominantly P2X₁-like kinetics were super-imposed. Abolishing P2X₁-receptor activity prevented most of the repetitive currents, consistent with their activation by secreted adenosine triphosphate (ATP). Recordings in P2Y₁-receptor–deficient megakaryocytes demonstrated an essential requirement of this receptor for activation of all ADP-evoked inward currents. However, P2Y₁₂ receptors, through the activation of PI3-kinase, played a synergistic role in both P2Y₁ and P2X₁-receptor–dependent currents. Thus, direct stimulation of P2Y₁ and P2Y₁₂ receptors, together with autocrine P2X₁ activation, is responsible for the activation of nonselective cation currents by the platelet agonistADP.