Non-apoptotic externalization of phosphatidylserine (PS) can act as a reactive surface for the efficient assembly of the prothrombinase complex leading to thrombin generation and coagulation. Here we show that extracellular ATP, acting at the macrophage P2X₇ receptor, drives the rapid Ca²⁺-dependent formation and release of PS-rich microvesicles that enhance the assembly of the prothrombinase complex and subsequent formation of thrombin. Incubation with P2X₇ receptor antagonists (KN-62 and Brilliant Blue G) attenuates ATP induced prothrombotic responses. Consistent with the hypothesis that exposed PS enhances prothrombinase activity; pre-incubation with annexin V blocks the increase in thrombin formation. The rapid translocation of PS and formation of pro-thrombotic microvesicles occurs in the absence of cell lysis. These data demonstrate that the pro-inflammatory P2X₇ receptor can also support and propagate rapid increases in thrombin formation.