Abstract-At sites of vascular injury, platelets come into contact with the subendothelial extracellular matrix which triggers their activation and the formation of a hemostatic plug. This process is crucial for normal hemostasis, but may also lead to pathological thrombus formation causing diseases such as myocardial infarction or stroke. The initial capture of flowing platelets is mediated by the interaction of the glycoprotein (GP) Ib-V-IX complex with von Willebrand factor (vWF) immobilized on exposed collagens. This interaction allows the binding of the collagen receptor GPVI to its ligand and to initiate cellular activation, a process that is reinforced by locally produced thrombin and soluble mediators released from platelets. These events lead to the shift of ␤1 and ␤3 integrins on the platelet surface from a low to a high affinity state, thereby enabling them to bind their ligands and to mediate firm adhesion, spreading, coagulant activity, and aggregation. This review summarizes the most important structural and functional properties of these adhesion receptors and briefly discusses their potential as targets for antithrombotic therapy.

Summary. Agonist-induced elevation in cytosolic Ca 2+ concentrations is essential for platelet activation in hemostasis and thrombosis. It occurs through Ca 2+ release from intracellular stores and Ca 2+ entry through the plasma membrane (PM). Ca 2+ store release is a well-established process involving phospholipase (PL)C-mediated production of inositol-1,4,5-trisphosphate (IP 3 ), which in turn releases Ca 2+ from the intracellular stores through IP 3 receptor channels. In contrast, the mechanisms controlling Ca 2+ entry and the significance of this process for platelet activation have been elucidated only very recently. In platelets, as in other non-excitable cells, the major way of Ca 2+ entry involves the agonist-induced release of cytosolic sequestered Ca 2+ followed by Ca 2+ influx through the PM, a process referred to as store-operated calcium entry (SOCE). It is now clear that stromal interaction molecule 1 (STIM1), a Ca 2+ sensor molecule in intracellular stores, and the four transmembrane channel protein Orai1 are the key players in platelet SOCE. The other major Ca 2+ entry mechanism is mediated by the direct receptor-operated calcium (ROC) channel, P2X 1 . Besides these, canonical transient receptor potential channel (TRPC) 6 mediates Ca 2+ entry through the PM. This review summarizes the current knowledge of platelet Ca 2+ homeostasis with a focus on the newly identified Ca 2+ entry mechanisms.

Platelet activation and aggregation at sites of vascular injury are essential for primary hemostasis, but are also major pathomechanisms underlying myocardial infarction and stroke. Changes in [Ca 2؉ ] i are a central step in platelet activation. In nonexcitable cells, receptormediated depletion of intracellular Ca 2؉ stores triggers Ca 2؉ entry through storeoperated calcium (SOC) channels. STIM1 has been identified as an endoplasmic reticulum (ER)-resident Ca 2؉ sensor that regulates store-operated calcium entry (SOCE) in immune cells and platelets, but the identity of the platelet SOC channel has remained elusive. Orai1 (CRACM1) is the recently discovered SOC (CRAC) channel in T cells and mast cells but its role in mammalian physiology is unknown. Here we report that Orai1 is strongly expressed in human and mouse platelets. To test its role in blood clotting, we generated Orai1-deficient mice and found that their platelets display severely defective SOCE, agonist-induced Ca 2؉ responses, and impaired activation and thrombus formation under flow in vitro. As a direct consequence, Orai1 deficiency in mice results in resistance to pulmonary thromboembolism, arterial thrombosis, and ischemic brain infarction, but only mild bleeding time prolongation. These results establish Orai1 as the long-sought platelet SOC channel and a crucial mediator of ischemic cardiovascular and cerebrovascular events. IntroductionAt sites of vascular injury the subendothelial extracellular matrix (ECM) is exposed to the flowing blood and triggers sudden platelet activation and the formation of a fibrin-containing thrombus. This process is essential to prevent excessive posttraumatic blood loss, but if it occurs at sites of atherosclerotic plaque rupture it can also lead to vessel occlusion and the development of myocardial infarction or ischemic stroke, which are among the leading causes of mortality and severe disability in industrialized countries. 1,2 Therefore, the inhibition of platelet activation has become an important strategy to prevent or treat such acute ischemic events. 3 Platelet activation can occur through different signaling pathways that culminate in the activation of phospholipase C (PLC) isoforms and production of diacylglycerol (DAG) and inositol 1,4,5-triphosphate (IP 3 ). IP 3 binds its receptors on the membrane of the intracellular Ca 2ϩ stores and mediates Ca 2ϩ release into the cytosol. In platelets, the dense tubular system, referred to as sarcoplasmic reticulum (SR), is thought to be the major Ca 2ϩ store. The resulting decline in Ca 2ϩ store content in turn triggers a sustained influx of extracellular Ca 2ϩ by a mechanism known as store-operated Ca 2ϩ entry (SOCE). 4,5 Although SOCE has pharmacologically been well defined for more than a decade, not much was known about the underlying molecular machinery until recently, when stromal interaction molecule 1 (STIM1) was identified as a sarco/ endoplasmic (SR/ER)-store calcium sensor that controls SOCE in T cells and mast cells. [6][7][8][9] Shortly after that, ...

Platelet activation and aggregation are essential to limit posttraumatic blood loss at sites of vascular injury but also contributes to arterial thrombosis, leading to myocardial infarction and stroke. Agonist-induced elevation of [Ca2+]i is a central step in platelet activation, but the underlying mechanisms are not fully understood. A major pathway for Ca2+ entry in nonexcitable cells involves receptor-mediated release of intracellular Ca2+ stores, followed by activation of store-operated calcium (SOC) channels in the plasma membrane. Stromal interaction molecule 1 (STIM1) has been identified as the Ca2+ sensor in the endoplasmic reticulum (ER) that activates Ca2+ release–activated channels in T cells, but its role in mammalian physiology is unknown. Platelets express high levels of STIM1, but its exact function has been elusive, because these cells lack a normal ER and Ca2+ is stored in a tubular system referred to as the sarcoplasmatic reticulum. We report that mice lacking STIM1 display early postnatal lethality and growth retardation. STIM1-deficient platelets have a marked defect in agonist-induced Ca2+ responses, and impaired activation and thrombus formation under flow in vitro. Importantly, mice with STIM1-deficient platelets are significantly protected from arterial thrombosis and ischemic brain infarction but have only a mild bleeding time prolongation. These results establish STIM1 as an important mediator in the pathogenesis of ischemic cardio- and cerebrovascular events.