Chemokines released by inflamed endothelium induce proaggregatory properties of platelets which represent an important link between vascular inflammation, thrombosis, and atherogenesis. The recently discovered chemokine CXCL16 has been proposed as a pathogenic mediator in atherosclerosis and coronary artery disease. CXCL16 is highly expressed in inflamed vascular tissue of atherosclerotic lesions, but its influence on platelet function remained elusive. Western blotting, confocal microscopy and FACS analysis revealed that the CXCL16-specific receptor CXCR6 (BONZO) is highly expressed in platelets. According to FACS-analysis platelet stimulation in vitro with increasing concentrations of CXCL16 significantly enhanced intracellular calcium, significantly upregulated expression of P-Selectin and activated integrin a_IIbb₃ on the platelet surface. Furthermore confocal microscopy and western blotting revealed a drastic change of shape and of cytoskeletal components. CXCL16-mediated effects on platelet activation was inhibited by wortmannin, LY294002, BX-912 and SH-6 suggesting involvement of phosphatidylinositol 3-kinase (PI3K), 3-phosphoinositide-dependent protein kinase 1 (PDK-1) and protein kinase B (PKB)/Akt. Additional flow cytometric measurements revealed that after CXCL16 treatment exposure of P-selectin and activation of integrin a_IIbb₃ were significantly lower in platelets from Akt1 deficient mice (akt1^-/-) than in platelets from respective wild type mice (akt1^+/+). In a flow chamber model of platelet adhesion, stimulation with CXCL16 significantly enhanced platelet tethering and firm adhesion on IL1b-stimulated endothelial cells. Enhanced adhesion could be prevented by inhibitors of PI3K/Akt signaling pathway. Our studies demonstrate a powerful effect of the inflammatory chemokine CXCL16 on platelet activation and adhesion suggesting a role for CXCL16 in the pathogenesis of atherosclerosis and thrombo-occlusive diseases.