Doxepin inhibits GPVI-dependent platelet Ca²⁺ signaling and collagen-dependent thrombus formation

Platelet adhesion, activation, and aggregation are essential for primary hemostasis, but are also critically involved in the development of acute arterial thrombotic occlusion. Stimulation of the collagen receptor glycoprotein VI (GPVI) leads to phospholipase Cγ2-dependent inositol triphosphate (IP₃) production with subsequent platelet activation, due to increased intracellular Ca²⁺ concentration ([Ca²⁺]_i). Although tricyclic antidepressants have been shown to potentially impair platelet activation, nothing is hitherto known about potential effects of the tricyclic antidepressant doxepin on platelet Ca²⁺ signaling and thrombus formation. As shown in the present study, doxepin significantly diminished the stimulatory effect of GPVI agonist collagen-related peptide (CRP) on intracellular Ca²⁺ release as well as subsequent extracellular Ca²⁺ influx. Doxepin was partially effective by impairment of CRP-dependent IP₃ production. Moreover, doxepin abrogated CRP-induced platelet degranulation and integrin α_IIbβ₃ activation and aggregation. Finally, doxepin markedly blunted in vitro platelet adhesion to collagen and thrombus formation under high arterial shear rates (1,700^-s). In conclusion, doxepin is a powerful inhibitor of GPVI-dependent platelet Ca²⁺ signaling, platelet activation, and thrombus formation.