The endocannabinoid 2-arachidonoylglycerol activates human platelets through non-CB₁/CB₂ receptors

Background: The endocannabinoid 2-arachidonoylglycerol (2-AG) is an endogenous lipid that acts through the activation of G-protein-coupled cannabinoid receptors and plays essential roles in many physiological contexts. In the cardiovascular system 2-AG is generated by both activated endothelial cells and platelets, and participates in the regulation of inflammation and thrombosis. Although human platelets actively metabolize endocannabinoids, 2-AG also binds to platelet surface and leads to cell activation. Objective: To investigate the biological consequence of 2-AG interactions with human platelets and to clarify the role of cannabinoid receptors. Methods: Gel-filtered platelets were stimulated with 2-AG inthe presence or absence of various inhibitors. Platelet aggregation and secretion were measured in a lumiaggregometer. Calcium ion movements were measured in FURA-2 loaded platelets. Thromboxane A₂ (TxA₂) generation was evaluated as Thromboxane B₂ accumulation with a commercial EIA assay. Results: 2-AG induced platelet shape change, aggregation and secretion with a dose-dependent mechanism that required engagement of platelet TxA₂ receptors. 2-AG caused also cytosolic calcium increase; however, it was totally dependent on availability of TxA₂. Indeed 2-AG was able to induce a robust generation of TxA₂ through the cyclooxygenase pathway. Treatment of platelets with inhibitors of monoacylglycerol lipase and fatty acid amide hydrolase did not affect the activation induced by 2-AG. Moreover, neither CB₁ and CB₂ proteins nor CB₁/ CB₂ mRNAs were detected in platelets. Conclusions: 2-AG canbe considered a new physiologic platelet agonist able to induce full platelet activation and aggregation with a non-CB₁/CB₂ receptor-mediated mechanism.