Integrin clustering enables anandamide-induced Ca²⁺ signaling in endothelial cells via GPR55 by protection against CB₁-receptor-triggered repression

Although the endocannabinoid anandamide is frequently described to act predominantly in the cardiovascular system, the molecular mechanisms of its signaling remained unclear. In human endothelial cells, two receptors for anandamide were found, which were characterized as cannabinoid 1 receptor (CB₁R; CNR1) and G-protein-coupled receptor 55 (GPR55). Both receptors trigger distinct signaling pathways. It crucially depends on the activation status of integrins which signaling cascade becomes promoted upon anandamide stimulation. Under conditions of inactive integrins, anandamide initiates CB₁R-derived signaling, including G_i-protein-mediated activation of spleen tyrosine kinase (Syk), resulting in NFB translocation. Furthermore, Syk inhibits phosphoinositide 3-kinase (PI3K) that represents a key protein in the transduction of GPR55-originated signaling. However, once integrins are clustered, CB₁R splits from integrins and, thus, Syk cannot further inhibit GPR55-triggered signaling resulting in intracellular Ca²⁺ mobilization from the endoplasmic reticulum (ER) via a PI3K-Bmx-phospholipase C (PLC) pathway and activation of nuclear factor of activated T-cells. Altogether, these data demonstrate that the physiological effects of anandamide on endothelial cells depend on the status of integrin clustering.