The effect of putative GPR18 agonists on intracellular Ca2+ mobilisation was assessed using FlexStation and single cell Ca2+ imaging techniques with different Ca2+ probes (fluo 4 and fura 2). NAGly induced ERK phosphorylation was quantified by immunoblotting.The tested panel of cannabinoid ligands did not observed to elicit a significant ligand mediated phosphorylation of ERK1/2 or mobilisation of intracellular Ca2+ in GPR18 expressing HEK293TR cells, BV 2 cells, INS 1 832/13 cells, Caco 2 cells and GPR18 transfected CHO cells heterologously expressing CB2 receptors.In this study recombinant human GPR18 receptor was not activated by NAGly or other cannabinoid ligand suggested by previous studies which indicated that NAGly may be not the natural endogenous ligand of GPR18 and argue against the deorphanization of GPR18 and assignment as a candidate cannabinoid receptor. Also it can concluded that the activation of GPR18 signalling mechanism may involve prodigious pathways not examined in the current study..
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