GPR55 activation by several cannabinoids increases intracellular calcium in HEK293 cells and DRG neurons. (A) The time course of changes in [Ca2+]i in HEK293 cells loaded with fura-2 and perfused with THC. The lines above the figure indicate drug application. The solid line indicates a representative HEK293 cell transiently expressing the human GPR55 (hGPR55-HEK293) perfused with THC, whereas the black dashed line indicates a representative nontransfected HEK293 cell (HEK293 wt). (B) Summary of calcium responses upon perfusion of cannabinoid agonists. Responses in HEK293 wt cells (n = 6, black bars) and hGPR55-HEK293 cells (open bars) were perfused with THC, methanandamide (MEA), anandamide (AEA), or JWH015. (C) Immunostaining of mouse DRGs with a rabbit anti-GPR55 antibody (1:2,000, red) and a guinea pig anti-CB1 antibody (1:2,000, green) show that GPR55 is preferentially expressed in large-diameter DRG neurons (arrowheads) whereas CB1 is more widely expressed (arrows). See SI Text for full methods. (Scale bar, 50 μm.) (D) Representative JWH015-induced calcium rises in a large (solid line) and small (dashed line) DRG neuron. (E) Summary of calcium responses in small and large DRG neurons upon perfusion of THC, JWH015, and LPI.