The vanilloid receptor type 1 (VR1) is a heat-activated ionophore preferentially expressed in nociceptive neurons of trigeminal and dorsal root ganglia (DRG). VR1, which binds and is activated by capsaicin and other vanilloid compounds, was noted to interact with the endocannabinoid anandamide (ANA) and certain inflammatory metabolites of arachidonic acid in a pH-dependent manner. At pH < or = 6.5 ANA induced (45)Ca(2+) uptake either in primary cultures of DRG neurons or cells ectopically expressing C-terminally tagged recombinant forms of VR1 with an EC(50) = approximately 10 microm at pH 5.5. Capsazepine, a potent antagonist of vanilloids, inhibited ANA-induced Ca(2+) transport in both cell systems. Vanilloids displaced [(3)H]ANA in VR1-expressing cells, suggesting competition for binding to VR1. Ratiometric determination of intracellular free calcium and confocal imaging of the VR1-green fluorescent fusion protein revealed that, at low pH (< or =6.5), ANA could induce an elevation of intracellular free Ca(2+) and consequent intracellular membrane changes in DRG neurons or transfected cells expressing VR1. These actions of ANA were similar to the effects determined previously for vanilloids. The ligand-induced changes in Ca(2+) at pH < or = 6.5 are consistent with the idea that ANA and other eicosanoids act as endogenous ligands of VR1 in a conditional fashion in vivo. The pH dependence suggests that tissue acidification in inflammation, ischemia, or traumatic injury can sensitize VR1 to eicosanoids and transduce pain from the periphery.