Abstract

[³H]Resiniferatoxin (RTX) binding and calcium uptake by rat dorsal root ganglion (DRG) neurons show distinct structure-activity relations, suggestive of independent vanilloid receptor (VR) subtypes. We have now characterized ligand binding to rat VR1 expressed in human embryonic kidney (HEK293) and Chinese hamster ovary (CHO) cells and compared the structure-activity relations with those for calcium mobilization. Human embryonic kidney cells (HEK293/VR1 cells) and Chinese hamster ovary cells transfected with VR1 (CHO/VR1 cells) bound [³H]RTX with affinities of 84 and 103 pM, respectively, and positive cooperativity (Hill numbers were 2.1 and 1.8). These parameters are similar to those determined with rat DRG membranes expressing native VRs (a K _d of 70 pM and a Hill number of 1.7). The typical vanilloid agonists olvanil and capsaicin inhibited [³H]RTX binding to HEK293/VR1 cells with K _i values of 0.4 and 4.0 μM, respectively. The corresponding values in DRG membranes were 0.3 and 2.5 μM. HEK293/VR1 cells and DRG membranes also recognized the novel vanilloids isovelleral and scutigeral with similarK _i values (18 and 20 μM in HEK293/VR1 cells; 24 and 21 μM in DRGs). The competitive vanilloid receptor antagonist capsazepine inhibited [³H]RTX binding to HEK293/VR1 cells with a K _i value of 6.2 μM and binding to DRG membranes with a K _i value of 8.6 μM. RTX and capsaicin induced calcium mobilization in HEK293/VR1 cells with EC₅₀ values of 4.1 and 82 nM, respectively. Thus, the relative potencies of RTX (more potent for binding) and capsaicin (more potent for calcium mobilization) are similar in DRG neurons and cells transfected with VR1. We conclude that VR1 can account for both the ligand binding and calcium uptake observed in rat DRG neurons.