Specific [3H]resiniferatoxin binding is thought to represent the postulated vanilloid (capsaicin) receptor. We have previously characterized [3H]resiniferatoxin binding to membranes from rat and pig dorsal root ganglia, which contain the cell bodies of capsaicin-sensitive primary afferent neurons. We now demonstrate specific binding of [3H]resiniferatoxin to particulate preparations from pig dorsal horn, which contains the central nerve endings of the capsaicin-sensitive primary afferent neurons. The Kd was 0.27 +/- 0.03 nM; the Bmax was 370 +/- 40 fmol/mg protein. Vanilloids of the capsaicin class (capsaicin, piperine, zingerone) and resiniferatoxin class (tinyatoxin, 12-deoxyphorbol 13-phenylacetate 20-homovanillate) inhibited binding with affinities consistent with their relative in vivo potencies. Given the interest in vanilloids as potential non-narcotic analgesic agents, this binding assay affords an attractive approach for characterization of structure-activity relations at spinal vanilloid receptors.