Zinc is concentrated in the dorsal horn of the spinal cord and has been proposed to alter excitability of primary afferent C-fibers, structures believed to be important in nociceptive transmission. Based on the inhibitory effect of zinc on the activity of various other neurotransmitters that play a role in nociception, we tested the hypothesis that zinc modulates pain transmission. To test this, we examined the effect of exogenous zinc, administered intrathecally (i.t.), on nociception in the mouse. We also assessed the impact of decreased concentrations of endogenously occurring zinc in the extracellular fluid brought about by an i.t. injection of either ethylenediaminetetraacetic acid disodium-calcium salt (Ca++EDTA), a calcium-saturated, membrane-impermeable chelator of divalent cations, or of dipicolinic acid, a zinc chelator. Injection of zinc produced a dose-related antinociceptive effect, optimal at 90 min in the writhing assay, but had no effect on tail-flick response latencies. In contrast, injection of either Ca++EDTA or dipicolinic acid produced a dose-related hyperalgesia in the tail-flick assay at 90 min after injection. Responses induced in the writhing assay were unaffected by Ca++EDTA. Although zinc had no effect on thermal nociception, the hyperalgesic effect of Ca++EDTA was antagonized by coadministration of Ca++EDTA with zinc. Similarly, the antinociceptive effect of zinc on writhing responses was attenuated when coadministered with Ca++EDTA. Zinc also inhibited primary afferent C-fiber activity because 10 ng of zinc i.t. inhibited the behavioral response induced by injection i.t. of 1 nmol of capsaicin. Neither zinc nor Ca++EDTA altered writhing or tail-flick latencies, respectively, when injected intracerebroventricularly. These findings support the hypothesis that endogenous zinc, localized in the dorsal horn of the spinal cord, plays a role in the regulation of pain.