Autoradiographic analyses of salmon calcitonin (sCT) binding in the rat mesencephalon revealed an exceptionally high concentration of receptors in the ventral and ventrolateral segments of the periaqueductal gray matter (PAG) extending along the entire rostral-caudal axis. Relatively heavy labeling was also seen along a band extending ventrolaterally through the mesencephalic reticular formation. Other receptor-rich areas include the nucleus linearis, pars compacta and lateralis of the substantia nigra, locus coeruleus, parabrachial nuclei and nucleus raphe pontis of the pontine reticular formation. Injections of sCT into the PAG induced a dose-dependent increase in hot-plate latencies. All rostral-caudal levels of these brain regions appeared to be equally responsive. Injections into the midline pontine reticular formation were also effective in increasing response latencies. Unilateral injections into the hypothalamus, medial thalamus, ventral thalamus and mesencephalic reticular formation proved to be ineffective. Human calcitonin (hCT) was considerably less potent. These biological effects are consistent with the potencies of both peptides in displacing 125I-sCT from slide-mounted sections of rat PAG. Naloxone failed to antagonize sCT-induced analgesia, suggesting an opiate independent mechanism for this peptide in eliciting analgesia.