The ontogeny of TRH and of a proposed TRH metabolite, histidyl proline diketopiperazine (His-Pro DKP), was determined in the rat central nervous system and pancreas as a means of studying the interrelationship of these peptides. Various regions of the rat brain, spinal cord, and pancreas were dissected from animals ranging in age from prenatal day 17 to adult. The tissues were extracted for TRH and His-Pro DKP, and tissue levels of the two peptides were measured by specific RIAs. We found increasing TRH levels in the hypothalamus, spinal cord, and multiple extrahypothalamic brain regions in the developing rat [e.g. from 21 +/- 3 (+/- SE) pg/hypothalamus on prenatal day 17 to 2606 +/- 296 pg/hypothalamus in the adult]. In the rat pancreas, however, TRH levels initially increased from 354 +/- 37 pg/pancreas on prenatal day 21 to 749 +/- 68 pg/pancreas on postnatal day 7, but from day 7 to adulthood, the TRH content fell dramatically, being undetectable in the adult rat pancreas. The His-Pro DKP content increased in nearly all tissues studied, with peak values occurring on postnatal days 10 and 28 and in the adult. There was little apparent correlation, however, between the anatomical distribution and ontogeny of TRH compared with those of His-Pro DKP. We conclude that His-Pro DKP and TRH have widespread distributions involving the hypothalamus, extrahypothalamic brain, spinal cord, and pancreas in the developing rat. TRH and His-Pro DKP, however, have differing patterns of ontogeny in the rat, suggesting that His-Pro DKP may be derived from sources other than just TRH.