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Members of the corticotropin-releasing factor (CRF) family of peptides play pivotal roles in the regulation of neuroendocrine, autonomic, and behavioral responses to physical and emotional stress. In amphibian tadpoles, CRF-like peptides stimulate both thy-roid and interrenal (adrenal) hormone secretion, and can thereby modulate the rate of metamorphosis. To better understand the reg-ulation of expression and actions of CRF in amphibians we developed a homologous radioimmunoassay (RIA) for Xenopus laevis CRF (xCRF). We validated this RIA and tissue extraction procedure for the measurement of brain CRF content in tadpoles and juveniles. We show that the CRF-binding protein, which is highly expressed in X. laevis brain, is largely removed by acid extraction and does not interfere in the RIA. We analyzed CRF peptide content in Wve microdissected brain regions in prometamorphic tad-poles and juveniles. CRF was detected throughout the brain, consistent with its role as both a hypophysiotropin and a neurotrans-mitter/neuromodulator. CRF content was highest in the region of the preoptic area (POa) and increased in all brain regions after metamorphosis. Exposure to 4 h of handling/shaking stress resulted in increased CRF peptide content in the POa in juvenile frogs. Injections of xCRF into prometamorphic tadpoles increased whole body corticosterone and thyroxine content, thus supporting Wnd-ings in other anuran species that this peptide functions as both a corticotropin- and a thyrotropin (TSH)-releasing factor. Further-more, treatment of cultured tadpole pituitaries with xCRF (100 nM for 24 h) resulted in increased medium content, but decreased pituitary content of TSH-immunoreactivity. Our results support the view that CRF functions as a stress neuropeptide in X. laevis a