Corticotropin-releasing hormone (CRH) plays a central role in the physiological regulation of the hypothalamus-pituitary-adrenal/interrenal axis mediating endocrine, behavioral, autonomic, and immune responses to stress. Despite the wealth of knowledge about the physiological roles of CRH, the genetic mechanisms by which CRH neurons arise during development are poorly understood. As a first step toward analyzing the molecular and genetic pathways involved in CRH lineage specification, we describe the developmental distribution of CRH neurons in the embryonic zebrafish, a model organism for functional genomics and developmental biology. We searched available zebrafish expressed sequence tag (EST) databases for CRH-like sequences and identified one EST that contained the complete zebrafish CRH open reading frame (ORF). The CRH precursor sequence contained a signal peptide, the CRH peptide, and a cryptic peptide with a conserved sequence motif. RT-PCR analysis showed crh expression in a wide range of adult tissues as well as during embryonic and larval stages. By whole-mount in situ hybridization histochemistry, discrete crh-expressing cell clusters were found in different parts of the embryonic zebrafish brain, including telencephalon, preoptic region, hypothalamus, posterior tuberculum, thalamus, epiphysis, midbrain tegmentum, and rostral hindbrain and in the neural retina. The localization of crh mRNA within the preoptic region is consistent with the central role of CRH in the teleost stress response through activation of the hypothalamic-pituitary-interrenal axis. The widespread distribution of CRH-synthesizing cells outside the preoptic region suggests additional functions of CRH in the embryonic zebrafish brain.