The corticotropin-releasing factor (CRF) superfamily of peptides includes the four paralogous vertebrate peptide lineages of CRF, urotensin-1/urocortin/sauvagine, urocortin 2 and urocortin 3, as well as the diuretic hormones (DH) and peptides of the arthropods. However, there are considerable sequence differences between the group of vertebrate peptides and those of the arthropods, notably insects. Because of the likely incidence of the formation of paralogous forms within the insects and the great variation in primary structures among these peptides, establishing the structure of the ancestral vertebrate version has not been possible. We screened the genomes of the tunicates, Ciona intestinalis and Ciona savignii, in silico, using the various conserved motifs of both the vertebrate CRF paralogues and the insect diuretic hormone sequences to identify the structure of the Ciona CRF/DH-like peptide genes. A single peptide gene was found in both genomes that possessed motifs reflective of both groups of peptides. These structures suggest a single CRF-like peptide was inherited by vertebrates and possibly chordates. Moreover, the conserved structure of the CRF peptide may have become constrained once it became associated with the regulation of the hypothalamus-pituitary-adrenal/interrenal axis.