The cDNA encoding prepro-thyrotropin-releasing hormone (ppTRH) in a teleost, medaka ( Oryzias latipes) was isolated and characterized. The medaka ppTRH cDNA codes for 270 amino acid residues including eight TRH progenitor sequences (-Lys/Arg-Arg-Gln-His-Pro-Gly-Lys/Arg-Arg-). In silico analyses of the medaka genome database predicted that the structure of the medaka ppTRH gene is similar to the ppTRH genes of the other vertebrate species studied to date; consisting of three exons and two introns. Identity of the medaka ppTRH with the other vertebrates is rather low except the sockeye salmon. A molecular phylogenic tree showed that the ppTRH sequences reflected the predicted pattern of species classification. RT-PCR analysis demonstrated ppTRH gene expression in the brain and retina. These results gave some insight into the molecular evolution of ppTRH and physiological functions of TRH in vertebrates.