Tyrosine hydroxylase (TH) is a rate-limiting enzyme in the biosynthesis of catecholamines. In zebrafish, two genes encoding TH have been identified. We cloned them and studied their expression in zebrafish. In adult tissues, th1 mRNA was more abundant than th2 mRNA in the brain and eyes, whereas th2 mRNA was more abundant in the liver, kidney, heart and gills. In developing brain, th1 mRNA was readily detected at 1 day post-fertilization using qPCR and in situ hybridization, whereas th2 mRNA appeared later. th1 was found in 17 catecholaminergic groups in larval brain, whereas th2 was found in four additional groups. A monoclonal antibody commonly used against TH detected preferentially TH1 protein. The two th genes, probably originated as a result of genome duplication, thus show complementary expression, although th1 is predominant in the brain and th2 in the periphery. th2 may be a novel essential factor in regulation of catecholamine synthesis in zebrafish.