Somatolactin (SL) is a pituitary hormone belonging to the growth hormone/prolactin family of adenohypophyseal hormones. In teleost fish, SL is encoded by one or two paralogous genes, namely SL-α and -β. Our previous studies have revealed that pituitary adenylate-cyclase-activating polypeptide stimulates SL release from cultured goldfish pituitary cells, whereas melanin-concentrating hormone suppresses this release. As in other fish, the goldfish possesses SL-α and -β. So far, however, no useful means of detecting the respective SLs immunologically in this species has been possible. In order to achieve this aim, we raised rabbit antisera against synthetic peptide fragments deduced from the goldfish SL-α and -β cDNA sequences. Using these antisera, we observed adenohypophyseal cells showing SL-α- and -β-like immunoreactivities in the goldfish pituitary, especially the pars intermedia (PI). Several cells in the PI showed the colocalization of SL-α- and -β-like immunoreactivities. Then, using single-cell polymerase chain reaction with laser microdissection, we examined SL-α and -β gene expression in adenohypophyseal cells showing SL-α- or -β-like immunoreactivity. Among cultured pituitary cells, we observed three types of cell: those that possess transcripts of SL-α, -β, or both. These results suggest a polymorphism of SL-producing cells in the goldfish pituitary.