In mammals, the pituitary hormone prolactin is also produced in various extrapituitary tissues and may act in an auto/paracrine fashion. To explore the comparative aspects of extrapituitary prolactin, the distributions of prolactin transcripts were investigated in extrapituitary organs of the goldfish, African clawed frog and mouse by reverse transcription-polymerase chain reaction (RT-PCR). For comparison, the amount of the transcript in mouse tissues was also estimated using competitive RT-PCR. In the goldfish, the transcript was detected in the ovary, testis, liver, kidney, spleen, gill, muscle and brain in slightly lower abundance than in the pituitary, but not detected in the intestine. In the frog, the transcript was detected in the following organs with an order of abundance: pituitary >> brain > testis and ovary. In the mouse, the transcript was detected in the pituitary, brain, testis, and ovary, and its copy number per μg of total RNA was estimated at ~10¹⁰ in the male pituitary, ~10⁴ in the placenta, hypothalamus and testis, ~10³ in the thymus and experimentally induced deciduoma, and ~10² in the ovary. These results suggest that the origin of extrapituitary prolactin goes back to the common ancestor of fish and tetrapods, but that distinct evolution has occurred in each lineage. The significance of extrapituitary PRL in non-mammalian species is also suggested.