To investigate the consequences of freshwater (FW) transfer, we studied the prolactin (PRL) cDNA sequence and its mRNA expression, and physiological responses in the black porgy (Acanthopagrus schlegeli). We cloned and characterized cDNA encoding its PRL from the pituitary gland. Black porgy PRL cDNA consists of 1492 bp and encodes a protein of 212 amino acids including 24 signal peptides. Reverse transcription-PCR showed the PRL mRNA expression in the pituitary gland. Expression of pituitary gland PRL mRNA was significantly higher during FW acclimation. Furthermore, we studied the stress responses and osmoregulatory abilities of black porgy in changing salinities. Plasma cortisol, glucose, aspartate aminotransferase (AST) and alanine aminotransferase (ALT) significantly increased in the fish immediately after transfer to FW. We also identified significant changes in the fish in terms of plasma ions (Na(+), Cl(-), Ca(2+)) and osmolality during the acclimation period. These results suggests that PRL plays an important role in hormonal regulation in osmoregulatory organs, thereby improving the hyperosmoregulatory ability of black porgy in freshwater.