Melanin-concentrating hormone (MCH) is a neuropeptide generated in neurons originating in the hypothalamus, from which axons project to the entire brain and neurohypophysis in fish. MCH has both central and peripheral roles such as food intake and body color change. Here we cloned two MCH receptors (MCH-R) from the barfin flounder, Verasper moseri, Pleuronectiformes. The phylogenetic analysis shows that these are orthologues to the mammalian MCH-R1 and MCH-R2 showing 49 and 30% amino acid sequence identity to the corresponding human receptors while they have 31% amino acid sequence identify between them. Essential amino acid residues for ligand binding, signal transduction and receptor conformation, which have been shown in mammalian MCH-R, are well conserved in the flounder MCH-Rs. MCH-R1 has one intron in the extracellular N-terminal region and MCH-R2 has one intron in the DRY motif, which is a homologous position to one of the five introns of human MCH-R2. Orthologues of MCH-R1 and MCH-R2 may have appeared by gene duplication of the ancestry of MCH-Rs having at least two introns, and then MCH-R1 and MCH-R2 inherited different introns in flounder strains. We also determined their tissue distribution and functional role in rearing condition. Reverse transcription PCR revealed that the expression of MCH-R1 is confined to the brain of the barfin flounder, while transcripts of MCH-R2 were detected in the brain, pituitary, eyeball, gill, atrium, ventricle, head kidney, body kidney, spleen, intestine, inclinator, skeletal muscle testis, ovary, eyed-side skin, and non-eyed-side skin. The expression of MCH-R2 in eyed-side skin was higher in fish reared in a black tank (121 days) than in a white tank while the expression levels of MCH in the brain were significantly greater in the group reared with the white background suggesting down-regulation of this receptor gene with increased levels of MCH. The results suggest that the MCH-R2 mediates the effect of MCH to control body color for background adaptation in the eyed-side skin of the barfin flounder.