Immunohistochemical detection of teleost- and mammalian-type melanin-concentrating hormone in goldfish brain with newly developed specific antibodies.

In teleosts, melanin-concentrating hormone (MCH) is involved in body color change, feeding, and sleep regulation. The distribution of MCH neurons in the brain and pituitary must be clarified to understand diverse functions of MCH in the brain. This study aimed to discriminate neurons producing teleost-type MCH (MCH-like, MCHL) and mammalian-type MCH (MCH) in the brain and pituitary of goldfish Carassius auratus by immunohistochemistry using specific antibodies. Since the mature MCH region is not available for specific peptide antigen, we selected peptides derived from non-MCH regions on proMCH (PMCH). Nano Liquid Chromatography with tandem mass spectrometry analysis showed that both PMCHL and PMCH are cleaved into smaller peptides at mono- and dibasic-amino acid as well as non-basic sites. Based on these findings, PMCHLa (72-90) and PMCHa (88-102) were synthesized and used for immunization. The results of immunohistochemistry using thus prepared antibodies showed that neurons containing PMCHLa and PMCHa are differentially distributed in the hypothalamus and pituitary of goldfish. PMCHLa-immunoreactive (ir) fibers were detected in several brain regions, while PMCHa-ir fibers were confined to the hypothalamic area. The antibodies developed in the present study would contribute to clarify the role of the teleost-type and mammalian-type MCH.