Immunohistochemical localization and biochemical characterization of two novel decapeptides derived from POMC-A in the trout hypothalamus

Several vertebrate species which underwent duplication of their genome, suc h as trout, salmon and Xenopus, possess two proopiomelanocortin (POMC) gene s. In the trout, one of the POMC molecules, called POMC-A, exhibits a uniqu e C-terminal extension of 25 amino acids which has no equivalent in other P OMCs characterized so far. This C-terminal peptide contains three pairs of basic residues, suggesting that it may be the source of novel regulatory pe ptides. The aim of the present study was to investigate the occurrence of t hese peptides in the brain of the trout Oncorhynchus mykiss by using specif ic antibodies raised against two epitopes derived from the C-terminal exten sion of POMC-A, i.e., EQWGREEGEE and YHFQ-NH2. Immunohistochemical labeling of brain sections revealed the presence of EQWGREEGEE- and YHFQ-NH2-immuno reactive cell bodies in the anterior part of the nucleus lateralis tuberis of the hypothalamus. Immunoreactive fibers were observed in the dorsal hypo thalamus, the thalamus, the telencephalon, the optic tectum and the medulla oblongata. In contrast, no labeling was detected using antibodies against the non-amidated peptide YHFQG. Biochemical characterization was performed by combining high-performance liquid chromatography (HPLC) analysis with ra dioimmunoassay (RIA) quantification. Two peptides exhibiting the same reten tion time as synthetic EQWGRE-EGEE and ALGERKYHFQ-NH2 were resolved. Howeve r, no peptide co-eluting with YHFQ-NH2 or YHFQG could be detected. These re sults demonstrate that, in the trout brain, post-translational processing o f POMC-A generates the two decapeptides EQWGREEGEE and ALGER-KYHFQ-NH2. The wide distribution of immunoreactive fibers in the diencephalon, telencepha lon, optic tectum and medulla ablongata suggests that these peptides may ex ert neurotransmitter and/or neuromodulator activities.