KAVANAUGH, S.I.*; SOWER, S.A.; Univ. of New Hampshire: Characterization of gonadotropin-releasing hormone (GnRH) in the Atlantic hagfish Myxine glutinosa

Hagfish are the oldest lineage of vertebrates with a lineage extending over 530 million years. They are in a key position between protochordates and vertebrates and they retain characteristics of extinct ancestral species that are common to their closest relatives. Gonadotropin-releasing hormone (GnRH) is the essential hypothalamic regulatory neurohormone controlling reproduction in all vertebrates. We recently showed a correlation of immunoreactive GnRH associated with gonadal maturation, estradiol, and progesterone production in the Atlantic hagfish Myxine glutinosa, however, the primary structure(s) of GnRH have not been identified in hagfish. Therefore, the objective of this study was to determine the primary structure(s) of GnRH present in M. glutinosa by proteomic methods using Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF). One thousand M. glutinosa were collected from the Gulf of Maine during October 2005. The brains were extracted in buffer using pressure cycling technology (Barocycler 2017, Pressure BioSciences, Inc.) and further purified using isoelectric focusing with immobilized pH gradient strips in conjunction with 2D gel electrophoresis. Immunoreactive bands were eluted, dried down and reconstituted for ZipTip purification, then subjected to analysis on Shimadzu�s Axima MALDI-TOF curved field reflectron (CFR) using post source decay and Shimadzu�s Axima MALDI-TOF quadrapole ion trap (QIT) collision induced dissociation. To date, the data suggest there are at least two distinct forms of GnRH in the hagfish brain which supports earlier immunocytochemical, and chromatographical studies. This research is funded by National Science Foundation Grant # 0421923 to SAS.