Three TRH-like molecules are released from rat hypothalamus in vitro

TRH-like immunoreactivity distinct from TRH is present in various tissues a nd fluids. In order to determine whether TRH-like molecules are secreted by the hypothalamus, we analyzed tissues and media from hypothalamic slices i ncubated in Krebs Ringer bicarbonate. Media from basal or high KCl conditio ns contained 3 TRH-like molecules evidenced by reverse phase high performan ce liquid chromatography followed by TRH radioimmunoassay. Peak I correspon ded to authentic TRH (73% of total immunoreactivity) and peaks II and III h ad a higher retention time. These additional TRH-like forms were neither de tected in hypothalamic tissue nor in tissue or medium from olfactory bulb. Gel filtration analysis of hypothalamic media revealed only one TRH-like pe ak eluting as TRH, suggesting that the molecular weights of peaks II and II I are similar to that of TRH. Peak II retention time was similar to that of pglu-phe-proNH(2). We analysed if they could be produced by post secretory metabolism of TRH. Incubation of hypothalamic slices with [H-3-Pro]-TRH di d not produce radioactive species comigrating with peaks II or III. However , it induced rapid degra dation to [H-3-Pro]-his-prodiketopiperazine ([H-3] -HPDKP). Inhibitor profile suggested that pyroglutamyl aminopeptidase II, b ut not pyroglutamyl aminopeptidase I, is responsible for [H-3]-HPDKP produc tion. These data are consistent with the hypothesis that pyroglutamyl amino peptidase II is the main aminopeptidase degrading TRH in hypothalamic extra cellular fluid. Furthermore, we suggest that the hypothalamus releases addi tional TRH-like molecules, one of them possibly pglu-phe-proNH(2), which ma y participate in control of adenohypophyseal secretions.