Gonadotropin releasing hormone binding sites in rat hippocampus: Different structure/binding relationships compared to the anterior pituitary

Abstract

In vitro autoradiography and radioreceptor assays were used to assess the binding characteristics of the gonadotropin releasing hormone (GnRH) binding sites in rat hippocampus. Competition studies with various GnRH agonists, antagonists, and fragments of GnRH indicate that the structure/binding requirement of the brain GnRH receptors is similar, but not identical, to that described in the pituitary. In the hippocampas, GnRH and the agonist d-Ala 6-GnRH exhibit an IC 50 for displacement of 125I-[ d-Ser(tBu) 6-Pro 9-NHEt]-GnRH (Buserelin, Hoechst) similar to that of the anterior pituitary; in contrast, the antagonist [ d- p-Glu 1- d-Phe 2- d-Trp 3,6]-GnRH bound only to the anterior pituitary GnRH receptor. Similarly, [His 5-Trp 7-Tyr 8]-GnRH (chicken II) recognized the pituitary GnRH receptor, but did not bind to hippocampal membranes. The difference in the binding affinities is likely not due to differential metabolism because of the stability of the tracer and of the antagonist and the addition of several enzyme inhibitors to the incubation buffer. The GnRH fragments [4–10]GnRH, [5–10]-GnRH, and [Ac5–10]-GnRH and [Gln 8]GnRH (chicken I), [Trp 7-Leu 8]-GnRH (salmon), and [Tyr 3-Leu 5-Glu 6-Trp 7-Lys 8]-GnRH (lamprey) did not displace the radiolabeled agonist binding in either the rat brain or the pituitary. Similarly, GTP and GTPγS had no effect on the binding of the GnRH agonist 125I-[ d-Ser(tBu) 6-Pro 9-NHEt]-GnRH (buserelin) to hippocampal or pituitary membrane preparations. Photoaffinity labeling of pituitary and hippocampal GnRH binding sites with 125I-[azidobenzoyl- d-Lys 6]-GnRH followed by one-dimensional gel electrophoresis in sodium dodecyl sulfate and by autoradiography revealed, in both tissues, two similar bands of proteins with apparent molecular weights of 29,000 and 60,000 Da. The results suggest that GnRH acts in the brain through specific binding sites that share some but not all of the characteristics of the pituitary receptor. Brain GnRH receptors are likely the site through which the neuropeptide can act to modify behavioral, sensory, and endocrine events.