Schwartzentruber R S, Omeljaniuk R J
Department of Biology, Lakehead University, Thunder Bay, Ontario, Canada.
Life Sci. 1994;55(10):751-9. doi: 10.1016/0024-3205(94)00558-3.
We investigated the existence and nature of specific [3H]pGlu-3-Me-His-Pro-NH2 ([3H]MeTRH) binding sites in juvenile rainbow trout (Oncorhynchus mykiss) hypothalamus. Washed hypothalamic membranes were incubated with [3H]MeTRH in the absence (B0) or presence of pGlu-His-Pro-NH2 (TRH) or MeTRH under various experimental paradigms; incubations were terminated by filtration and bound radioactivity was determined by liquid scintillation spectroscopy. Specific binding (Bsp) was tissue dependent, associable, dissociable, and thermolabile. Estimated rates of association (k+1) and dissociation (k-1) were 1.64 x 10(7) M-1 min-1 and 1.98 x 10(-2) min-1, respectively, providing a kinetically derived dissociation rate constant (Kd) of 1.21 x 10(-9) M. [3H]MeTRH binding was displaceable; LIGAND-analysis of three independent homologous displacement experiments consistently indicated a single class of binding sites with an average Kd = 6.91 (+/- 4.32) x 10(-9) M and average maximum binding capacity (Bmax) of 8.84 (+/- 2.72) x 10(-15) mol/mg protein. Native TRH also displaced the radiolabel in a dose dependent manner; LIGAND-estimates for Kd and Bmax were 1.52 (+/- 0.12) x 10(-9) M and 3.79 (+/- 0.99) x 10(-15) mol/mg protein (n = 3 experiments), respectively. Our data indicate that presence of a single class of specific high-affinity TRH-binding sites in the rainbow trout hypothalamus; these findings suggest a role for TRH in regulating the release of hypophysiotrophic factors in the teleost hypothalamus.
