Properties of [3H](3-Me-His2)TRH binding to apparent TRH receptors in the sheep central nervous system

Abstract

[3H](3-methyl-His2)thyrotropin releasing hormone ([3H]MeTRH) binds to sites in the sheep central nervous system (CNS) whose properties closely resemble both those of CNS binding sites for [3H]TRH and those of pituitary binding sites for [3H]MeTRH. Detailed studies for binding of [3H]MeTRH in the sheep nucleus accumbens and retina have yielded equilibrium dissociation constants of about 4 nM and densities of binding sites of about 3 and 2 pmol/g wet weight, respectively. The binding affinity of [3H]MeTRH was 8- to 10-fold higher than that of [3H]TRH, resulting in much lower non-specific binding with the new ligand. The association reaction had a rate constant of about 2-3 x 10(7) M-1 min-1, while the biphasic dissociation reaction had rate constants of 8-9 x 10(-2) min-1 for the fast phase and 1-2 x 10(-2) min-1 for the slow phase. The regional distribution of binding in the sheep CNS was similar to that observed previously with [3H]TRH. Highest binding outside the pituitary was in the nucleus accumbens area and retina, with another peak in the amygdala-temporal cortex area. Binding was widely distributed, so that no CNS region appeared totally devoid of binding. Nineteen TRH analogs, ranging in potency over 6 orders of magnitude, showed nearly identical abilities to complete for binding of [3H]MeTRH in the CNS areas and in the sheep anterior pituitary gland in side-by-side experiments. These findings argue strongly for identification of [3H]MeTRH binding sites in the CNS as TRH receptors.