Binding and effects of catecholestrogens on adenylate cyclase activity, and adrenoceptors, benzodiazepine and GABA receptors in guinea-pig hypothalamic membranes.
Catecholestrogen (CE) binding to guinea-pig hypothalamic membranes was assessed by using [3H]2-hydroxyestrone (2-OHE1) as a ligand. Binding was maximal at pH 7.4 and 37 degrees C, and after 10 min incubations. A high affinity binding site with dissociation constant (KD) = 0.20 +/- 0.02 nM and site concentration (Bmax) = 38 +/- 2 fmol/mg protein, and a low affinity binding site with KD = 235 +/- 10 nM and Bmax = 4.2 +/- 1.0 pmol/mg protein (n = 7) were detected. The order of affinity (Ki, microM) for displacement of 10 nM [3H]2-OHE1 from hypothalamic binding sites was 2-OHE1 (0.8), 2-hydroxyestradiol (2-OHE2) (1.0), epinephrine (5.9), norepinephrine (NE) (7.7), dopamine (270), estradiol, estrone, propranolol, phentolamine, domperidone (greater than 10 000). NE inhibition of 2-OHE1 binding was non-competitive, Only 0.5 mM 2-OHE2 depressed in a non-competitive way the hypothalamic beta-adrenoceptor binding (measured by using [3H]dihydroalprenolol) without affecting alpha-adrenoceptor binding (measured by using [3H]dihydroergocryptine). Both 2-OHE2 and 2-OHE1 impaired NE-stimulated adenylate cyclase activity in hypothalamic membranes with EC50 of about 5 and 10 microM, respectively. CE decreased [3H]gamma-aminobutyric acid binding by hypothalamic membranes with Ki = 8 microM (2-OHE2) and 50 microM (2-OHE1). The binding of [3H]flunitrazepam to the same membrane preparation was not affected by CE. These results support the existence of significant CE binding and effects in guinea-pig hypothalamic membranes.
D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D002393:Estrogens, Catechol / Q000378:metabolism* / Q000494:pharmacology; D006168:Guinea Pigs; D007031:Hypothalamus / Q000201:enzymology / Q000378:metabolism*; D066298:In Vitro Techniques; D009638:Norepinephrine / Q000494:pharmacology; D011941:Receptors, Adrenergic / Q000378:metabolism*; D011963:Receptors, GABA-A / Q000378:metabolism*
D000067956:Adenylyl Cyclase Inhibitors; D002393:Estrogens, Catechol; D011941:Receptors, Adrenergic; D011963:Receptors, GABA-A; D009638:Norepinephrine