Cholinergic muscarinic receptor binding was measured in the preoptic area (POA) and whole hypothalamus (HTH) of adult Sprague-Dawley rats using the tritiated antagonist quinuclidinyl benzilate ([3H]QNB) as the ligand. Binding of [3H]QNB expressed as fmol/mg protein was 30% higher in POA than in HTH from gonadectomized rats. Cyclic changes were observed in the POA with the highest binding at proestrus and the lowest binding at diestrus. In HTH, no significant changes occurred over the estrous cycle. Estrogen treatment (10 micrograms of estradiol benzoate (EB)/120 g b. wt./48 and 24 h before sacrifice) increased [3H]QNB binding by 42% in the POA and 17% in HTH, relative to the ovariectomized controls. The enhancement of [3H]QNB binding in POA as compared with controls was evident with both the filtration and the centrifugation methods, although binding levels were higher when centrifugation assay was used. A lower estrogen dose (2 micrograms EB/rat/48 and 24 h before sacrifice) which is routinely used to activate lordotic behavior in female rats increased muscarinic binding by 26% in the POA but had no appreciable effect in HTH. A significant sex difference was found in the ability of estrogen to induce [3H]QNB binding in the central nervous system (CNS). Estrogen was ineffective in altering [3H]QNB binding in either brain region of castrated males, although the level and pattern of cholinergic binding between untreated gonadectomized males and females were similar.2+ These data suggest that physiological changes in estrogen secretion over the estrous cycle are capable of modulating cholinergic muscarinic binding in the POA and these changes may be of physiological relevance.(ABSTRACT TRUNCATED AT 250 WORDS)