The existence of typical muscarinic receptors in membranes of sheep anterior pituitary was detected by binding of [3H]quinuclidinyl benzilate ([3H]QNB), a potent and specific muscarinic antagonist. [3H]QNB binding sites in anterior pituitary had an equilibrium dissociation constant of about 20-40 pM, a rate constant for association at 37 degrees C of about 2 x 10(8) M-1 min-1, a rate constant for dissociation at 37 degrees C of about 4 x 10(-3) min-1, and the expected specificities for a variety of cholinergic and other drugs. The concentration of [3H]QNB binding sites in anterior pituitary, about 4-6 pmol/g tissue, was at least twice that in posterior pituitary, but less than a quarter that in the hypothalamus. Since there is no established cholinergic or other innervation of the anterior pituitary, the presence there of typical muscarinic receptors, if we assume that they are functional, suggests that acetylcholine reaches the tissue through the hypophyseal portal circulation, and that it may thus have a role in the regulation of pituitary function.