Both within the brain and in the periphery, M1muscarinic receptors function primarily as postsynaptic receptors and M2muscarinic receptors function primarily as presynaptic autoreceptors. In addition to classical parasympathetic effectors, cholinergic stimulation of central muscarinic receptors influences the release of adrenocorticotrophic hormone (ACTH) and corticosterone. We previously reported that oxotremorine administration to male and female M2receptor knockout and wild‐type mice increased ACTH to a significantly greater degree in knockout males compared to all other groups, and that M2knockout mice of both sexes were significantly more responsive to the mild stress of saline injection than were wild‐type mice. These results accord with the primary function of M2receptors as presynaptic autoreceptors. In the present study, we explored the role of the M1receptor in pituitary–adrenal responses to oxotremorine and saline in male and female M1knockout and wild‐type mice. Because these mice responded differently to the mild stress of saline injection than did the M2knockout and wild‐type mice, we also determined hormone responses to restraint stress in both M1and M2knockout and wild‐type mice. Male and female M1knockout and wild‐type mice were equally unresponsive to the stress of saline injection. Oxotremorine increased both ACTH and corticosterone in M1wild‐type mice to a significantly greater degree than in knockout mice. In both M1knockout and wild‐type animals, ACTH responses were greater in males compared to females, and corticosterone responses were greater in females compared to males. Hormone responses to restraint stress were increased in M2knockout mice and decreased in M1knockout mice compared to their wild‐type counterparts. These findings suggest that M1and M2muscarinic receptor subtypes differentially influence male and female pituitary–adrenal responses to cholinergic stimulation and stress. The decreased pituitary–adrenal sensitivity to oxotremorine and restraint stress noted in M1knockout mice is consistent with M1being primarily a postsynaptic receptor. Conversely, the increased pituitary–adrenal sensitivity to these challenges noted in M2knockout mice is consistent with M2being primarily a presynaptic autoreceptor.