To examine the role of interleukin-1 (IL-1) in the regulation of the hypothalamo-pituitary-adrenal (HPA) axis, mice with knockout of the IL-1 receptor type I (IL-1rKO) were exposed to psychological and metabolic stressors. When exposed to mild stressors (auditory stress or a low dose of 2-deoxyglucose), IL-1rKO mice displayed a significantly diminished corticosterone secretion, compared with wild-type (WT) controls. In response to more severe stressors (60-min restraint or a high dose of 2-deoxyglucose), both groups exhibited a similar increase in corticosterone secretion. To examine the role of IL-1 in HPA axis feedback regulation, serum ACTH levels were measured following adrenalectomy (ADX) in IL-1rKO mice and in mice with transgenic over-expression of IL-1 receptor antagonist within the brain (IL-1raTG). As expected, WT controls exhibited ADX-induced ACTH hyper-secretion, whereas in IL-1rKO and IL-1raTG mice showed no increase in ACTH levels, suggesting that brain IL-1 has a critical role in ADX-associated ACTH hyper-secretion. Similarly, WT mice that were chronically exposed to IL-1ra in utero displayed a diminished ADX-induced ACTH hyper-secretion, compared with vehicle-treated controls, suggesting a developmental role of IL-1 in HPA axis regulation. In conclusion, our results suggest that endogenous IL-1 plays a critical role in HPA axis activation following stress and adrenalectomy.