1. The effects of stress, adrenalectomy and corticosterone treatment on the functional activity of the hypothalamo-pituitary-adrenocortical system have been studied using highly sensitive and precise bio-assay methods for the determination of ACTH and CRH.2. Adrenalectomy resulted in a rise and corticosterone treatment a fall in the hypothalamic CRH content and the plasma ACTH concentration.3. Stress caused a fall followed by a rise in the hypothalamic CRH content, a rise in the pituitary and plasma ACTH concentrations and an increase in the plasma corticosterone concentration.4. The stress-induced changes in hypothalamic CRH content and pituitary and plasma ACTH were exaggerated in adrenalectomized rats but normal in adrenalectomized rats treated with corticosterone. They were also normal in animals injected I.P. with corticosterone 15 min before the stress but inhibited in those similarly treated 60 min previously.5. The ability of adenohypophyses and hypothalami to synthesize and release in vitro ACTH and CRH respectively in response to trophic stimuli was exaggerated in glands removed from adrenalectomized rats and reduced in those removed from corticosterone-treated rats.6. Addition of corticosterone to the pre-incubation medium reduced the capacities of adenohypophyses and hypothalami removed from untreated rats to synthesize and release ACTH and CRH respectively.