In 29 consecutive patients with adrenocorticotropin (ACTH)-dependent Cushing's syndrome, we compared the usefulness of multiple baseline ACTH evaluations (10/29), multiple hormone evaluation (29/29) and ACTH-releasing hormone (CRH) stimulation (21/29) during simultaneous and bilateral inferior petrosal sinus sampling. The basal inferior petrosal sinus/periphery ratio for ACTH concentrations was greater than 2 in 18 of the 29 patients and CRH challenge caused the appearance of an inferior petrosal sinus/periphery ratio greater than 3 in 6 other patients. The presence of an ACTH-secreting adenoma was surgically proven in all the 24 patients who had an ACTH inferior petrosal sinus/periphery ratio greater than 2 basally or greater than 3 after the CRH test but also in 1 patient who had an inferior petrosal sinus/periphery ratio lower than 2 basally or 3 after the CRH test. In 4 patients, both the very high peripheral ACTH levels, the inferior petrosal sinus/periphery ratio and the complete lack of ACTH increase after CRH indicated the presence of an ectopic ACTH syndrome: a bronchial carcinoid was found in 2 patients, whereas the site of the tumor is still unknown in the remaining 2. An ACTH intersinus gradient greater than 1.4 was found in 23 patients. Among these 23 patients, the side of the adenoma was correctly predicted in 19 patients and wrongly in 4.(ABSTRACT TRUNCATED AT 250 WORDS)