Bilateral simultaneous inferior petrosal sinus sampling, associated with the oCRH stimulation test (100 micrograms i.v. as a bolus) was performed in 22 patients with Cushing's syndrome and no signs of pituitary abnormalities. Catheters were inserted into both femoral veins. More than one site in the superior and inferior vena cava was sampled before reaching the inferior petrosal sinuses. Blood samples for ACTH and beta-endorphin were gently aspirated from both petrosal sinuses and from a peripheral vein simultaneously. Blood was drawn at 0, 5, 10 and 15 min after oCRH injection. Seventeen of 22 patients showed an ipsilateral to peripheral vein ratio higher than 1.5, and 12 patients showed a lateralization of ACTH levels after oCRH stimulation. Seventeen patients underwent transsphenoidal pituitary surgery. Nine patients had a pituitary adenoma at the expected side; 1 at the contralateral side, while in 2 it was central. Three of 4 patients in whom the ipsilateral/peripheral ratio was less than 1.5 had the highest ACTH levels at the superior or inferior vena cava, not responsive to oCRH stimulation. One of these had a mediastinal and one a pulmonary mass. The third one, with an occult ectopic source, is still under investigation. At immunohistochemical and biological in vitro studies, both tumors were shown to secrete ACTH. In 13 patients in whom both beta-endorphin and ACTH measurements were performed, these hormones showed similar patterns of response. In conclusion, simultaneous bilateral petrosal sinus catheterization is a useful tool in the differential diagnosis of Cushing's syndrome as concerning pituitary and ectopic forms.(ABSTRACT TRUNCATED AT 250 WORDS)