Multihormonal bilateral petrosal sinus sampling (BPSS) has been proposed to improve corticotroph microadenomas prediction of lateralisation. Few series have simultaneously compared data of pituitary MRI, surgical findings and multihormonal BPSS. Seven patients (6F/1M) with Cushing's disease, mean age at diagnosis of 35 years (range 24-55) were prospectively studied to compare radiological and multihormonal BPSS data with surgical and pathological findings. In untreated patients, simultaneous measures of ACTH, TSH and prolactine (PRL) were done at time 0, 7, 15, 22 minutes after CRH (500 microgram) and TRH (200 microgram) stimulation. An intersinus gradient of 1.4 was considered as a lateralisation. All microadenomas were identified during surgery, diameters ranged from 2 to 7 mm. All patients were in long-term surgical remission. Pathological studies confirmed a tumoral tissue with ACTH immunostaining in 6/7 cases and PRL in 3/7 cases. Pituitary MRI correctly identified tumors in 4 cases, the remaining tumors were not seen. Basal and stimulated intersinus gradients of ACTH, TSH and PRL were homolateral in 6/7 cases and were coincident with surgical findings in 4/7 cases. The other three cases were contralateral to MRI and surgical data. In conclusion, simultaneous gradient of ACTH, PRL and TSH did not improve lateralisation prediction in this series. Hormonal hypersecretion was homolateral in six cases whereas pathological studies demonstrated a mixed secretion in only three cases. A preferential pituitary draining could explain these discordances. Data from our series and from others (done with CRH stimulation and ACTH-PRL measures) strongly suggest a paracrine interaction between tumoral and normal pituitary tissue.