Rat brain natriuretic peptide (BNP) was detected by radioimmunoassay in heart atria and ventricles and in plasma. We have investigated its localization in atria and the possibility of cosecretion of atrial natriuretic factor (ANF) and BNP into the circulation. BNP was detected by chromatographic analysis and immunoblotting in the isolated atrial granules together with ANF: It consisted of two immunoreactive proteins of 14,000 and 2,500 apparent molecular weight. By immunohistochemical methods, BNP was particularly found in the perinuclear region of atrial cardiocytes. Double-labeling immunocytochemical methods colocalized BNP and ANF in the same atrial secretory granules. Basal plasma BNP levels ranged from 2.6 to 4.4 fmol/ml. After stimuli by morphine injection or an aortocaval shunt, BNP levels increased by 4- and 7-fold, respectively, whereas ANF levels rose by 50- and 6-fold, respectively. Depending on the stimulus, BNP release into the circulation is not necessarily proportional to ANF, indicating that BNP may originate not only from the atrial granules but also from other tissues such as the ventricles. These results suggest that BNP may participate with ANF in blood pressure control and salt and water homeostasis.