The aim of the present study was to correlate in cardiomyopathic hamsters with congestive heart failure the levels of atrial and ventricular atrial natriuretic factor (ANF) messenger RNA (mRNA) with immunoreactive ANF (IR-ANF) plasma levels and the relative amount of IR-ANF released by the whole heart versus isolated ventricles in the Langendorff preparation. High-performance liquid chromotagraphy analysis of the forms of ANF present in plasma and in the Langendorff effluent of whole heart versus isolated ventricles was also performed. As previously found for cardiac IR-ANF, the levels of ANF mRNA decreased gradually in atria and increased in an analogous fashion in ventricles with the severity of congestive heart failure. Plasma IR-ANF levels (C-terminal) were more elevated in moderate than in severe congestive heart failure, as were the IR-ANF levels in the Langendorff effluent of the whole heart. On the contrary, the effluent of isolated ventricles from animals in severe heart failure yielded more IR-ANF than that from hamsters in moderate heart failure. Thus, while the isolated ventricles from controls contributed 35.8% of IR-ANF released by the whole heart, ventricles from hamsters in moderate heart failure contributed 17.5%, and those from hamsters in severe heart failure contributed 73.9%. These results indicate that atrial cardiocytes contribute more IR-ANF than their ventricular counterpart in moderate heart failure and that ventricles are a major source of plasma IR-ANF in severe heart failure. Analysis of IR-ANF from plasma and the Langendorff efituent from whole hearts and isolated ventricles revealed that the ventricles are the major source of the propeptide (and of its cleaved products) found in the circulation of cardiomyopathic hamsters. These results suggest that ANF synthesis and secretion do not increase conjointly in atria but do increase in ventricles during congestive heart failure.