The cardiac and plasma levels of immunoreactive (IR-) atrial natriuretic factor (ANF) in cardiomyopathic hamsters with moderate and severe congestive heart failure were measured, compared with those of controls, and correlated by HPLC analysis of IR-ANF in atria, ventricles, and plasma and with the ultrastructure of atrial and ventricular cells. Congestive heart failure in the hamster produced a significant increase in plasma IR-ANF, a significant decrease in atrial IR-ANF, and a marked increase in ventricular IR-ANF. The HPLC pattern of IR-ANF was of the high mol wt type in atria and ventricles of control and cardiomyopathic animals. High mol wt forms of ANF appeared in the plasma of animals with severe congestive heart failure, but not in controls. Severe congestive heart failure produced a tremendous increase in the size of the Golgi complex, with a decrease in the number and size of secretory granules in atrial cardiocytes. Ventricular cardiocytes also showed a less marked increase in the size of the Golgi complex. Secretory-like granules indistinguishable from lysosomes were present in about 1% of ventricular cardiocytes of control hamsters; in hamsters with severe congestive heart failure, secretory granules, identical to those of atrial cardiocytes, were present in greater number in about 20% of ventricular cardiocytes. Immunocytochemistry (immunogold technique) revealed that secretory granules containing IR-ANF are not present in control ventricular cardiocytes but are localized in relatively large number in about 20% of ventricular cardiocytes in hamsters with severe congestive heart failure. These results suggest that the increased IR-ANF levels (including the high mol wt forms) in animals with congestive heart failure may come from hypersecretion of both atrial and ventricular cardiocytes.