To examine variations in immunoreactivity of angiotensinogen and cathepsins B and H in hepatocytes over 24 hr, rat liver was examined immunohistochemically. Immunoreactivity of angiotensinogen and cathepsins B and H in periportal and perivenous hepatocytes varied significantly over 24 hr, when analyzed by an image analyzer. In periportal and perivenous hepatocytes, immunoreactivity of angiotensinogen was highest at 0800 hr and lowest at 2000 hr or 0000 hr, whereas that of cathepsins B and H was maximal at 1600 hr and minimal at 0400 hr or 0800 hr. Proteolytic activities of cathepsins B and H in liver extracts varied in parallel to the variations in immunoreactivity of these enzymes. Localization of angiotensinogen in the liver acinus was inversely correlated to that of cathepsins B and H; angiotensinogen was predominantly localized in periportal hepatocytes, but cathepsins B and H were in perivenous hepatocytes at each time point examined. These results suggest that angiotensinogen in hepatocytes is actively synthesized and secreted early in the light period, whereas proteolytic activities in lysosomes of hepatocytes are augmented late in the light period.