Mice were fed a lithogenic diet containing 1% cholesterol and 0.5% cholic acid, and the sequence of morphologic changes in the gallbladder was followed. Histologic changes were studied with light microscopy and cell proliferation with autoradiography. Mucus glycoprotein production was studied with a spectrum of glycoprotein histochemical stains and electron-microscopic morphometry of membrane-bound mucus secretory granules. There was a rapid and early increase in cell proliferation of the epithelial cells and an increase in mucus production, which took place long before any observable cholesterol crystal precipitation. The gallbladder epithelial cells showed early focal pseudostratification and large numbers of degenerative cells. These changes culminated in glandular metaplasia and the formation of Rokitansky-Aschoff sinuses later in the gallstone induction phase. These observations clearly indicated that gallbladder epithelial injury had taken place before gallstone formation. The authors hypothesize that the consequences of these cellular changes play a contributory role in causing gallstone formation.