Expression of the c-Ha-ras proto-oncogene is specifically enhanced during liver regeneration, in parallel with increased DNA replication, which suggests that c-Ha-ras may play a role in the control of regeneration. In this study, an in situ hybridization technique was applied for analysis of expression of the c-Ha-ras gene at the cellular level during liver regeneration induced by CCl4 administration. The in situ hybridization was compared with the observation for the p21c-Ha-ras protein, the corresponding protein of the c-Ha-ras gene, by immunohistochemistry. In normal rat liver, a few hepatocytes expressed the mRNAs and the corresponding proteins without any preferential localization. Zonal heterogeneity of c-Ha-ras gene expression first became evident at 12 hr after CCl4 administration, a higher number of gene products being detected in the pericentral zone than in the periportal zone. This heterogeneity became maximal at 24 hr after CCl4 administration. Zonal heterogeneity in the level of the p21c-Ha-ras protein paralleled that in the level of gene expression. Furthermore, both hepatocytes and nonparenchymal cells participated in expression of the c-Ha-ras gene during liver regeneration.