Age-related changes in the biological activity of osteoblastic cells have been studied extensively using histomorphometry, especially in relation to osteoporosis. Nevertheless, the changes occurring in the biological activity of individual osteoblastic cells are not sufficiently clarified by this technique. In the present study, age-related changes in the expression of bone matrix protein mRNAs in individual osteoblastic cells were analyzed in vivo by in situ hybridization using undecalcified bone sections. In the femurs of 8-week-old male rats, strong expression of type I collagen and osteocalcin mRNAs was detected in cuboidal osteoblasts on the bone formation surface. Osteopontin mRNA was detected in some of the mononuclear cells and osteoclasts on the bone resorption surface, and also in osteocytes. In the femurs of 60-week-old and 100-week-old male rats, expression of these bone matrix protein mRNAs was markedly decreased. Histomorphometrical analysis of 8-week-old and 60-week-old rats indicated that both the activity and number of osteoblasts expressing type I collagen mRNA, as well as the number of osteoclasts, were reduced in these tissues in older animals. These results indicate age-related reductions in both biological activity and numbers of osteoblasts.