The mechanism by which sex steroids influence very low density hepatic lipoprotein triglyceride production has not been fully elucidated. In previous studies we showed that [(14)C]oleate utilization and incorporation into triglycerides were greater in hepatocyte suspensions from adult female rats than from males. The sex differences were not related to activities of the enzymes of triglyceride biosynthesis, whereas fatty acid binding protein (FABP) concentration in liver cytosol was greater in females. These findings suggested that sex differences in lipoprotein could reflect a sex steroid influence on the availability of fatty acids for hepatocellular triglyceride biosynthesis. In the present studies, sex steroid effects on hepatocyte [(14)C]oleate utilization and FABP concentration were investigated directly. Hepatocytes from immature (30-d-old) rats exhibited no sex differences in [(14)C]oleate utilization. With maturation, total [(14)C]oleate utilization and triglyceride biosynthesis increased moderately in female cells and decreased markedly in male cells; the profound sex differences in adults were maximal by age 60 d. Fatty acid oxidation was little affected. Rats were castrated at age 30 d, and received estradiol, testosterone, or no hormone until age 60 d, when hepatocyte [(14)C]oleate utilization was studied. Castration virtually eliminated maturational changes and blunted the sex differences in adults. Estradiol or testosterone largely reproduced the appropriate adult pattern of [(14)C]oleate utilization regardless of the genotypic sex of the treated animal. In immature females and males, total cytosolic FABP concentrations were similar. In 60-d-old animals, there was a striking correlation among all groups (females, males, castrates, and hormone-treated) between mean cytosolic FABP concentration on the one hand, and mean total [(14)C]oleate utilization (r = 0.91) and incorporation into triglycerides (r = 0.94) on the other. In 30-d-old animals rates of [(14)C]oleate utilization were greater, relative to FABP concentrations, than in 60-d-old animals. The sex differences that characterize fatty acid utilization in adult rat hepatocytes are not present in cells from immature animals, and reflect in part the influence of sex steroids. It remains to be determined whether the observed relationship of hepatic FABP concentration to [(14)C]oleate utilization in adult cells is causal or secondary to changes in cellular fatty acid uptake effected through another mechanism. In either case, modulation of triglyceride-rich lipoprotein production by six steroids appears to be mediated to a significant extent by their effects on hepatic fatty acid utilization.