Insulin-like growth factors (IGFs) are well known as peptide mitogens and important growth factors in fetal as well as in early postnatal development. In particular, IGF II is strongly expressed during fetal life and in neonatal animals. Very little is known about the regulation of IGF II gene expression. In order to study in detail the regulation of IGF II mRNA levels in the liver by the potent nonaromatizable androgen dihydrotestosterone (DHT), we have used quantitative in situ hybridization to detect the mRNA encoding for this growth factor. Pups were separated into 4 groups and injected twice a day immediately after birth with 3 different doses of DHT: 0.1 mg DHT/day, 0.25 mg DHT/day, 0.5 mg DHT/day for 4 and 7 days, and the control groups were injected with the vehicle alone. Animals were perfused with 4% paraformaldehyde and sections from the liver, heart, kidneys and brain were cut with a cryostat. A [35S]-labeled cDNA probe was used to detect IGF II mRNA levels. After hybridization, sections were autoradiographed with X-ray films and then coated with liquid photographic emulsion. Densitometric measurement revealed that, at 4 days of age, IGF II mRNA levels were lower in DHT-treated rats than in control animals. No statistically significant differences in IGF II mRNA levels were observed among the three groups treated with the different doses of DHT, thus revealing that even the lowest dose of DHT (0.1 mg/day) used was sufficient to inhibit IGF II gene expression in neonatal rats. Moreover, at 7 days of age, DHT-treated rats showed the same levels of IGF II mRNA as those observed in rats treated with DHT for 4 days. These results suggest that DHT may play an important role in the regulation of IGF II gene expression in the rat liver during the neonatal period.