Most of the nerve growth factor (NGF) protein in the rat and mouse brain is readily extractable in the presence of guanidine hydrochloride as is the case of brain-derived neurotrophic factor. In the present study, we measured amounts of NGF that could be extracted in the presence and absence of 1 M guanidine hydrochloride from various regions of the brains of male and female mice. About 14% of the total NGF in the hippocampus from female mice at 4 months of age could be extracted without 1 M guanidine hydrochloride (designated loosely bound NGF; about 32% in the rat hippocampus) and the remainder only in its presence (designated tightly bound NGF). The molecular masses of the NGF-immunoreactive protein in both cases were approximately 14 kDa. There were significant differences in respective concentrations of total NGF (the loosely bound plus tightly bound NGF) in the hypothalamus and hypophysis, but not in other brain regions, between male and female mice at 4 months of age. However, levels of loosely bound NGF in the cerebellum and olfactory bulb from males were significantly higher than those in the same regions from females. This difference resulted in two-fold higher ratios of the concentrations of loosely bound to total NGF in males as compared to females. On the other hand, the ratio in the hypophysis was close to unity in both sexes. The concentrations of loosely bound NGF in the hippocampus and cerebral cortex decreased slightly with age in both males and females. Levels of loosely bound NGF increased significantly from 2 to 12 months after birth in the whole brain, olfactory bulb, cerebellum, hypothalamus and hypophysis to a greater extent in males than in females. Thus, it is suggested that high ratios of loosely bound to total NGF in selected regions of brains from male mice are due to an enhanced conversion from tightly to loosely bound form, which is considered to be regulated by androgens (see Brain Res. 322, 112-117, 1990). They may also influence the total NGF expression.