Disappearance of Sex Difference in the Anxiety-Like Behavior in Progranulin-Deficient Mice.

Abstract

It is well established that the mammalian brain possesses sexually dimorphic structure, which is reflected by functional sexual differences in the control of specific types of behavior. Sexual differentiation of the brain in rodents is achieved by estrogens, which are converted from androgens in the brain, during the perinatal period. We have identified the progranulin (PGRN) gene as one of the sex steroid-inducible genes in the hypothalamus of neonatal rats, and suggested that PGRN and/or PGRN-derived peptides (granulins) are involved in the organization of neuronal circuits responsible for the expression of male sexual behavior. We have further generated a line of mice with targeted disruption of the PGRN gene, and observed a decrease in ejaculation incidence and an increase in aggression. Recently, we also suggested the involvement of PGRN in estrogen-induced neurogenesis in adult rat hippocampus. To further clarify the role of PGRN in the sexual differentiation of the brain, we analyzed the anxiety-like behavior and anxiety-related brain nuclei using PGRN-deficient mice. We used 7-week old male and female wild type (Grn+/+) and PGRN-deficient (Grn-/-) mice of C57BL/6J background. To evaluate the anxiety-like behavior, open field test was performed. The open field was composed of 50 x 50 cm square surrounded by 40 cm-height walls. Each mouse was introduced in the center of the square and its behavior was recorded on videotape for 10 min. The duration when the mouse stayed in the peripheral zone (7.5 cm from the wall) or the center, and the distance traveled were analyzed using a computerized technique with Ethovision 3.0 software. Then, 50-µm thick sections were made from 4% paraformaldehyde-fixed brain and stained with 0.1% cresyl violet. The volume and number of cells in the locus ceruleus (LC) and paraventricular nucleus (PVN), both of which are known to be involved in exhibiting anxiety-like behavior, were determined from the digital images of sections using Image J software. Male Grn+/+ mice spent significantly longer time in the center of open filed than female Grn+/+ mice, suggesting that the anxiety is lower in males than in females. Male Grn-/- mice spent significantly shorter time in the center of open field than male Grn+/+ mice, and the duration staying in the center was not different between male and female Grn-/- mice. Distances traveled in both areas were not significantly different between the sexes nor genotypes. Histological analysis revealed that the volume and the number of cells in the LC of Grn-/- mice were significantly larger than those of Grn+/+ mice. On the other hand, there were no significant differences in the volume and cell numbers in the PVN between the sexes nor genotypes. These results suggest that there are sex differences in anxiety to the novel environment in wild type mice of the strain used in the present study, i.e., females exhibit enhanced anxiety than males, and that PGRN plays an important role in the formation of this sex difference. It is probable that PGRN produces the sex difference in anxiety by affecting the organization of neuronal structure in the LC. Given that PGRN is involved in estrogen-induced neurogenesis, PGRN may suppress proliferation and/or the survival of neural cells in the LC during the perinatal period, thereby affects the behaviors such as aggression and anxiety in adulthood.