Activating Transcription Factor 3 Stimulates Follicle-Stimulating Hormone-ß Expression In Vitro But Is Dispensable for Follicle-Stimulating Hormone Production in Murine Gonadotropes In Vivo.

Follicle-stimulating hormone (FSH), a dimeric glycoprotein produced by pituitary gonadotrope cells, regulates spermatogenesis in males and ovarian follicle growth in females. Hypothalamic gonadotropin-releasing hormone (GnRH) stimulates FSHß subunit gene (Fshb) transcription, though the underlying mechanisms are poorly understood. To address this gap in knowledge, we examined changes in pituitary gene expression in GnRH-deficient mice (hpg) treated with a regimen of exogenous GnRH that increases pituitary Fshb but not luteinizing hormone ß (Lhb) messenger RNA levels. Activating transcription factor 3 (Atf3) was among the most upregulated genes. Activating transcription factor 3 (ATF3) can heterodimerize with members of the activator protein 1 family to regulate gene transcription. Co-expression of ATF3 with JunB stimulated murine Fshb, but not Lhb, promoter-reporter activity in homologous LßT2b cells. ATF3 also synergized with a constitutively active activin type I receptor to increase endogenous Fshb expression in these cells. Nevertheless, FSH production was intact in gonadotrope-specific Atf3 knockout [conditional knockout (cKO)] mice. Ovarian follicle development, ovulation, and litter sizes were equivalent between cKOs and controls. Testis weights and sperm counts did not differ between genotypes. Following gonadectomy, increases in LH secretion were enhanced in cKO animals. Though FSH levels did not differ between genotypes, post-gonadectomy increases in pituitary Fshb and gonadotropin α subunit expression were more pronounced in cKO than control mice. These data indicate that ATF3 can selectively stimulate Fshb expression in vitro but is not required for FSH production in vivo.

Strain differences in post-castration sexual and aggressive behavior in male mice.

The degree to which male sexual behavior and territorial aggression are regulated by gonadal steroid hormones depends strongly on species and experience. While castration abolishes male sexual behavior in most laboratory rodents, approximately one third of B6D2F1 mice retain the full repertoire of male sexual behaviors long term ("maters"). It is not yet known whether maters retain other behaviors that typically rely on gonadal steroids to a greater extent than non-maters. In this study, we tested aggressive behavior in B6D2F1 males and males of each parental strain (C57BL/6J and DBA/2J) in the resident intruder paradigm before and after castration, as well as male sexual behavior after castration. Before castration, B6D2F1 residents displayed more attacks compared to DBA/2J males (p < 0.05). There was no difference in attack frequency between B6D2F1 and C57BL/6J males nor between DBA/2J and C57BL/6J males (p > 0.2). A greater proportion of hybrid males demonstrated intromissions and the ejaculatory reflex compared to males of either parental strain (p < 0.01). After castration, B6D2F1 residents attacked more than C57BL/6J males, but not DBA/2 J males (p < 0.05; p > 0.2). There was no difference in post-castration attack frequency between maters and non-maters (p > 0.7). Finally, residents that attacked during all 3 pre-castration resident intruder tests displayed more attacks post-castration than animals that attacked during 1 pre-castration test (p < 0.05). These data suggest that strain and experience influence the expression of aggressive behavior after castration and warrant future study in experience-induced transient increases in extragonadal testosterone.

B6D2F1 mice that retain sexual behavior long term after castration outperform those that cease in the radial arm maze.

In rodents, gonadal steroids play a critical yet variable role in behaviors such as social interaction and cognitive performance. Gonadal steroids organize sex differences observed in spatial working memory, while the absence of activational effects induced by castration generally impedes spatial learning and memory. Although male sexual behavior is typically inhibited following castration, a significant proportion of gonadectomized B6D2F1 hybrid males retains the complete repertoire of male reproductive behavior. In a prior study, amyloid precursor protein and tau, proteins involved in cognitive behavior, facilitated steroid-independent male sex behavior in B6D2F1 hybrid male mice. We used this strain to investigate the relationship between gonadal steroid-independent male sexual behavior and cognition. After identifying "maters" (animals retaining steroid-independent male sex behavior) and "non-maters," we tested spatial memory in an 8-arm radial arm maze. Although neither group demonstrated a decrease in errors as a function of time, maters committed fewer errors compared to non-maters overall (p < 0.05). Maters also completed the maze more quickly than non-maters (p < 0.05). We measured mRNA expression of APP and MAPT as well as LEPR and D2R to probe potential roles of metabolism and motivation. Uniquely among maters, increased relative expression of D2R and LEPR in the hippocampus was associated with a longer latency to complete the maze during the last 3 or across all trials, respectively. These data demonstrate that maters outperform non-maters in the radial arm maze, warranting further study of potential differences in acquisition of spatial memory tasks or learning strategy between these groups.

Peripubertal gonadal steroids are necessary for steroid-independent male sexual behavior in castrated B6D2F1 male mice.

Gonadal steroids play an integral role in male sexual behavior, and in most rodent models, this relationship is tightly coupled. However, many other species, including humans, continue to demonstrate male sex behavior in the absence of gonadal steroids, and the mechanisms that regulate steroid-independent male sex behavior are not well understood. Approximately 30% of castrated male B6D2F1 hybrid mice display male sex behavior many months after castration, allowing for the investigation of individual variation in steroidal regulation of male sex behavior. During both the perinatal and peripubertal periods of development, the organizational effects of gonadal steroids on sexual differentiation of the neural circuits controlling male sex behavior are well-documented. Several factors can alter the normal range of gonadal steroids or their receptors which may lead to the disruption of the normal processes of masculinization and defeminization. It is unknown whether the organizational effects of gonadal hormones during puberty are necessary for steroid-independent male sex behavior. However, gonadal steroids during puberty were not necessary for either testosterone or estradiol to activate male sex behavior in adulthood. Furthermore, activation of male sex behavior was initiated sooner in hybrid male mice castrated prior to puberty that were administered estradiol in adulthood compared to those that were provided testosterone. The underlying mechanisms by which gonadal hormones, during both the perinatal and peripubertal developmental periods of sexual differentiation, organize the normal maturation of neural circuitry that regulates steroid-independent male sex behavior in adult castrated B6D2F1 male mice warrants further investigation.