Control of Cell Number in the Bed Nucleus of the Stria Terminalis of Mice: Role of Testosterone Metabolites and Estrogen Receptor Subtypes

Abstract

Introduction The bed nucleus of the stria terminalis (BNST) exhibits several sex differences that may be related to male sexual behavior and gender identity. In mice and rats, sex differences in the principal nucleus of the BNST (BNSTp) are due to sexually dimorphic cell death during perinatal life. Although testosterone treatment of newborn female rats increases BNSTp cell number, the relevant hormone metabolite(s) are not known, and the effect of testosterone on the development of BNSTp cell number in mice has not been examined. Aim To identify the sex hormone metabolites and receptors controlling cell number, volume, and cell size in the BNSTp of mice. Methods In the first experiment, C57BL/6J male mice were injected on the day of birth with peanut oil; females were injected with testosterone propionate (TP), estradiol benzoate (EB), dihydrotestosterone propionate (DHTP), or oil alone, and the BNSTp of all animals was examined in adulthood. In the second experiment, to compare effects of EB to the effects of estrogen receptor subtype specific agonists, newborn female mice were injected with EB, propyl-pyrazole-triol (PPT, a selective estrogen receptor alpha [ERα] agonist), or diarylpropionitrile (DPN, a selective estrogen receptor beta [ERβ] agonist). Main Outcome Measures Nuclear volume measurements and stereological cell counts in the BNSTp in adulthood. Results TP treatment of newborn females completely masculinized both BNSTp volume and cell number. EB masculinized neuron number, whereas DHTP had no effect on volume or cell number. In the second experiment, EB again fully masculinized neuron number in the BNSTp and in this study also masculinized BNSTp volume. PPT and DPN each significantly increased cell number, but neither completely mimicked the effects of EB. Conclusions We conclude that estrogenic metabolites of testosterone control sexually dimorphic cell survival in the BNSTp and that activation of both ERα and ERβ may be required for complete masculinization of this brain region.

Effects of Aromatase or Estrogen Receptor Gene Deletion on Masculinization of the Principal Nucleus of the Bed Nucleus of the Stria Terminalis of Mice

“…Also, a study using female mice did not clearly determine whether postnatal EB treatment can masculinize the volume of female BNSTp because it showed that BNSTp volume in adulthood was increased by subcutaneous injection of EB into newborn females, but it also showed no effect of this same treatment [18]. However, this previous report clearly demonstrated that EB treatment could increase the number of BNSTp neurons in female mice [18]. There are several possible hypotheses that could explain why postnatal EB treatment did not restore BNSTp volume in male ArKO mice.…”

“…This finding suggested that estrogen functions during the early postnatal period to maintain neuron number of the BNSTp in male mice. Also, in female mice, postnatal EB treatment has been shown to increase the number of BNSTp neurons [18]. Neurons expressing aromatase are located in the BNSTp of rats during the late prenatal period to the early postnatal period [30].…”

“…Thus, estrogen actions via both ERα and ERβ may be required for masculinization of the BNSTp in mice. However, Hisasue et al [18] also showed that neonatal EB treatment was ineffective in increasing BNSTp volume in female mice. Therefore, it remains unclear whether the estrogen that is synthesized from testicular androgen by aromatase has all the functions necessary for masculinization of the BNSTp in male mice.…”

“…Moreover, in rodents, it is generally accepted that perinatal testicular androgen exerts significant effects on organization of male-typical structures on the brain after it is converted to estrogen in the brain by the cytochrome P450 enzyme aromatase [1,2], although sexual differentiation of the brain is proposed to be partially sex chromosome-dependent [16,17]. Hisasue et al [18] recently reported that treatment with estradiol benzoate (EB) or a selective agonist of estrogen receptor (ER)-α or -β into newborn female mice increases the number of neuron in the BNSTp and enlarges BNSTp volume in adulthood, although activation of either ERα or ERβ is not sufficient for masculinization of the BNSTp. Thus, estrogen actions via both ERα and ERβ may be required for masculinization of the BNSTp in mice.…”

Epigenetics and sex differences in the brain: A genome-wide comparison of histone-3 lysine-4 trimethylation (H3K4me3) in male and female mice

“…Newborn male, female, and androgenized female mice were given saline or valproic acid and effects on the principal nucleus of the BNST (BNSTp) were determined at weaning (Murray et al, 2009). The BNSTp exhibits a number of morphological and neurochemical sex differences in rodents and humans (Guillamon et al, 1988; Hines et al, 1985; Forger et al, 2004; Allen & Gorski, 1990), and in mice these differences can be eliminated by giving females a single injection of testosterone propionate on the day of birth (Hisasue et al, 2010). Valproic acid treatment transiently increased histone acetylation in the brain and prevented masculinization of BNSTp cell number in both males and testosterone-treated females (Murray et al, 2009).…”