Estrogen receptor-dependent sexual differentiation of dopaminergic neurons in the preoptic region of the mouse

Abstract

Although it has been known for some time that estrogen exerts a profound influence on brain development a definitive demonstration of the role of the classical estrogen receptor (ERalpha) in sexual differentiation has remained elusive. In the present study we used a sexually dimorphic population of dopaminergic neurons in the anteroventral periventricular nucleus of the hypothalamus (AVPV) to test the dependence of sexual differentiation on a functional ERalpha by comparing the number of tyrosine hydroxylase (TH)-immunoreactive neurons in the AVPV of wild-type (WT) mice with that of mice in which the ERalpha had been disrupted by homologous recombination (ERKOalpha). Only a few ERalpha-immunoreactive neurons were detected in the AVPV of ERKOalpha mice, and the number of TH-immunoreactive neurons was three times that of WT mice, suggesting that disruption of the ERalpha gene feminized the number of TH-immunoreactive neurons. In contrast, the AVPV contains the same number of TH-immunoreactive neurons in testicular feminized male mice as in WT males, indicating that sexual differentiation of this population of neurons is not dependent on an intact androgen receptor. The number of TH-immunoreactive neurons in the AVPV of female ERKOalpha mice remained higher than that of WT males, but TH staining appeared to be lower than that of WT females. Thus, the sexual differentiation of dopamine neurons in the AVPV appears to be receptor specific and dependent on the perinatal steroid environment.

Figure 1

ERα expression in the AVPV: Images of immunohistochemically stained sections through the AVPV to illustrate the density and distribution of neurons that express ERα (arrows) in the AVPV of WT (+/+) female (A), WT (+/+) male (B), or ERKOα male (C) mice (×100).

Figure 2

TH expression in the AVPV of the ERKOα mouse. Images of immunohistochemically stained sections through the AVPV to illustrate the density and distribution of neurons that express TH in the AVPV of WT (+/+) female (A), WT (+/+) male (B), ERKOα male (C), or ERKOα female (D) mice. WT females have significantly more TH-immunoreactive neurons relative to that of WT males. Disruption of the ERα in the male (C) prevents the development of this sexually dimorphic pattern of TH expression. Cellular staining is noticeably reduced in the AVPV of the ERKOα female (D), but the number of TH-immunoreactive neurons is similar to that of WT females (see Fig. 3; ×200).

Figure 3

Genetic influences on TH expression in the AVPV. Graphical comparison of the number of TH-immunoreactive neurons detected in the AVPV (unilaterally) of WT and ERKOα mice. (A) The number of TH-immunoreactive neurons in WT (C57BL/6J) male and female mice were compared with that of ERKOα male mice (experiment 1). (B) In a separate experiment. TH staining was compared between WT (+/+) male or female littermates and ERKOα female mice (experiment 2).

Figure 4

(A and B) Development of dopaminergic neurons in Tfm male mice. (C) Images to illustrate the density and distribution of TH-immunoreactive neurons in the AVPV of WT (C57BL/6J) male (A), and Tfm male (B) mice.

Figure 5

Development of PENK mRNA-containing neurons in the AVPV of mice. Graphical comparison of the number of neurons detected in the AVPV (unilaterally) that express PENK mRNA. Neither a sex difference, nor an effect of ERα gene disruption, was observed.