Sex differences in hypothalamic astrocyte response to estradiol stimulation.

Background: Reproductive functions controlled by the hypothalamus are highly sexually differentiated. One of the most dramatic differences involves estrogen positive feedback, which leads to ovulation. A crucial feature of this positive feedback is the ability of estradiol to facilitate progesterone synthesis in female hypothalamic astrocytes. Conversely, estradiol fails to elevate hypothalamic progesterone levels in male rodents, which lack the estrogen positive feedback-induced luteinizing hormone (LH) surge. To determine whether hypothalamic astrocytes are sexually differentiated, we examined the cellular responses of female and male astrocytes to estradiol stimulation.

Methods: Primary adult hypothalamic astrocyte cultures were established from wild type rats and mice, estrogen receptor-α knockout (ERKO) mice, and four core genotype (FCG) mice, with the sex determining region of the Y chromosome (Sry) deleted and inserted into an autosome. Astrocytes were analyzed for Sry expression with reverse transcription PCR. Responses to estradiol stimulation were tested by measuring free cytoplasmic calcium concentration ([Ca2+]i) with fluo-4 AM, and progesterone synthesis with column chromatography and radioimmunoassay. Membrane estrogen receptor-α (mERα) levels were examined using surface biotinylation and western blotting.

Results: Estradiol stimulated both [Ca2+]i release and progesterone synthesis in hypothalamic astrocytes from adult female mice. Male astrocytes had a significantly elevated [Ca2+]i response but it was significantly lower than in females, and progesterone synthesis was not enhanced. Surface biotinylation demonstrated mERα in both female and male astrocytes, but only in female astrocytes did estradiol treatment increase insertion of the receptor into the membrane, a necessary step for maximal [Ca2+]i release. Regardless of the chromosomal sex, estradiol facilitated progesterone synthesis in astrocytes from mice with ovaries (XX and XY-), but not in mice with testes (XY-Sry and XXSry).

Conclusions: Astrocytes are sexually differentiated, and in adulthood reflect the actions of sex steroids during development. The response of hypothalamic astrocytes to estradiol stimulation was determined by the presence or absence of ovaries, regardless of chromosomal sex. The trafficking of mERα in female, but not male, astrocytes further suggests that cell signaling mechanisms are sexually differentiated.