Long-term space missions expose astronauts to an altered gravitational field (microgravity) that significantly affects human physiology. Although it is critical to assess the effect of microgravity on female reproductive health, this aspect remains a poorly investigated one. To date, interest in the effects on the main ovary hormones (estrogens) has been in depth for its role in bone loss, while the impact of altered estrogen availability on fertility and ovarian function is unclear. The female reproductive apparatus depends on a finely tuned hormonal network involving the hypothalamic-pituitary-ovarian axis. At the heart of this system are the ovarian follicles, composed of granulosa and theca cells, which cooperate in the synthesis of steroid hormones in supporting oocyte maturation. Folliculogenesis and estrogen production are tightly regulated by gonadotropins—FSH and LH—which act on these two distinct cell types. Any disruption in this delicate endocrine cross talk may impair oocyte quality, ovulatory function, and overall reproductive health. New evidence suggests that microgravity may interfere with this delicate hormonal balance, potentially reducing estrogen synthesis and impairing follicular development mostly by downregulating aromatase activity. As the regulation of ovarian function is distributed across different tiers, investigation is particularly delicate. Nevertheless, molecular assessment of relevant parameters can be performed upon cells/tissues cultures growing in real and simulated weightlessness in order to collect data and drawing explicative models.