Ovarian granulosa cells display strong androgen receptor (AR) expression, suggesting a functional role for direct AR-mediated actions within developing mammalian follicles. By crossing AR-floxed and anti-Müllerian hormone (AMH)-Cre recombinase mice, we generated granulosa cell-specific androgen receptor knockout mice (GCARKO). Cre expression, assessed by lacZ activity, localized to 70%-100% of granulosa cells in most preantral to antral follicles, allowing for selected evaluation of granulosa cell AR-dependent actions during follicle development. Relative to wild-type (WT) females, GCARKO females were subfertile, producing a 24% reduction in the number of litters (P < 0.05) over 6 mo and an age-dependent decrease in total number of pups born, evident from 6 mo of age (P < 0.05). Follicle dynamics were altered in GCARKO ovaries at 3 mo of age, with a significant reduction in large preantral and small antral follicle numbers compared to WT ovaries (P < 0.05). Global premature follicle depletion was not observed, but increased follicular atresia was evident in GCARKO ovaries at 6 mo of age, with an 81% increase in unhealthy follicles and zona pellucida remnants (P < 0.01). Cumulus cell expansion was decreased (P < 0.01) and oocyte viability was diminished in GCARKO females, with a significant reduction in the percentage of oocytes fertilized after natural mating and, thus, in the rate of progression to the two-cell embryo stage (P < 0.05). In addition, compared with age-matched WT females, 6-mo-old GCARKO females exhibited significantly prolonged estrous cycles (P ≤ 0.05), suggesting altered hypothalamic-pituitary-gonadal feedback signaling. In conclusion, our findings revealed that selective loss of granulosa cell AR actions during preantral and antral stages of development leads to a premature reduction in female fecundity through reduced follicle health and oocyte viability.

Homozygous androgen receptor (AR)-knockout (ARKO) female mice are subfertile due to both intra-and extraovarian (neuroendocrine) defects as defined by ovary transplantation. Using ARKO mice, this study set out to reveal the precise ARregulated pathways required for optimal androgen-regulated ovulation and fertility. ARKO females exhibit deficient neuroendocrine negative feedback, with a reduced serum luteinizing hormone (LH) response to ovariectomy (OVX) (P Ͻ 0.01). Positive feedback is also altered as intact ARKO females, at late proestrus, exhibit an often mistimed endogenous ovulatory LH surge. Furthermore, at late proestrus, intact ARKO females display diminished preovulatory serum estradiol (E 2; P Ͻ 0.01) and LH (P Ͻ 0.05) surge levels and reduced Kiss1 mRNA expression in the anteroventral periventricular nucleus (P Ͻ 0.01) compared with controls. However, this reduced ovulatory LH response in intact ARKO females can be rescued by OVX and E 2 priming or treatment with endogenous GnRH. These findings reveal that AR regulates the negative feedback response to E 2, E2-positive feedback is compromised in ARKO mice, and AR-regulated negative and positive steroidal feedback pathways impact on intrahypothalamic control of the kisspeptin/GnRH/LH cascade. In addition, intraovarian AR-regulated pathways controlling antral to preovulatory follicle dynamics are disrupted because adult ARKO ovaries collected at proestrus have small antral follicles with reduced oocyte/follicle diameter ratios (P Ͻ 0.01) and increased proportions of unhealthy large antral follicles (P Ͻ 0.05) compared with controls. As a consequence of aberrant follicular growth patterns, proestrus ARKO ovaries also exhibit fewer preovulatory follicle (P Ͻ 0.05) and corpora lutea numbers (P Ͻ 0.01). However, embryo development to the blastocyst stage is unchanged in ARKO females, and hence, the subfertility is a consequence of reduced ovulations and not altered embryo quality. These findings reveal that the AR has a functional role in neuroendocrine regulation and timing of the ovulatory LH surge as well as antral/ preovulatory follicle development. androgen receptor; ovulation; female reproduction A HEALTHY PREOVULATORY FOLLICLE produces a maturing oocyte and secretes estradiol (E 2 ), leading to a preovulatory surge, which in turn triggers the ovulatory luteinizing hormone (LH) surge and the subsequent follicular rupture and release of the oocyte (18). Throughout most of the murine estrous cycle, E 2 exerts negative feedback on gonadotropin-releasing hormone (GnRH) and LH secretion until the afternoon of proestrus, when there is a switch to positive feedback, with rising follicular E 2 secretion evoking an abrupt release of the preovulatory GnRH surge and then the LH surge that triggers ovulation (22,30). Although follicle development and ovulation are physiologically well-characterized events, the underlying molecular pathways are still being unraveled.The androgen receptor (AR), a member of the nuclear receptor superfamily encoded by the X chromosom...