PROJECT SUMMARY We have identified FSH as an actionable target for osteoporosis, obesity and Alzheimer's disease (AD)— diseases that affect millions of men and women worldwide [e.g. Cell, 2006, PMID: 16630814; Nature, 2017, PMID: 28538730; Nature, 2022, PMID: 35236988]1-8. SWAN (NIA) shows that these diseases track together during the menopausal transition when FSH rises while estrogen remains unperturbed—implicating FSH as a disease driver9-13. FSH blockade in mouse models of osteoporosis, obesity and AD prevents disease onset4- 6,8,14. We thus developed a lead therapeutic—MS-Hu6—which is currently undergoing preclinical evaluation [PNAS, 2020, PMD: 33127753; eLife, 2022, PMID: 36125123]2,15. Project 1 will focus on testing the efficacy of MS-Hu6 in models of established disease toward future clinical studies. Mechanistically, however, several issues arise from prior studies, which will be the focus of Project 2. First, while we and others have established FSH receptor (FSHR) expression in bone cells4-6,16, we are unclear which of the three cell types—osteoblasts, osteocytes or osteoclasts—mediate FSH action on bone mass. While global FSHR haploinsufficiency results in impaired osteoclastic bone resorption6, FSH–blocking antibodies also unmask potent bone anabolic effects [PNAS, 2012, PMID: 22908268; PNAS, 2018, PMID: 29440419]2,4,14. In wild type mice, where resorption is not ordinarily elevated, MS-Hu6 increases osteoblastic bone formation2. Furthermore, prior studies were performed in young mice, leaving open a further question on how bone responds to FSH blockade over the lifetime of a mouse—this is particularly relevant to older women, where we find that FSH levels correlate with bone mineral density and fracture risk [JCEM, 2021, PMID: 33326040]17. Thus, in Specific Aim 1, to determine which bone cell is the primary FSH target, we will delete FSHRs selectively in osteoblasts, osteocytes or osteoclasts by crossing Fshrfl/fl mice with tamoxifen–inducible Cre lines. We will then phenotype the skeletons of the mutants over their lifetime and following gonadectomy. We have also shown that FSH is a pro–obesity hormone and that genetic or pharmacologic ablation induces a lean, thermogenic phenotype5. While FSHRs are expressed in adipocytes5, we also find expression in hypothalamic and hindbrain nuclei that transmit sympathetic signals peripherally to adipose tissue [eLife, 22022, PMID: 36052994]18. It is therefore imperative that we understand the contribution of peripheral versus central effects of FSH on body composition. In Specific Aim 2, we will delete Fshr selectively in adipocytes by crossing Fshrfl/fl mice with doxycycline–inducible Adipoq-CrertTA,tetO mice, and use UCP1–reporter ThermoMice to study beiging. In parallel, we will inject siFshr stereotactically into FSHR–rich hypothalamic nuclei18. Following Fshr deletion, mutant mice will be put on a high–diet and undergo detailed body composition and metabolic phenotyping. In all, these ...