Ovarian ageing is a naturally occurring physiological process, marked by dynamic changes in ovarian function and hormone secretion. A key endocrine regulator of ovarian function is the heterodimeric glycoprotein hormone, follicle stimulating hormone (FSH). FSH is secreted as two glycosylation variants: partially glycosylated FSH (FSH21) and fully glycosylated FSH (FSH24). These variants have different in-vitro activities, with FSH21 more bioactive than FSH24. Interestingly, analysis of human pituitary extracts has shown that the ratio of FSH21:FSH24 changes with age, with FSH21 predominant in women of reproductive prime, and FSH24 predominant in menopausal women. However, how differential FSH glycosylation modulates ovarian functions remains unknown. This study therefore aimed to determine the effects of FSH21 and FSH24 on follicle growth and survival. To do this, mouse ovarian follicles were isolated from 3-5wk-old-C57/BL6 mice and treated +/- 10ng/ml, FSH21 (n=85), FSH24 (n=80), a ratio of FSH21:FSH24 at 80:20 (to mimic reproductive prime; n=77), FSH21:FSH24 at 50:50 (n=53), or FSH21:FSH24 at 20:80 (to mimic late peri-menopause; n=78). Follicles were cultured for up to 96hrs and imaged daily to evaluate follicle morphology, and were snap frozen at 24-hour time intervals for qPCR analysis. In the presence of FSH21 dominant conditions, follicle growth was markedly increased at all time points, in comparison to control and FSH24 alone and 20:80 FSH21:FSH24 conditions. Treatment of follicles with FSH24 or 20:80 FSH21:FSH24 resulted in increased basement membrane rupture and oocyte extrusion, with survival rates significantly decreased. qPCR analysis revealed markers of apoptosis were increased in follicles treated with FSH24 alone and 20:80 FSH21:FSH24, while FSH-responsive genes including hormone receptors and steroidogenic enzymes were increased in FSH21 or 80:20 FSH21:FSH24 conditions. These data suggest that the nature of FSH glycosylation modulates the follicular microenvironment to control follicle growth and survival.