In females, germ cells are maintained in ovarian structures called primordial follicles. The number of primordial follicles in the ovarian reserve is a critical determinant of the length of the fertile lifespan. Despite this significance, knowledge of the precise physiological mechanisms that regulate primordial follicle number is lacking. In this study we show that a wave of primordial follicle depletion occurs during the transition to adulthood in mice. We demonstrate that this sudden and dramatic loss of primordial follicles is hormonally triggered and identify the pro-apoptotic BH3-only protein, BCL-2 modifying factor (BMF), as essential for this process, implicating the intrinsic apoptotic pathway as a key mechanism. The elimination of primordial follicles during puberty is not only a striking developmental event, it is also physiologically important because it ultimately reduces the availability of primordial follicles and determines the duration of fertility. Collectively, these findings show that puberty is a critical developmental window for the regulation of the size of ovarian reserve, impacting on female fertility and reproductive longevity.