Development and validation of a method for precise dating of female puberty in laboratory rodents: The puberty ovarian maturation score (Pub-Score)

Abstract

Puberty is a key developmental event whose primary regulatory mechanisms remain poorly understood. Precise dating of puberty is crucial for experimental (preclinical) studies on its complex neuroendocrine controlling networks. In female laboratory rodents, external signs of puberty, such as vaginal opening (VO) and epithelial cell cornification (i.e., first vaginal estrus, FE), are indirectly related to the maturational state of the ovary and first ovulation, which is the unequivocal marker of puberty. Whereas in rats, VO and FE are almost simultaneous with the first ovulation, these events are not so closely associated in mice. Moreover, external signs of puberty can be uncoupled with first ovulation in both species under certain experimental conditions. We propose herein the Pubertal Ovarian Maturation Score (Pub-score), as novel, reliable method to assess peripubertal ovarian maturation in rats and mice. This method is founded on histological evaluation of pre-pubertal ovarian maturation, based on antral follicle development, and the precise timing of first ovulation, by retrospective dating of maturational and regressive changes in corpora lutea. This approach allows exact timing of puberty within a time-window of at least two weeks after VO in both species, thus facilitating the identification and precise dating of advanced or delayed puberty under various experimental conditions.

Metabolic regulation of female puberty via hypothalamic AMPK–kisspeptin signaling

“…Of note, documentation of the delay in puberty onset was achieved by a combination of phenotypic and hormonal markers that, admittedly, did not include first estrus. This was mainly due to the fact that the delay of vaginal opening, as a robust and reliable sign of puberty onset in rodents (37), which was associated with activation of AMPK, caused a substantial number of animals to not undergo complete canalization of the vagina by the time of termination of some experiments. In any event, our findings disclose a pubertal dimension of brain AMPK signaling, which is compatible with previous fragmentary evidence suggesting that activation of AMPK might repress adult reproductive function, eventually via inhibition of key components of the reproductive brain (e.g., GnRH neurons) in adverse metabolic conditions, such as extreme glucoprivation, as suggested by electrophysiological studies (24).…”

SIRT1 mediates obesity- and nutrient-dependent perturbation of pubertal timing by epigenetically controlling Kiss1 expression

“…Somatic and reproductive indices of pubertal development were evaluated as previously described 50 , including (a) body weight (BW) gain; (b) age of vaginal opening (VO), a consensus external marker of puberty in female rodents; (c) uterine and ovarian weight; and (d) serum LH and FSH levels. Based on previous data on the normal timing of puberty in female rodents 50 – 52 , VO was monitored daily from PND25 until termination of the experiment, time at which uterine and ovarian weight were recorded and serum hormone levels were assayed. Further assessment of the precise age of completion of puberty was achieved by histological analysis of the ovary, as described below.…”

“…Serial (7 μm) sections were stained with hematoxylin and eosin, and evaluated under a microscope. Pubertal progression was estimated using a scoring method (Pub-Score), recently validated by one of our groups, based on histometric analyses of follicular development and development of corporal lutea 52 . This method dates pubertal maturation based on the combined analysis of follicular development and corpus luteum dynamics (the latter, for animals that has completed first ovulation).…”

Developmental decrease in parvalbumin‐positive neurons precedes increase in flurothyl‐induced seizure susceptibility in the Brd2+/− mouse model of juvenile myoclonic epilepsy

“…Developmentally, in Brd2 +/‐ mice, there was an increase in seizure susceptibility just before puberty (P30), 29 suggesting possible effects of the changing sex steroid levels occurring before and around puberty. For example, testosterone (in males) surges around P25 30 and might play a role in seizure susceptibility (eg, it may represent a trigger factor in the Brd2 +/‐ genotype).…”