Daily-, estral- and age-dependent regulation of RFRP-3 neurons and their role in luteinizing hormone secretion in female mice

Female reproductive success relies on proper integration of circadian- and ovarian- signals to the hypothalamic-pituitary-gonadal axis in order to synchronize the pre-ovulatory LH surge with the onset of the main active period. In this study, we assessed in female mice whether hypothalamic neurons expressing the gonadotropin inhibitory peptide, RFRP-3, exhibit daily-, estrous stage-, and age-dependent variations. Furthermore, we investigated whether arginine vasopressin (AVP) and vasoactive intestinal peptide (VIP), two circadian peptides produced by the suprachiasmatic nucleus, regulate RFRP-3 neuronal activity. In young mice, the number of activated (c-Fos–positive) RFRP-3 neurons was reduced at the day-to-night transition with no difference between diestrus and proestrus. Contrastingly, RFRP neuron firing rate was higher in proestrus as compared to diestrus, independently of the time of the day. AVP fibers contacted RFRP neurons with the highest density observed during the late afternoon of diestrus and proestrus, while AVP application increased RFRP neuron firing in the afternoon of diestrus, but not of proestrus. By contrast, we found no daily variation and no effect of VIP on RFRP neurons. Moreover, we demonstrated an age-dependent decrease in the total and activated number of RFRP-3 neurons. Finally, we reported that the daily variations in the activated RFRP-3 neurons and number of RFRP-3 neurons with close AVP and VIP fiber appositions were abolished in old mice. In conclusion, RFRP neurons integrate both daily and estrogenic signals in young, but not in old mice, a phenomenon which could be implicated in the aging of the reproductive axis.