Direct evidence that KNDy neurons maintain gonadotropin pulses and folliculogenesis as the GnRH pulse generator

Abstract

The gonadotropin-releasing hormone (GnRH) pulse is fundamental for mammalian reproduction: GnRH pulse regimens are needed as therapies for infertile women as continuous GnRH treatment paradoxically inhibits gonadotropin release. Circumstantial evidence suggests that the hypothalamic arcuate KNDy neurons expressing kisspeptin (encoded by Kiss1), neurokinin B (encoded by Tac3), and dynorphin A serve as a GnRH pulse generator; however, no direct evidence is currently available. Here, we show that rescuing >20% KNDy neurons by transfecting Kiss1 inside arcuate Tac3 neurons, but not outside of these neurons, recovered folliculogenesis and luteinizing hormone (LH) pulses, an indicator of GnRH pulses, in female global Kiss1 knockout (KO) rats and that >90% conditional arcuate Kiss1 KO in newly generated Kiss1-floxed rats completely suppressed LH pulses. These results first provide direct evidence that KNDy neurons are the GnRH pulse generator, and at least 20% of KNDy neurons are sufficient to maintain folliculogenesis via generating GnRH/gonadotropin pulses.

Role of KNDy Neurons Expressing Kisspeptin, Neurokinin B, and Dynorphin A as a GnRH Pulse Generator Controlling Mammalian Reproduction

“…This is because plasma LH/FSH levels were undetectable in global Kiss1 knockout rats ( 58 ), in which Tac3 and Pdyn gene were abundantly expressed in the ARC as shown in wild-type female rats. Further, Kiss1 rescue into the ARC Tac3 -expressing cells but not out of the Tac3 -expressing cells rescued LH pulses in the global Kiss1 knockout female rats ( 123 ). These findings indicate that ARC neurokinin B/dynorphin A neurons without Kiss1 could not drive GnRH/LH pulse generation.…”

“…To prove that KNDy neurons serve as the GnRH pulse generator, we rescued KNDy neurons by infecting viral vectors expressing Kiss1 mRNA targeted into the ARC Tac3 -expressing neurons in global Kiss1 knockout female rats ( 123 ). Pulsatile LH release was recovered in KNDy-rescued rats in which 20-50% ARC Tac3 -expressing neurons exhibited Kiss1 expression.…”

“…To confirm the notion obtained by the KNDy rescue experiment, we evaluated the effect of conditional ARC Kiss1 knockout on GnRH pulse generation in newly generated Kiss1 -floxed rats. By using the Cre-loxP system, we engineered conditional ARC Kiss1 knockout rats ( 123 ). Pulsatile LH release was completely suppressed in conditional ARC Kiss1 knockout female rats in which >90% Kiss1 -expressing cells disappeared in the ARC.…”

Deletion of Stim1 in hypothalamic arcuate nucleus Kiss1 neurons potentiates synchronous GCaMP activity and protects against diet-induced obesity

“…Stim1 kko mice exhibited more estrous-type vaginal cytology, which may be indicative of higher levels of circulating estrogens due to increased synchronous firing of kisspeptin neurons and excitatory drive to GnRH neurons (Qiu et al, 2016;Clarkson et al, 2017). It is important to note that synchronous firing of "pulse generator" Kiss1 ARH neurons is a failsafe system for maintaining gonadotropin pulses and folliculogenesis in female rodents (Nagae et al, 2021).…”

Reduction of arcuate kappa-opioid receptor-expressing cells increased luteinizing hormone pulse frequency in female rats

“…Loss of function studies of kisspeptin or GPR54 by genetic mutation or conventional knockout showed hypogonadotropic hypogonadism with low to undetectable luteinizing hormone (LH) levels, which clearly demonstrates that kisspeptin-GPR54 signaling is a fundamental factor for stimulating GnRH/LH secretion in rodents and humans [16][17][18]. Kisspeptin neurons located in the hypothalamic arcuate nucleus (ARC) coexpress neurokinin B (NKB) and dynorphin A (Dyn) in rats [19][20][21], mice [22], guinea pigs [23], sheep [24], and goats [25], and are therefore referred to as KNDy neurons [9], and suggested to be the GnRH/LH pulse generator [25][26][27][28][29][30][31].…”