Chemogenetic Depletion of Hypophysiotropic GnRH Neurons Does Not Affect Fertility in Mature Female Zebrafish

Abstract

The hypophysiotropic gonadotropin-releasing hormone (GnRH) and its neurons are crucial for vertebrate reproduction, primarily in regulating luteinizing hormone (LH) secretion and ovulation. However, in zebrafish, which lack GnRH1, and instead possess GnRH3 as the hypophysiotropic form, GnRH3 gene knockout did not affect reproduction. However, early-stage ablation of all GnRH3 neurons causes infertility in females, implicating GnRH3 neurons, rather than GnRH3 peptides in female reproduction. To determine the role of GnRH3 neurons in the reproduction of adult females, a Tg(gnrh3:Gal4ff; UAS:nfsb-mCherry) line was generated to facilitate a chemogenetic conditional ablation of GnRH3 neurons. Following ablation, there was a reduction of preoptic area GnRH3 neurons by an average of 85.3%, which was associated with reduced pituitary projections and gnrh3 mRNA levels. However, plasma LH levels were unaffected, and the ablated females displayed normal reproductive capacity. There was no correlation between the number of remaining GnRH3 neurons and reproductive performance. Though it is possible that the few remaining GnRH3 neurons can still induce an LH surge, our findings are consistent with the idea that GnRH and its neurons are likely dispensable for LH surge in zebrafish. Altogether, our results resurrected questions regarding the functional homology of the hypophysiotropic GnRH1 and GnRH3 in controlling ovulation.

Cholecystokinin gates reproduction in zebrafish by controlling gonadotropin secretion

“…Our results indicate that while fish gonadotrophs were segregated into two different populations and placed under the control of two distinct neuropeptides during evolution, a common hypothalamic pathway gates the secretion of both gonadotropins. Interestingly, in contrast to GnRH, the novel CCK regulation identified under the current study has a more substantial effect on the gonadotrophic axis, as revealed in our mutants, while the disruption of GnRH and its receptors didn’t lead to any drastic effect on reproduction ( 24, 40 ).…”

“…The stimulus was applied manually as a pulse of 300 μl of 30 μg/μl peptide during the first 30 seconds of the imaging session. Hormones were measured in the fractions before and after stimulation using ELISA developed for common carp, which was established in the Levavi-Sivan lab using recombinant carp gonadotropins produced in yeast, and had been previously shown to be suitable for zebrafish (24). Tissue viability was validated by monitoring the morphology and activity of the cells, looking at granulation and calcium changes.…”

“…In some species, such as the zebrafish, that express only two forms of GnRH (GnRH2 and GnRH3), the gene encoding GnRH1 has been lost, and GnRH3 has become the dominant hypophysiotropic form (23). For a yet unknown reason, in the zebrafish, even a complete absence of GnRH does not impair ovulation, as adult zebrafish with loss of function (LOF) of GnRH signaling are fertile (24)(25)(26), suggesting that GnRH activity is either replaced by a compensation mechanism or it is dispensable for the control of gonadotropin release overall. In other species, such as medaka, the effects of GnRH are limited to the control of final oocyte maturation and ovulation via LH secretion (2).…”

Hormonal dynamics reveal a stimulatory role for secretoneurin in zebrafish ovulation

“…Therefore a multitude of hypothalamic neurons, including those that produce the fish Gnrh and Kiss peptides, terminate near, or directly innervate, the distinct gonadotroph cells releasing Lh or Fsh (9), thus conferring control over them without a tetrapodlike median eminence-portal blood system. In zebrafish, Gnrh2/Gnrh3 single or double mutants (10,11) and those with chemogenic depletion of Gnrh3 are fertile (12). Medaka harboring mutations in gnrh1 have well-developed ovaries but fail to ovulate.…”