Knock-out of vasotocin reduces reproductive success in female zebrafish, Danio rerio.

The vertebrate nonapeptide vasotocin/vasopressin is evolutionarily highly conserved and acts as neuromodulator and endocrine/paracrine signaling molecule. Circumstantial and mechanistic evidence from pharmacological manipulations of the vasotocin system in several teleost fishes suggest sex- and species-specific reproductive roles of vasotocin. While effects of vasotocin on teleost reproductive physiology involve both courtship behaviors and the regulation of the hypothalamic-pituitary-gonadal (HPG) axes, comprehensive studies investigating behavioral and physiological reproductive consequences of genetic ablation of vasotocin in a genetically tractable fish model, such as the zebrafish, are currently lacking. Here, we report the generation of homozygous CRISPR/Cas9-based vasotocin gene knock-out zebrafish. Breeding pairs of vasotocin knock-out fish produce significantly fewer fertilized eggs per clutch compared to wildtype fish, an effect coincident with reduced female quivering courtship behavior. Crossbreeding experiments reveal that this reproductive phenotype is entirely female-dependent, as vasotocin-deficient males reproduce normally when paired with female wild-type fish. Histological analyses of vasotocin knock-out ovaries revealed an overall reduction in oocytes and differential distribution of oocyte maturation stages, demonstrating that the reproductive phenotype is linked to oocyte maturation and release. Ovarian hormone quantification and gene expression analysis in mutant fish indicated reduced synthesis of Prostaglandin F2α, a hormone involved in ovarian maturation, egg release and regulation of female courtship behavior in some cyprinids. However, acute injection of vasotocin did not rescue the female mutant reproductive phenotype, suggesting a contribution of organizational effects of vasotocin. Together, this study provides further support for emerging roles of vasotocin in female teleost reproduction in an important teleost model species.

Glucocorticoid receptor activation reduces food intake independent of hyperglycemia in zebrafish.

Chronic cortisol exposure suppresses food intake in fish, but the central mechanism(s) involved in appetite regulation are unclear. Stress and the associated increase in cortisol levels increase hepatic gluconeogenesis, leading to hyperglycemia. As hyperglycemia causes a reduction in food intake, we tested the hypothesis that cortisol-induced hyperglycemia suppresses feeding in zebrafish (Danio rerio). We first established that stress-independent hyperglycemia suppressed food intake, and this corresponded with a reduction in the phosphorylation of the nutrient sensor, AMP-activated protein kinase (AMPK) in the brain. Chronic cortisol exposure also led to hyperglycemia and reduced food intake, but the mechanisms were distinct. In cortisol-exposed fish, there were no changes in brain glucose uptake or AMPK phosphorylation. Also, the phosphorylation of Akt and mTOR was reduced along with an increase in redd1, suggesting an enhanced capacity for proteolysis. Loss of the glucocorticoid receptor did not rescue cortisol-mediated feeding suppression but did increase glucose uptake and abolished the changes seen in mTOR phosphorylation and redd1 transcript abundance. Taken together, our results indicate that GR activation enhances brain proteolysis, and the associated amino acids levels, and not hyperglycemia, maybe a key mediator of the feeding suppression in response to chronic cortisol stimulation in zebrafish.