Department of Biology, University of Ottawa, Ottawa, ON, Canada.
Front Endocrinol (Lausanne). 2023 Aug 21;14:1151299. doi: 10.3389/fendo.2023.1151299. eCollection 2023.
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 F, 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.
脊椎动物非肽血管升压素/加压素在进化上高度保守，作为神经调节剂和内分泌/旁分泌信号分子发挥作用。对几种硬骨鱼的血管升压素系统进行药理学操作的间接和机制证据表明，血管升压素具有性别和物种特异性的生殖作用。虽然血管升压素对硬骨鱼生殖生理学的影响既涉及求偶行为，又涉及下丘脑-垂体-性腺（HPG）轴的调节，但目前缺乏在遗传上可操作的鱼类模型（如斑马鱼）中，全面研究遗传消融血管升压素对行为和生理生殖后果的研究。在这里，我们报告了基于同源重组/CRISPR/Cas9 的血管升压素基因敲除斑马鱼的产生。与野生型鱼类相比，血管升压素敲除鱼的繁殖对产生的受精卵数量明显减少，这与雌性颤抖求偶行为减少有关。杂交实验表明，这种生殖表型完全是雌性依赖性的，因为缺乏血管升压素的雄性与雌性野生型鱼类配对时可以正常繁殖。血管升压素敲除卵巢的组织学分析显示卵母细胞总体减少，卵母细胞成熟阶段的分布存在差异，表明生殖表型与卵母细胞成熟和释放有关。突变鱼的卵巢激素定量和基因表达分析表明，参与卵母细胞成熟、卵子释放以及某些鲤科鱼类雌性求偶行为调节的前列腺素 F 的合成减少。然而，血管升压素的急性注射不能挽救雌性突变体的生殖表型，这表明血管升压素的组织效应有一定的贡献。总之，这项研究为血管升压素在重要硬骨鱼模型物种中的雌性硬骨鱼生殖中的新兴作用提供了进一步的支持。
