| Popis: |
The nonapeptide family of hormones is deeply conserved in evolution. In teleost fishes, as in all vertebrates, two nonapeptide families exist. These are vasotocin (avp) and oxytocin (oxt). While vasotocin has been shown to regulate individual aspects of reproductive physiology in several teleost species, an integrative assessment of its role on male and female reproduction is currently lacking even in widely used fish models, such as the zebrafish (Danio rerio). Taking advantage of the genetic tractability of the zebrafish, and its emerging status as model to study reproductive physiology, I generated avp -/- mutants using a CRISPR/Cas9 based approach to determine reproductive consequences in female and male zebrafish. Following the identification of a female-specific reproductive phenotype which manifests as a reduction in oocyte release and decreased quivering behaviour, I investigated the potential mechanistic basis at the level of the gonad. In avp -/- ovaries, significantly fewer eggs were present compared to WT fishes. When comparing the distribution of oocyte maturation stages, a significantly lower percentage of stage I and higher percentage of stage V oocytes was present in avp-/- ovaries. The altered distribution in oocyte maturation stages coincided with significant decreases in ovarian transcript abundance of nanos2, a germ-cell specific marker suggesting a possible role for Avp in germ-cell maintenance. Additionally, I observed a decrease in the ovarian concentration of the prostaglandin PGF2, which coincided with a reduction in ovarian transcript abundance of pla2g4ab, a paralogue of the phospholipase A2 involved in mobilizing arachidonic acid, a precursor of PGF2,. Together, these finding suggests a role for Avp in PGF2 -mediated ovulation. Because Avp has pleiotropic effects and may thus affect female reproductive physiology indirectly, we assessed somatic growth, a key regulator of sexual maturation in zebrafish, as well as aspects of the endocrine stress axis known to affect oocyte growth in avp -/- mutants. While avp -/- mutants did not exhibit differences in somatic growth up to sexual maturation or GSI, mutants exhibited hypercortisolism. While other zebrafish knock-out mutants exhibiting persistent hypercortisolism do not share the observed reproductive phenotype, future studies investigating potential contributions of pleiotropic Avp effects are nevertheless warranted. Overall, I demonstrate that avp, while not essential, affects female reproductive success, at least iii in part by regulating oocyte maturation. This finding is in line with the recent findings from other vertebrate and invertebrate species, suggesting an evolutionarily ancient role in these processes. It is anticipated that such novel insights into the regulation of female oocyte maturation have in addition to increasing our understanding of female reproduction, translational potential for captive breeding (aquaculture, species conservation) and ecotoxicology (insight into mode of action of specific EDCs). |
