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Reproductive Consequences of CRISPR/Cas9-Based avp Knock-Out in Zebrafish (Danio rerio)Ramachandran, Divya 06 December 2022 (has links)
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
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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).
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Mise au point d'une approche de sélection génétique de peptides inhibiteurs d'interactions protéiques fonctionnelles en cellules de mammifèresOstiguy, Alexandre January 2003 (has links)
Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.
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Cooperation in a dynamic social environmentDimitriadou, Sylvia January 2018 (has links)
Cooperative behaviour among unrelated individuals is an evolutionary paradox. Research suggests that an individual’s propensity to cooperate and its response to experiencing cooperation or defection from its social environment consistently varies among individuals and as a function of external factors. The biological and psychological underpinnings of such behavioural variation remain unknown; they can, however, provide more insight into the evolution and maintenance of cooperation among non-kin. This thesis explores the proximate effects of experiences of cooperation or defection from the social environment, as well as possible proximate drivers of cooperative behaviour, using the Trinidadian guppy (Poecilia reticulata) as a study system. Firstly, the behavioural rules underpinning an individual’s decision to cooperate or not with unfamiliar individuals in the presence of specific or non-specific information were explored. When fish had information about their social partner’s cooperativeness, they behaved in a manner consistent with direct reciprocity, copying their partner’s last move. When paired with an ostensibly novel partner, a different, or at least additional, behavioural rule seemed to be employed. In order to help understand the drivers of individual variation in cooperative behaviour, phenotypic selection on cooperativeness was carried out over three filial generations, resulting in fish of high cooperativeness (HC) and low cooperativeness (LC). The divergence of individual cooperativeness observed between the two phenotypic selection lines suggests that cooperative behaviour in the context of predator inspection is at least in part heritable. Cooperative behaviour of F3 fish was found not to correlate with boldness or exploratory behaviour; HC and LC fish did, however, differ in some aspects of sociability and agonistic behaviour. Possible proximate neuromodulatory mechanisms underlying these differences in cooperativeness were also explored, focusing on brain expression patterns for the isotocin receptor (itr) gene in F3 females. HC females were found to have higher mid-section itr expression levels than LC females. Finally, I explored the effects of experiencing cooperation or defection on monoaminergic neurotransmission, which is thought to instantiate the effects of such experiences on the individual’s internal state. My findings suggest that experiencing cooperation or defection from the social environment affects internal state; this phenomenon may be crucial for the appropriate adjustment of the behavioural response to such experiences, and for the emergence of behavioural rules such as generalised reciprocity. Taken together these results suggest that neuromodulatory mechanisms are pivotal for the perception of stimuli from the social environment in the tested cooperative context and that variation in cooperative behaviour may be underpinned by individual differences in the structural properties of such systems. They also provide insight into how behavioural input may affect the behavioural response to such experiences, and ultimately how such mechanisms may lead to the evolution and maintenance of cooperation.
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