Two distinct nonapeptide systems, consisting of the vasotocin- and oxytocin-related peptides have evolved in vertebrates, and their role in male reproduction is well-described in mammals. In contrast, their comparative role in reproduction in basal vertebrate species, and teleost fishes in particular, has not been investigated in great detail. Using two cyprinid species, the zebrafish (D. rerio) and the goldfish (C. auratus), I address the hypothesis that the teleost nonapeptides vasotocin and isotocin stimulate male cyprinid reproductive physiology by affecting central neuronal and/or peripheral endocrine pathways. To test this hypothesis in zebrafish, an indeterminate breeder, I conducted pharmacological inhibition experiments employing vasotocin and isotocin-specific antagonists in males, a treatment predicted to inhibit reproductive success in mating trials. Because nonapeptides can act both as central peptide neuromodulators and as secreted hormone, I further quantified indices of male courtship behavior (nudging, circling and chasing) and major androgens (testosterone and 11-keto-testosterone) as key endocrine indices of the male reproductive axis. Together, these experiments revealed a dose-dependent, differential inhibition of spawning success, with significant reductions (-65%) in egg fertilization rate observed in pairs in which males had been i.p. injected with 5 ng/g vasotocin and significant reductions (-79%) observed at 500 ng/g i.p injected isotocin. In either case, these partial inhibitions of reproductive success were correlated with significant decreases in specific indices of male courtship behavior, but not endocrine indices, suggesting that individual nonapeptides mediate their effects via central modulation of behavioural neurocircuits. Interestingly, a co-administration of vasotocin and isotocin antagonists completely abolished reproductive success, however this effect was neither correlated with decreases in male courtship behavior, nor endocrine indices, suggesting a separate mode of action, possibly at the level of male pheromone release. To further probe the role of nonapeptides in male zebrafish reproduction, I subsequently tested the hypothesis that nonapeptide systems are acutely activated by key reproductive cues, specifically the releaser pheromone PGF2α, which serves as a chemoattractant and acutely stimulates male reproductive behavior in male cyprinids. Using a chemoattractant choice assay in conjunction with immunohistochemistry and gene expression approaches, I determined whether male zebrafish are attracted to pheromonal cues and acutely activate isotocinergic neurons in the short term and/or regulate nonapeptide gene expression in the longer term. My results show that individual male zebrafish are attracted to PGF2α in an acute choice test. Furthermore, an increase in p-ERK immunoreactivity, a marker of neuronal activation, was observed in the olfactory bulb 10 min following exposure, suggesting a specific response to the pheromone compared to EtOH vehicle. However, no co-localization of p-ERK and IT-positive perikarya was observed in the preoptic area (POA), refuting the hypothesis that PGF2α exposure acutely activates isotocinergic neurons in zebrafish. Analysis of whole brain relative mRNA transcript abundance revealed that PGF2α exposure time-dependently regulates whole brain isotocin, but not vasotocin transcript abundance, suggesting secondary longer-term effects of PGF2α exposure on the isotocinergic system. Using an analogous experimental approach, I further tested the hypothesis that nonapeptides stimulate male reproductive physiology in goldfish, a determinate breeder. Sexually mature male goldfish pretreated with saline or vasotocin or isotocin antagonists were exposed to saline or PGF2α-injected stimulus females and male courtship behavior (chasing, circling), endocrine indices (circulating testosterone) and milt release were quantified. Both nonapeptide antagonists reduced strippable male milt quantity in response to PGF2α-injected females, suggesting a neuronal or hormonal action of both nonapeptides on goldfish milt release. Together, I show that nonapeptides contribute to male reproductive physiology in two species of cyprinids with different reproductive tactics. However, the mode of action may differ from one species to another, with evidence suggesting that nonapeptides play a role in the regulation of reproductive behavior and, possibly, male pheromone, release in zebrafish, while effects on male goldfish seem to be exclusively related to the release of milt. Future studies should compare other teleost species with specific reproductive biology and focus on the gonadal roles of nonapeptides in sperm maturation and/or release.

The ability of an animal to respond and adapt to stimuli is necessary for its survival and involves plasticity and coordination of multiple levels of biological organization, including behavior, tissue organization, hormones, and gene expression. Each of these levels of response is complex, and none of them responds to stimuli in isolation. Thus, to understand how each system responds, it is necessary to consider its role in the context of the entire organism. Here, I have used the African cichlid fish Astatotilapia burtoni and its extraordinary phenotypic plasticity to investigate how animals respond to a change in social status from subordinate to dominant and attempted to integrate these multiple levels of biological response, as well as the roles of several candidate neuromodulators,. First, I have described how male A. burtoni become more aggressive and reproductive during their transition to dominance as well as increasing circulating levels of testosterone and estradiol and the histological organization of their testes. I then mapped the distribution of expression of two behaviorally relevant neuropeptides, arginine vasotocin and isotocin, and their respective receptors, throughout the A. burtoni brain, and found that they were highly expressed in several brain areas important for social behavior and decision-making. I then investigated the role of arginine vasotocin in social status and behavior via pharmacological manipulation and qPCR, showing the importance of arginine vasotocin in controlling the transition to dominance. Lastly, I investigated the role of aromatase, testosterone, and estradiol in male A. burtoni, both in stable dominant males and in males as they transition to dominance, using pharmacological manipulation and quantitative radioactive in situ hybridization, illustrating that estradiol synthesis during dominance is dependent on aromatase activity and necessary for aggressive behavior. / text

<p>For my doctoral research I examined social decision-making in a cooperatively breeding cichlid fish, <em>Neolamprologus pulcher</em> with a focus on affiliation and aggression. I investigated the role that the nonapeptide hormone, isotocin, plays in modulating social decisions in these contexts. I show that <em>N. pulcher</em> males prefer to join larger groups regardless of the rank at which they will join, whereas females prefer larger groups only when they can join a group in a high rank (Chapter 2). I examined decision-making during resource contests in<em> </em>(Chapter 3) and found that <em>N. pulcher</em> are sensitive to the size of their opponents, making fighting decisions depending on their opponents’ body size. I also found that smaller <em>N. pulcher</em> are more motivated to persist within contests, showing a shorter latency to resume fighting following interruption (Chapter 4). In Chapters 5 and 6, I explored the role of isotocin (the teleost fish homologue of oxytocin) in regulating social behaviour. I discovered that an increase in isotocin increased responsiveness to social information. Fish treated with isotocin were more sensitive to their opponent’s size in contests and were more submissive to dominant individuals within their social group (Chapter 5). Unexpectedly, I found that exogenous isotocin reduced sociality in <em>N. pulcher, </em>and that an isotocin receptor antagonist increased it (Chapter 6). These results suggest that the relationship between isotocin and social behaviour is both complex and context specific. In my final data chapter, I used social network analysis to explore the role of dominance interactions in determining the structure of <em>N. pulcher</em> social groups. I found that <em>N. pulcher</em> dominance hierarchies are highly linear, but that dominance interactions are not predicted by sex or body size asymmetry (Chapter 7). I found that conflict within <em>N. pulcher</em> social groups is greatest at the top of the dominance hierarchy. Taken together the results of my thesis helps to elucidate the behavioural and hormonal basis of social decision-making in a cooperatively breeding vertebrate and help to illuminate the evolution of social behaviour.</p> / Doctor of Philosophy (PhD)

No description available.

No description available.