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.

Neuroendocrine mechanisms contributing to the coevolution of sociality and communication.

Communication is inherently social, so signaling systems should evolve with social systems. The 'social complexity hypothesis' posits that social complexity necessitates communicative complexity and is generally supported in vocalizing mammals. This hypothesis, however, has seldom been tested outside the acoustic modality, and comparisons across studies are confounded by varying definitions of complexity. Moreover, proximate mechanisms underlying coevolution of sociality and communication remain largely unexamined. In this review, we argue that to uncover how sociality and communication coevolve, we need to examine variation in the neuroendocrine mechanisms that coregulate social behavior and signal production and perception. Specifically, we focus on steroid hormones, monoamines, and nonapeptides, which modulate both social behavior and sensorimotor circuits and are likely targets of selection during social evolution. Lastly, we highlight weakly electric fishes as an ideal system in which to comparatively address the proximate mechanisms underlying relationships between social and signal diversity in a novel modality.

Losing a parent in early-life impairs flock size discrimination and lowers oxytocin receptor abundance in a medial amygdala homologue of adult zebra finches.

Experiencing inadequate parental care during early-life diminishes adult social competencies. For example, low parental care impairs adult socio-cognitive abilities (e.g., recognizing familiar conspecifics) and affiliation (e.g., close social proximity); outcomes attributed to diminished medial amygdala nonapeptide functioning in rodents. Whether parental care has effects beyond familiarity, and if siblings have similar effects to parents, is unclear. Here, zebra finches were used to explore if parent and/or sibling number shape adult recognition and preference of small versus large flocks and nonapeptide (oxytocin, vasotocin) receptor expression in an avian homologue of the mammalian medial amygdala. Chicks were raised by single mothers or fathers in small broods or paired parents in small or large broods matched to single parents for chicks per nest or per parent, respectively. Pair-raised birds had preferred flock sizes as adults, but birds raised by single parents had equal preference for either size. Oxytocin receptor expression was lower in birds raised by single parents versus paired parents, but vasotocin receptor levels were unaffected. Such results highlight parents as formative antecedents of their offspring's social competencies related to group size preference and their nonapeptide mechanisms, outcomes that influence an animal's ability to live in social groups.

Vasopressin mediates nonapeptide and glucocorticoid signaling and social dynamics in juvenile dominance hierarchies of a highly social cichlid fish.

Early-life social experience can strongly affect adult behavior, yet the behavioral mechanisms underlying developmental trajectories are poorly understood. Here, we use the highly social cichlid, Burton's Mouthbrooder (Astatotilapia burtoni) to investigate juvenile social status and behavior, as well as the underlying neuroendocrine mechanisms. We placed juveniles in pairs or triads and found that they readily establish social status hierarchies, with some group structural variation depending on group size, as well as the relative body size of the group members. Next, we used intracerebroventricular injections to test the hypothesis that arginine vasopressin (AVP) regulates juvenile social behavior and status, similar to adult A. burtoni. While we found no direct behavioral effects of experimentally increasing (via vasotocin) or decreasing (via antagonist Manning Compound) AVP signaling, social interactions directed at the treated individual were significantly altered. This group-level effect of central AVP manipulation was also reflected in a significant shift in whole brain expression of genes involved in nonapeptide signaling (AVP, oxytocin, and oxytocin receptor) and the neuroendocrine stress axis (corticotropin-releasing factor (CRF), glucocorticoid receptors (GR) 1a and 1b). Further, social status was associated with the expression of genes involved in glucocorticoid signaling (GR1a, GR1b, GR2, mineralocorticoid receptor), social interactions with the dominant fish, and nonapeptide signaling activity (AVP, AVP receptor V1aR2, OTR). Together, our results considerably expand our understanding of the context-specific emergence of social dominance hierarchies in juveniles and demonstrate a role for nonapeptide and stress axis signaling in the regulation of social status and social group dynamics.

Uncovering the seasonal brain: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) as a biochemical approach for studying seasonal social behaviors.

Many animals show pronounced changes in physiology and behavior across the annual cycle, and these adaptations enable individuals to prioritize investing in the neuroendocrine mechanisms underlying reproduction and/or survival based on the time of year. While prior research has offered valuable insight into how seasonal variation in neuroendocrine processes regulates social behavior, the majority of these studies have investigated how a single hormone influences a single behavioral phenotype. Given that hormones are synthesized and metabolized via complex biochemical pathways and often act in concert to control social behavior, these approaches provide a limited view of how hormones regulate seasonal changes in behavior. In this review, we discuss how seasonal influences on hormones, the brain, and social behavior can be studied using liquid chromatography-tandem mass spectrometry (LC-MS/MS), an analytical chemistry technique that enables researchers to simultaneously quantify the concentrations of multiple hormones and the activities of their synthetic enzymes. First, we examine studies that have investigated seasonal plasticity in brain-behavior interactions, specifically by focusing on how two groups of hormones, sex steroids and nonapeptides, regulate sexual and aggressive behavior. Then, we explain the operations of LC-MS/MS, highlight studies that have used LC-MS/MS to study the neuroendocrine mechanisms underlying social behavior, both within and outside of a seasonal context, and discuss potential applications for LC-MS/MS in the field of behavioral neuroendocrinology. We propose that this cutting-edge technology will provide a more comprehensive understanding of how the multitude of hormones that comprise complex neuroendocrine networks affect seasonal variation in the brain and behavior.

Reciprocal processes of sensory perception and social bonding: an integrated social-sensory framework of social behavior.

Organisms filter the complexity of natural stimuli through their individual sensory and perceptual systems. Such perceptual filtering is particularly important for social stimuli. A shared "social umwelt" allows individuals to respond appropriately to the expected diversity of cues and signals during social interactions. In this way, the behavioral and neurobiological mechanisms of sociality and social bonding cannot be disentangled from perceptual mechanisms and sensory processing. While a degree of embeddedness between social and sensory processes is clear, our dominant theoretical frameworks favor treating the social and sensory processes as distinct. An integrated social-sensory framework has the potential to greatly expand our understanding of the mechanisms underlying individual variation in social bonding and sociality more broadly. Here we leverage what is known about sensory processing and pair bonding in two common study systems with significant species differences in their umwelt (rodent chemosensation and avian acoustic communication). We primarily highlight that (1) communication is essential for pair bond formation and maintenance, (2) the neural circuits underlying perception, communication and social bonding are integrated, and (3) candidate neuromodulatory mechanisms that regulate pair bonding also impact communication and perception. Finally, we propose approaches and frameworks that more fully integrate sensory processing, communication, and social bonding across levels of analysis: behavioral, neurobiological, and genomic. This perspective raises two key questions: (1) how is social bonding shaped by differences in sensory processing?, and (2) to what extent is sensory processing and the saliency of signals shaped by social interactions and emerging relationships?

Social partner cooperativeness influences brain oxytocin transcription in Trinidadian guppies (Poecilia reticulata).

For non-kin cooperation to be maintained, individuals need to respond adaptively to the cooperative behaviour of their social partners. Currently, however, little is known about the biological responses of individuals to experiencing cooperation. Here, we quantify the neuroregulatory response of Trinidadian guppies (Poecilia reticulata) experiencing cooperation or defection by examining the transcriptional response of the oxytocin gene (oxt; also known as isotocin), which has been implicated in cooperative decision-making. We exposed wild-caught females to social environments where partners either cooperated or defected during predator inspection, or to a control (non-predator inspection) context, and quantified the relative transcription of the oxt gene. We tested an experimental group, originating from a site where individuals are under high predation threat and have previous experience of large aquatic predators (HP), and a control group, where individuals are under low predation threat and naïve to large aquatic predators (LP). LP, but not HP, fish showed different behavioural responses to the behaviour of their social environment, cooperating with cooperative partners and defecting when paired with defecting ones. In HP, but not LP, fish brain mid-section oxt relative transcription varied depending on social partner behaviour. HP fish experiencing cooperation during predator inspection had lower oxt transcription than those experiencing defection. This effect was not present in the control population or in the control context, where the behaviour of social partners did not affect oxt transcription. Our findings provide insight into the neuromodulation underpinning behavioural responses to social experiences, and ultimately to the proximate mechanisms underlying social decision-making.

Toxoplasma gondii Infection Causes an Atypical Abundance of Oxytocin and Its Receptor in the Female Rat Brain.

Infection with the protozoan Toxoplasma gondii causes loss of innate fear of cat odors in both male and female rats. This behavioral change is presumed to reflect a parasitic manipulation that increases transmission of the parasite from its intermediate to definitive host. The host behavioral change in male rats is dependent on gonadal steroids. In contrast, the loss of fear in female rats is not accompanied by greater gonadal steroids and cannot be rescued by gonadectomy. This disparity suggests that proximate mechanisms of the post infection host behavioral change in rats are sexually dimorphic. Here, we report that female rats infected with Toxoplasma gondii exhibit greater abundance of messenger RNA for oxytocin and oxytocin receptors in the paraventricular nucleus of the hypothalamus and posterodorsal medial amygdala, respectively. Brain oxytocin is critical for sex-typical social and sexual behaviors in female rodents. The change in oxytocin and its receptor could potentially alter activity in the social salience circuits, leading to a reduction in defensive behaviors and an increase in approach to ambivalent environmental cues. Our results argue that sexually dimorphic neural substrates underpin sexually monomorphic host behavioral change in this host-parasite association.

Arginine vasopressin in the medial amygdala causes greater post-stress recruitment of hypothalamic vasopressin neurons.

Arginine vasopressin (AVP) is expressed in both hypothalamic and extra-hypothalamic neurons. The expression and role of AVP exhibit remarkable divergence between these two neuronal populations. Polysynaptic pathways enable these neuronal groups to regulate each other. AVP neurons in the paraventricular nucleus of the hypothalamus increase the production of adrenal stress hormones by stimulating the hypothalamic-pituitary-adrenal axis. Outside the hypothalamus, the medial amygdala also contains robust amounts of AVP. Contrary to the hypothalamic counterpart, the expression of extra-hypothalamic medial amygdala AVP is sexually dimorphic, in that it is preferentially transcribed in males in response to the continual presence of testosterone. Male gonadal hormones typically generate a negative feedback on the neuroendocrine stress axis. Here, we investigated whether testosterone-responsive medial amygdala AVP neurons provide negative feedback to hypothalamic AVP, thereby providing a feedback loop to suppress stress endocrine response during periods of high testosterone secretion. Contrary to our expectation, we found that AVP overexpression within the posterodorsal medial amygdala increased the recruitment of hypothalamic AVP neurons during stress, without affecting the total number of AVP neurons or the number of recently activated neurons following stress. These observations suggest that the effects of testosterone on extra-hypothalamic AVP facilitate stress responsiveness through permissive influence on the recruitment of hypothalamic AVP neurons.

Medial Amygdala Arginine Vasopressin Neurons Regulate Innate Aversion to Cat Odors in Male Mice.

Aversion to environmental cues of predators is an integral part of defensive behaviors in many prey animals. It enhances their survival and probability of future reproduction. At the same time, animals cannot be maximally defended because imperatives of defense usually trade-off with behaviors required for sexual reproduction like display of dominance and production of sexual pheromones. Here, we approach this trade-off through the lens of arginine vasopressin (AVP) neurons within the posterodorsal medial amygdala (MePD) of mice. This neuronal population is known to be involved in sexual behaviors like approach to sexually salient cues. We show that chemogenetic partial ablation of this neuronal population increases aversion to predator odors. Moreover, overexpression of AVP within this population is sufficient to reduce aversion to predator odors. The loss of fear of the predator odor occurs in parallel with increased recruitment of AVP neurons within the MePD. These observations suggest that AVP neurons in the medial aspect of the extended amygdala are a proximate locus for the reduction in innate fear during life stages dominated by reproductive efforts.

Female reproductive state is associated with changes in distinct arginine vasotocin cell types in the preoptic area of Astatotilapia burtoni.

Nonapeptides play a crucial role in mediating reproduction, aggression, and parental care across taxa. In fishes, arginine vasotocin (AVT) expression is related to social and/or reproductive status in most male fishes studied to date, and is linked to territorial defense, paternal care, and courtship. Despite a plethora of studies examining AVT in male fishes, relatively little is known about how AVT expression varies with female reproductive state or its role in female social behaviors. We used multiple methods for examining the AVT system in female African cichlid fish Astatotilapia burtoni, including immunohistochemistry for AVT, in situ hybridization for avt-mRNA, and quantitative PCR. Ovulated and mouthbrooding females had similar numbers of parvocellular, magnocellular, and gigantocellular AVT cells in the preoptic area. However, ovulated females had larger magnocellular and gigantocellular cells compared to mouthbrooding females, and gigantocellular AVT cell size correlated with the number of days brooding, such that late-stage brooding females had larger AVT cells than mid-stage brooding females. In addition, we found that ventral hypothalamic cells were more prominent in females compared to males, and were larger in mouthbrooding compared to ovulated females, suggesting a role in maternal care. Together, these data indicate that AVT neurons change across the reproductive cycle in female fishes, similar to that seen in males. These data on females complement studies in male A. burtoni, providing a comprehensive picture of the regulation and potential function of different AVT cell types in reproduction and social behaviors in both sexes.

Male-specific features are reduced in Mecp2-null mice: analyses of vasopressinergic innervation, pheromone production and social behaviour.

Deficits in arginine vasopressin (AVP) and oxytocin (OT), two neuropeptides closely implicated in the modulation of social behaviours, have been reported in some early developmental disorders and autism spectrum disorders. Mutations in the X-linked methyl-CpG-binding protein 2 (MECP2) gene are associated to Rett syndrome and other neuropsychiatric conditions. Thus, we first analysed AVP and OT expression in the brain of Mecp2-mutant mice by immunohistochemistry. Our results revealed no significant differences in these systems in young adult Mecp2-heterozygous females, as compared to WT littermates. By contrast, we found a significant reduction in the sexually dimorphic, testosterone-dependent, vasopressinergic innervation in several nuclei of the social brain network and oxytocinergic innervation in the lateral habenula of Mecp2-null males, as compared to WT littermates. Analysis of urinary production of pheromones shows that Mecp2-null males lack the testosterone-dependent pheromone darcin, strongly suggesting low levels of androgens in these males. In addition, resident-intruder tests revealed lack of aggressive behaviour in Mecp2-null males and decreased chemoinvestigation of the intruder. By contrast, Mecp2-null males exhibited enhanced social approach, as compared to WT animals, in a 3-chamber social interaction test. In summary, Mecp2-null males, which display internal testicles, display a significant reduction of some male-specific features, such as vasopressinergic innervation within the social brain network, male pheromone production and aggressive behaviour. Thus, atypical social behaviours in Mecp2-null males may be caused, at least in part, by the effect of lack of MeCP2 over sexual differentiation.

Gene expression correlates of social evolution in coral reef butterflyfishes.

Animals display remarkable variation in social behaviour. However, outside of rodents, little is known about the neural mechanisms of social variation, and whether they are shared across species and sexes, limiting our understanding of how sociality evolves. Using coral reef butterflyfishes, we examined gene expression correlates of social variation (i.e. pair bonding versus solitary living) within and between species and sexes. In several brain regions, we quantified gene expression of receptors important for social variation in mammals: oxytocin (OTR), arginine vasopressin (V1aR), dopamine (D1R, D2R) and mu-opioid (MOR). We found that social variation across individuals of the oval butterflyfish, Chaetodon lunulatus, is linked to differences in OTR,V1aR, D1R, D2R and MOR gene expression within several forebrain regions in a sexually dimorphic manner. However, this contrasted with social variation among six species representing a single evolutionary transition from pair-bonded to solitary living. Here, OTR expression within the supracommissural part of the ventral telencephalon was higher in pair-bonded than solitary species, specifically in males. These results contribute to the emerging idea that nonapeptide, dopamine and opioid signalling is a central theme to the evolution of sociality across individuals, although the precise mechanism may be flexible across sexes and species.

Nonapeptides mediate trade-offs in parental care strategy.

Parental care represents a suite of distinct behaviors performed by parents to maximize fitness. Dynamic shifts in parental care behaviors, such as between nest defense and direct provisioning of the offspring, are required in response to environmental variation. However, the neural mechanisms which mediate such behavioral shifts remain a mystery. The anemonefish, Amphiprion ocellaris, represents an experimentally valuable model in social neuroscience which is conducive to manipulating the environment while simultaneously measuring parental care. The goal of this study was to determine the extent to which arginine vasotocin (AVT) and isotocin (IT) signaling are necessary for males to shift between direct egg care and aggressive nest defense in the presence of intruders, Domino damselfish (Dascyllus trimaculatus). The IT receptor antagonist desGly-NH2-d(CH2)5[D-Tyr2,Thr4]OVT, significantly reduced direct egg care, while at the same time increased levels of aggressive nest defense relative to vehicle. Conversely, blockade of AVT using the antagonist d(CH2)5[Tyr(Me)2]AVP, reduced aggression and tended to increase egg care. Results demonstrate that male anemonefish alter their parental strategy in response to allospecific intruders, and that IT and AVT signaling oppositely regulate parental care displays of aggression versus egg care.

Effects of oxytocin-family peptides and substance P on locomotor activity and filial preferences in visually naïve chicks.

Nonapeptides from the vasopressin/oxytocin family have been implicated in a wide variety of social behaviours across vertebrates. Experimental manipulations that alter nonapeptide levels or receptor function in the brain have provided evidence for understanding how nonapeptides influence responses to social stimuli in adults. While behaviours in adults have been extensively studied, much less in known about roles of nonapeptides in early life and the development of affiliative social behaviours. We examined an experience-independent preference (social predisposition) that is present at hatching and is characterized by the tendency of visually naïve chicks (Gallus gallus) to prefer to approach a stuffed hen stimulus over a control stimulus in a choice test. Among chicks that show the social predisposition preference, bilateral intracranial mesotocin injections resulted in higher mean hen preference scores compared with saline-injected controls. Equimolar doses of mesotocin and vasotocin injections had different effects on locomotor activity: vasotocin, but not mesotocin, resulted in hypoactivity. We also tested whether intraperitoneal substance P had an effect on hen preference scores because previous research has proposed that vasotocin effects on social approach are mediated by peripheral release of substance P, but found no significant effect. All together, our data suggest that mesotocin signalling may be important for social predispositions and can potentially enhance the perceived salience of social stimuli soon after hatching. Specifically, mesotocin release and signalling in the brain may regulate the ability to recognize naturalistic stimuli and/or to act on the motivation to approach naturalistic stimuli.

The oxytocin system of mice and men-Similarities and discrepancies of oxytocinergic modulation in rodents and primates.

Nonapeptides and their respective receptors have been conserved throughout evolution and display astonishing similarities among the animal kingdom. They can be found in worms, birds, fish, amphibians, reptiles and mammals, including rodents, non-human primates and humans. In particular, the neuropeptide oxytocin (OT) has attracted the attention of scientists due to its profound effects on social behavior. However, although both the neuropeptide and its receptor are identical in rodents and primates, the effects of OT vary greatly in the two species. Here, we provide a brief overview about OT's role in the evolution of mammals and provide reasons for the manifold effects of OT within the brain with a particular focus on the discrepancy of OT's effects in rodents and primates. In addition, we suggest new approaches towards improvement of translatability of scientific studies and highlight the most recent advances in animal models for autism spectrum disorder, a disease, in which the normal function of the OT system seems to be impaired.

Corrigendum: Commentary: Arginine Vasotocin Preprohormone Is Expressed in Surprising Regions of the Teleost Forebrain.

[This corrects the article on p. 63 in vol. 9, PMID: 29545774.].

The Neurobiology of Mutualistic Behavior: The Cleanerfish Swims into the Spotlight.

One of the most notorious examples of cooperation between different species happens in the cleaner-client fish mutualism. The best known cleaner fish species, the bluestreak Indo-Pacific cleaner wrasse Labroides dimidiatus has been a model system to study the evolution of cooperation between unrelated animals and between distinct species during the last couple of decades. Given that the cleanerfish mutualism is well-established for behavioral studies of cooperation, it offered an outstanding opportunity to identify the link between cooperation, social cognition, and to undertake proximate studies, which were severely in need. This review surveys the current achievements of several recent studies, pointing towards the potential of the cleanerfish mutualism as a relevant model system for future accomplishments in neuroendocrine research.

Sensitive Periods, Vasotocin-Family Peptides, and the Evolution and Development of Social Behavior.

Nonapeptides, by modulating the activity of neural circuits in specific social contexts, provide an important mechanism underlying the evolution of diverse behavioral phenotypes across vertebrate taxa. Vasotocin-family nonapeptides, in particular, have been found to be involved in behavioral plasticity and diversity in social behavior, including seasonal variation, sexual dimorphism, and species differences. Although nonapeptides have been the focus of a great deal of research over the last several decades, the vast majority of this work has focused on adults. However, behavioral diversity may also be explained by the ways in which these peptides shape neural circuits and influence social processes during development. In this review, I synthesize comparative work on vasotocin-family peptides during development and classic work on early forms of social learning in developmental psychobiology. I also summarize recent work demonstrating that early life manipulations of the nonapeptide system alter attachment, affiliation, and vocal learning in zebra finches. I thus hypothesize that vasotocin-family peptides are involved in the evolution of social behaviors through their influence on learning during sensitive periods in social development.