A brain-wide analysis maps structural evolution to distinct anatomical module.

The vertebrate brain is highly conserved topologically, but less is known about neuroanatomical variation between individual brain regions. Neuroanatomical variation at the regional level is hypothesized to provide functional expansion, building upon ancestral anatomy needed for basic functions. Classically, animal models used to study evolution have lacked tools for detailed anatomical analysis that are widely used in zebrafish and mice, presenting a barrier to studying brain evolution at fine scales. In this study, we sought to investigate the evolution of brain anatomy using a single species of fish consisting of divergent surface and cave morphs, that permits functional genetic testing of regional volume and shape across the entire brain. We generated a high-resolution brain atlas for the blind Mexican cavefish Astyanax mexicanus and coupled the atlas with automated computational tools to directly assess variability in brain region shape and volume across all populations. We measured the volume and shape of every grossly defined neuroanatomical region of the brain and assessed correlations between anatomical regions in surface fish, cavefish, and surface × cave F2 hybrids, whose phenotypes span the range of surface to cave. We find that dorsal regions of the brain are contracted, while ventral regions have expanded, with F2 hybrid data providing support for developmental constraint along the dorsal-ventral axis. Furthermore, these dorsal-ventral relationships in anatomical variation show similar patterns for both volume and shape, suggesting that the anatomical evolution captured by these two parameters could be driven by similar developmental mechanisms. Together, these data demonstrate that A. mexicanus is a powerful system for functionally determining basic principles of brain evolution and will permit testing how genes influence early patterning events to drive brain-wide anatomical evolution.

Novel Husbandry Practices Result in Rapid Rates of Growth and Sexual Maturation Without Impacting Adult Behavior in the Blind Mexican Cavefish.

Animal model systems are dependent on the standardization of husbandry protocols that maximize growth and reduce generation time. The Mexican tetra, Astyanax mexicanus, exists as eyed surface and blind cave dwelling populations. The opportunity for comparative approaches between independently evolved populations has led to the rapid growth of A. mexicanus as a model for evolution and biomedical research. However, a slow and inconsistent growth rate remains a major limitation to the expanded application of A. mexicanus. Fortunately, this temporal limitation can be addressed through husbandry changes that accelerate growth rates while maintaining optimal health outcomes. Here, we describe a husbandry protocol that produces rapid growth rates through changes in diet, feeding frequency, growth sorting and progressive changes in tank size. This protocol produced robust growth rates and decreased the age of sexual maturity in comparison to our previous protocol. To determine whether changes in feeding impacted behavior, we tested fish in exploration and schooling assays. We found no difference in behavior between the two groups, suggesting that increased feeding and rapid growth will not impact the natural variation in behavioral traits. Taken together, this standardized husbandry protocol will accelerate the development of A. mexicanus as a genetic model.

Convergence on reduced aggression through shared behavioral traits in multiple populations of Astyanax mexicanus.

BACKGROUND: Aggression is observed across the animal kingdom, and benefits animals in a number of ways to increase fitness and promote survival. While aggressive behaviors vary widely across populations and can evolve as an adaptation to a particular environment, the complexity of aggressive behaviors presents a challenge to studying the evolution of aggression. The Mexican tetra, Astyanax mexicanus exists as an aggressive river-dwelling surface form and multiple populations of a blind cave form, some of which exhibit reduced aggression, providing the opportunity to investigate how evolution shapes aggressive behaviors. RESULTS: To define how aggressive behaviors evolve, we performed a high-resolution analysis of multiple social behaviors that occur during aggressive interactions in A. mexicanus. We found that many of the aggression-associated behaviors observed in surface-surface aggressive encounters were reduced or lost in Pachón cavefish. Interestingly, one behavior, circling, was observed more often in cavefish, suggesting evolution of a shift in the types of social behaviors exhibited by cavefish. Further, detailed analysis revealed substantive differences in aggression-related sub-behaviors in independently evolved cavefish populations, suggesting independent evolution of reduced aggression between cave populations. We found that many aggressive behaviors are still present when surface fish fight in the dark, suggesting that these reductions in aggression-associated and escape-associated behaviors in cavefish are likely independent of loss of vision in this species. Further, levels of aggression within populations were largely independent of type of opponent (cave vs. surface) or individual stress levels, measured through quantifying stress-like behaviors, suggesting these behaviors are hardwired and not reflective of population-specific changes in other cave-evolved traits. CONCLUSION: These results reveal that loss of aggression in cavefish evolved through the loss of multiple aggression-associated behaviors and raise the possibility that independent genetic mechanisms underlie changes in each behavior within populations and across populations. Taken together, these findings reveal the complexity of evolution of social behaviors and establish A. mexicanus as a model for investigating the evolutionary and genetic basis of aggressive behavior.

Cavefish brain atlases reveal functional and anatomical convergence across independently evolved populations.

Environmental perturbation can drive behavioral evolution and associated changes in brain structure and function. The Mexican fish species, Astyanax mexicanus, includes eyed river-dwelling surface populations and multiple independently evolved populations of blind cavefish. We used whole-brain imaging and neuronal mapping of 684 larval fish to generate neuroanatomical atlases of surface fish and three different cave populations. Analyses of brain region volume and neural circuits associated with cavefish behavior identified evolutionary convergence in hindbrain and hypothalamic expansion, and changes in neurotransmitter systems, including increased numbers of catecholamine and hypocretin/orexin neurons. To define evolutionary changes in brain function, we performed whole-brain activity mapping associated with behavior. Hunting behavior evoked activity in sensory processing centers, while sleep-associated activity differed in the rostral zone of the hypothalamus and tegmentum. These atlases represent a comparative brain-wide study of intraspecies variation in vertebrates and provide a resource for studying the neural basis of behavioral evolution.