Differential Neuroanatomical, Neurochemical, and Behavioral Impacts of Early-Age Isolation in a Eusocial Insect.

INTRODUCTION: Social experience early in life appears to be necessary for the development of species-typical behavior. Although isolation during critical periods of maturation has been shown to impact behavior by altering gene expression and brain development in invertebrates and vertebrates, workers of some ant species appear resilient to social deprivation and other neurobiological challenges that occur during senescence or due to loss of sensory input. It is unclear if and to what degree neuroanatomy, neurochemistry, and behavior will show deficiencies if social experience in the early adult life of worker ants is compromised. METHODS: We reared newly-eclosed adult workers of Camponotus floridanus under conditions of social isolation for 2 to 53 days, quantified brain compartment volumes, recorded biogenic amine levels in individual brains, and evaluated movement and behavioral performance to compare the neuroanatomy, neurochemistry, brood-care behavior, and foraging (predatory behavior) of isolated workers with that of workers experiencing natural social contact after adult eclosion. RESULTS: We found that the volume of the antennal lobe, which processes olfactory inputs, was significantly reduced in workers isolated for an average of 40 days, whereas the size of the mushroom bodies, centers of higher-order sensory processing, increased after eclosion and was not significantly different from controls. Titers of the neuromodulators serotonin, dopamine, and octopamine remained stable and were not significantly different in isolation treatments and controls. Brood care, predation, and overall movement were reduced in workers lacking social contact early in life. CONCLUSION: These results suggest that the behavioral development of isolated workers of C. floridanus is specifically impacted by a reduction in the size of the antennal lobe. Task performance and locomotor ability therefore appear to be sensitive to a loss of social contact through a reduction of olfactory processing ability rather than change in the size of the mushroom bodies, which serve important functions in learning and memory, or the central complex, which controls movement.

Differential Neuroanatomical, Neurochemical, and Behavioral Impacts of Early-Age Isolation in a Eusocial Insect.

Social experience early in life appears to be necessary for the development of species-typical behavior. Although isolation during critical periods of maturation has been shown to impact behavior by altering gene expression and brain development in invertebrates and vertebrates, workers of some ant species appear resilient to social deprivation and other neurobiological challenges that occur during senescence or due to loss of sensory input. It is unclear if and to what degree neuroanatomy, neurochemistry, and behavior will show deficiencies if social experience in the early adult life of worker ants is compromised. We reared newly-eclosed adult workers of Camponotus floridanus under conditions of social isolation for 2 to 53 days, quantified brain compartment volumes, recorded biogenic amine levels in individual brains, and evaluated movement and behavioral performance to compare the neuroanatomy, neurochemistry, brood-care behavior, and foraging (predatory behavior) of isolated workers with that of workers experiencing natural social contact after adult eclosion. We found that the volume of the antennal lobe, which processes olfactory inputs, was significantly reduced in workers isolated for an average of 40 days, whereas the size of the mushroom bodies, centers of higher-order sensory processing, increased after eclosion and was not significantly different from controls. Titers of the neuromodulators serotonin, dopamine, and octopamine remained stable and were not significantly different in isolation treatments and controls. Brood care, predation, and overall movement were reduced in workers lacking social contact early in life. These results suggest that the behavioral development of isolated workers of C. floridanus is specifically impacted by a reduction in the size of the antennal lobe. Task performance and locomotor ability therefore appear to be sensitive to a loss of social contact through a reduction of olfactory processing ability rather than change in the size of the mushroom bodies, which serve important functions in learning and memory, or the central complex, which controls movement.

White Paper: An Integrated Perspective on the Causes of Hypometric Metabolic Scaling in Animals.

Larger animals studied during ontogeny, across populations, or across species, usually have lower mass-specific metabolic rates than smaller animals (hypometric scaling). This pattern is usually observed regardless of physiological state (e.g. basal, resting, field, maximally-active). The scaling of metabolism is usually highly correlated with the scaling of many life history traits, behaviors, physiological variables, and cellular/molecular properties, making determination of the causation of this pattern challenging. For across-species comparisons of resting and locomoting animals (but less so for across populations or during ontogeny), the mechanisms at the physiological and cellular level are becoming clear. Lower mass-specific metabolic rates of larger species at rest are due to a) lower contents of expensive tissues (brains, liver, kidneys), and b) slower ion leak across membranes at least partially due to membrane composition, with lower ion pump ATPase activities. Lower mass-specific costs of larger species during locomotion are due to lower costs for lower-frequency muscle activity, with slower myosin and Ca++ ATPase activities, and likely more elastic energy storage. The evolutionary explanation(s) for hypometric scaling remain(s) highly controversial. One subset of evolutionary hypotheses relies on constraints on larger animals due to changes in geometry with size; for example, lower surface-to-volume ratios of exchange surfaces may constrain nutrient or heat exchange, or lower cross-sectional areas of muscles and tendons relative to body mass ratios would make larger animals more fragile without compensation. Another subset of hypotheses suggests that hypometric scaling arises from biotic interactions and correlated selection, with larger animals experiencing less selection for mass-specific growth or neurolocomotor performance. A additional third type of explanation comes from population genetics. Larger animals with their lower effective population sizes and subsequent less effective selection relative to drift may have more deleterious mutations, reducing maximal performance and metabolic rates. Resolving the evolutionary explanation for the hypometric scaling of metabolism and associated variables is a major challenge for organismal and evolutionary biology. To aid progress, we identify some variation in terminology use that has impeded cross-field conversations on scaling. We also suggest that promising directions for the field to move forward include: 1) studies examining the linkages between ontogenetic, population-level, and cross-species allometries, 2) studies linking scaling to ecological or phylogenetic context, 3) studies that consider multiple, possibly interacting hypotheses, and 4) obtaining better field data for metabolic rates and the life history correlates of metabolic rate such as lifespan, growth rate and reproduction.

Social Brain Energetics: Ergonomic Efficiency, Neurometabolic Scaling, and Metabolic Polyphenism in Ants.

Metabolism, a metric of the energy cost of behavior, plays a significant role in social evolution. Body size and metabolic scaling are coupled, and a socioecological pattern of increased body size is associated with dietary change and the formation of larger and more complex groups. These consequences of the adaptive radiation of animal societies beg questions concerning energy expenses, a substantial portion of which may involve the metabolic rates of brains that process social information. Brain size scales with body size, but little is understood about brain metabolic scaling. Social insects such as ants show wide variation in worker body size and morphology that correlates with brain size, structure, and worker task performance, which is dependent on sensory inputs and information-processing ability to generate behavior. Elevated production and maintenance costs in workers may impose energetic constraints on body size and brain size that are reflected in patterns of metabolic scaling. Models of brain evolution do not clearly predict patterns of brain metabolic scaling, nor do they specify its relationship to task performance and worker ergonomic efficiency, two key elements of social evolution in ants. Brain metabolic rate is rarely recorded and therefore the conditions under which brain metabolism influences the evolution of brain size are unclear. We propose that studies of morphological evolution, colony social organization, and worker ergonomic efficiency should be integrated with analyses of species-specific patterns of brain metabolic scaling to advance our understanding of brain evolution in ants.

Eusociality and Senescence: Neuroprotection and Physiological Resilience to Aging in Insect and Mammalian Systems.

Are eusociality and extraordinary aging polyphenisms evolutionarily coupled? The remarkable disparity in longevity between social insect queens and sterile workers-decades vs. months, respectively-has long been recognized. In mammals, the lifespan of eusocial naked mole rats is extremely long-roughly 10 times greater than that of mice. Is this robustness to senescence associated with social evolution and shared mechanisms of developmental timing, neuroprotection, antioxidant defenses, and neurophysiology? Focusing on brain senescence, we examine correlates and consequences of aging across two divergent eusocial clades and how they differ from solitary taxa. Chronological age and physiological indicators of neural deterioration, including DNA damage or cell death, appear to be decoupled in eusocial insects. In some species, brain cell death does not increase with worker age and DNA damage occurs at similar rates between queens and workers. In comparison, naked mole rats exhibit characteristics of neonatal mice such as protracted development that may offer protection from aging and environmental stressors. Antioxidant defenses appear to be regulated differently across taxa, suggesting independent adaptations to life history and environment. Eusocial insects and naked mole rats appear to have evolved different mechanisms that lead to similar senescence-resistant phenotypes. Careful selection of comparison taxa and further exploration of the role of metabolism in aging can reveal mechanisms that preserve brain functionality and physiological resilience in eusocial species.

The neuroplasticity of division of labor: worker polymorphism, compound eye structure and brain organization in the leafcutter ant Atta cephalotes.

Our understanding of how sensory structure design is coupled with neural processing capacity to adaptively support division of labor is limited. Workers of the remarkably polymorphic fungus-growing ant Atta cephalotes are behaviorally specialized by size: the smallest workers (minims) tend fungi in dark subterranean chambers while larger workers perform tasks outside the nest. Strong differences in worksite light conditions are predicted to influence sensory and processing requirements for vision. Analyzing confocal scans of worker eyes and brains, we found that eye structure and visual neuropils appear to have been selected to maximize task performance according to light availability. Minim eyes had few ommatidia, large interommatidial angles and eye parameter values, suggesting selection for visual sensitivity over acuity. Large workers had larger eyes with disproportionally more and larger ommatidia, and smaller interommatidial angles and eye parameter values, indicating peripheral sensory adaptation to ambient rainforest light. Optic lobes and mushroom body collars were disproportionately small in minims. Within the optic lobe, lamina and lobula relative volumes increased with worker size whereas medulla volume decreased. Visual system phenotypes thus correspond to task specializations in dark or light environments and illustrate a functional neuroplasticity underpinning division of labor in this socially complex agricultural ant.

Correction: Division of labor and brain evolution in insect societies: Neurobiology of extreme specialization in the turtle ant Cephalotes varians.

[This corrects the article DOI: 10.1371/journal.pone.0213618.].

Social Complexity and Brain Evolution: Comparative Analysis of Modularity and Integration in Ant Brain Organization.

The behavioral demands of living in social groups have been linked to the evolution of brain size and structure, but how social organization shapes investment and connectivity within and among functionally specialized brain regions remains unclear. To understand the influence of sociality on brain evolution in ants, a premier clade of eusocial insects, we statistically analyzed patterns of brain region size covariation as a proxy for brain region connectivity. We investigated brain structure covariance in young and old workers of two formicine ants, the Australasian weaver ant Oecophylla smaragdina, a pinnacle of social complexity in insects, and its socially basic sister clade Formica subsericea. As previously identified in other ant species, we predicted that our analysis would recognize in both species an olfaction-related brain module underpinning social information processing in the brain, and a second neuroanatomical cluster involved in nonolfactory sensorimotor processes, thus reflecting conservation of compartmental connectivity. Furthermore, we hypothesized that covariance patterns would reflect divergence in social organization and life histories either within this species pair or compared to other ant species. Contrary to our predictions, our covariance analyses revealed a weakly defined visual, rather than olfactory, sensory processing cluster in both species. This pattern may be linked to the reliance on vision for worker behavioral performance outside of the nest and the correlated expansion of the optic lobes to meet navigational demands in both species. Additionally, we found that colony size and social organization, key measures of social complexity, were only weakly correlated with brain modularity in these formicine ants. Worker age also contributed to variance in brain organization, though in different ways in each species. These findings suggest that brain organization may be shaped by the divergent life histories of the two study species. We compare our findings with patterns of brain organization of other eusocial insects.

Division of labor and brain evolution in insect societies: Neurobiology of extreme specialization in the turtle ant Cephalotes varians.

Strongly polyphenic social insects provide excellent models to examine the neurobiological basis of division of labor. Turtle ants, Cephalotes varians, have distinct minor worker, soldier, and reproductive (gyne/queen) morphologies associated with their behavioral profiles: small-bodied task-generalist minors lack the phragmotic shield-shaped heads of soldiers, which are specialized to block and guard the nest entrance. Gynes found new colonies and during early stages of colony growth overlap behaviorally with soldiers. Here we describe patterns of brain structure and synaptic organization associated with division of labor in C. varians minor workers, soldiers, and gynes. We quantified brain volumes, determined scaling relationships among brain regions, and quantified the density and size of microglomeruli, synaptic complexes in the mushroom body calyxes important to higher-order processing abilities that may underpin behavioral performance. We found that brain volume was significantly larger in gynes; minor workers and soldiers had similar brain sizes. Consistent with their larger behavioral repertoire, minors had disproportionately larger mushroom bodies than soldiers and gynes. Soldiers and gynes had larger optic lobes, which may be important for flight and navigation in gynes, but serve different functions in soldiers. Microglomeruli were larger and less dense in minor workers; soldiers and gynes did not differ. Correspondence in brain structure despite differences in soldiers and gyne behavior may reflect developmental integration, suggesting that neurobiological metrics not only advance our understanding of brain evolution in social insects, but may also help resolve questions of the origin of novel castes.

Synaptic organization and division of labor in the exceptionally polymorphic ant Pheidole rhea.

Social insect polyphenisms provide models to examine the neural basis of division of labor and anatomy of the invertebrate social brain. Worker size-related behavior is hypothesized to enhance task performance, raising questions concerning the integration of morphology, behavior, and cellular neuroarchitecture, and how variation in sensory inputs and cognitive demands of behaviorally differentiated workers is reflected in higher-order processing ability. We used the highly polymorphic ant Pheidole rhea, which has three distinct worker size classes - minors, soldiers, and supersoldiers - to examine variation in synaptic circuitry across worker size and social role. We hypothesized that the density and size of synaptic complexes (microglomeruli, MG) would be positively associated with behavioral repertoire and the relative size of the mushroom bodies (MB). Supersoldiers had significantly larger and less dense MG in the lip (olfactory region) of the MB calyx (MBC), and larger MG in the collar (visual region) compared to minors. Soldiers were intermediate in synaptic phenotype: they did not differ significantly in MG density from minors and supersoldiers, had MG of similar size to minors in the lip, and did not differ from these two worker groups in MG size in the collar. Results suggest a complex relationship between MG density, size, behavior, and worker body size involving a conserved and plastic neurobiological development plan, although workers show strong variation in size and social role.

Interspecific comparison of mushroom body synaptic complexes of dimorphic workers in the ant genus Pheidole.

Social insects may have morphologically and behaviorally specialized workers that vary in requirements for sensory information processing, making them excellent systems to examine the relationship between brain structure and behavior. The density and size of synaptic complexes (microglomeruli, MG) in the mushroom bodies (MB) have served as proxies for processing ability and synaptic plasticity, and have been shown to vary among insect species that differ in behavioral complexity. To understand the relationship between behavioral specialization and synaptic structure, we examined age-related changes in MG density and size between minor worker and soldier subcastes in two species of Pheidole ants, P. dentata and P. morrisi, that differ in behavior. We hypothesized that task-diverse minor workers would have more densely packed MG than soldiers, and that species-specific differences in soldier repertories would be reflected in MG structure. We also examined MG variation in young and mature minor workers and soldiers, predicting that as workers age and develop behaviorally, MG would decrease in density in both subcastes due to synaptic pruning. Results support the hypothesis that MG density in the lip (olfactory) and collar (visual) regions of the MBs decrease with age in association with increases in bouton size in the lip. However, minors had significantly lower densities of MG in the lip than soldiers, suggesting MG may not show structural variation according to subcaste-related differences in cognitive demands in either species.

Origins of Aminergic Regulation of Behavior in Complex Insect Social Systems.

Neuromodulators are conserved across insect taxa, but how biogenic amines and their receptors in ancestral solitary forms have been co-opted to control behaviors in derived socially complex species is largely unknown. Here we explore patterns associated with the functions of octopamine (OA), serotonin (5-HT) and dopamine (DA) in solitary ancestral insects and their derived functions in eusocial ants, bees, wasps and termites. Synthesizing current findings that reveal potential ancestral roles of monoamines in insects, we identify physiological processes and conserved behaviors under aminergic control, consider how biogenic amines may have evolved to modulate complex social behavior, and present focal research areas that warrant further study.

Behavioral Performance and Neural Systems Are Robust to Sensory Injury in Workers of the Ant Pheidole dentata.

Miniaturized nervous systems have been thought to limit behavioral ability, and animals with miniaturized brains may be less flexible when challenged by injuries resulting in sensory deficits that impact the development, maintenance, and plasticity of small-scale neural networks. We experimentally examined how injuries to sensory structures critical for olfactory ability affect behavioral performance in workers of the ant Pheidole dentata, which have minute brains (0.01 mm3) and primarily rely on the perception and processing of chemical signals and cues to direct their social behavior. We employed unilateral antennal denervation to decrease the olfactory perception ability of workers and quantified consequential neuroanatomical and behavioral performance effects. Postablation neuroanatomical metrics revealed a 25% reduction in the volume of the antennal lobe ipsilateral to the antennal lesion relative to the contralateral lobe, indicating atrophy of the input-deprived tissue. However, antennectomy did not affect the volumes of the mushroom body or its subcompartments or the number of mushroom body synaptic complexes (microglomeruli) in either brain hemisphere. Synapsin immunoreactivity, however, was significantly higher in the ipsilateral mushroom body calyces, which could reflect presynaptic potentiation and homeostatic compensation in higher-order olfactory regions. Despite tissue loss caused by antennal lesioning and resulting unilateral sensory deprivation, the ability of workers to perform behaviors that encompass the breadth of their task repertoire and meet demands for colony labor remained largely intact. The few behavioral deficits recorded were restricted to pheromone trail-following ability, a result that was expected due to the need for bilateral olfactory input to process spatial odor information. Our macroscopic and cellular neuroanatomical measurements and assessments of task performance demonstrate that the miniaturized brains of P. dentata workers and their sensorimotor functions are remarkably robust to injury-related size reduction and remain capable of generating behaviors required to respond appropriately to chemical social signals and effectively nurse immatures, as well as participate in coordinated foraging.

Worker brain development and colony organization in ants: Does division of labor influence neuroplasticity?

Brain compartment size allometries may adaptively reflect cognitive needs associated with behavioral development and ecology. Ants provide an informative system to study the relationship of neural architecture and development because worker tasks and sensory inputs may change with age. Additionally, tasks may be divided among morphologically and behaviorally differentiated worker groups (subcastes), reducing repertoire size through specialization and aligning brain structure with task-specific cognitive requirements. We hypothesized that division of labor may decrease developmental neuroplasticity in workers due to the apparently limited behavioral flexibility associated with task specialization. To test this hypothesis, we compared macroscopic and cellular neuroanatomy in two ant sister clades with striking contrasts in worker morphological differentiation and colony-level social organization: Oecophylla smaragdina, a socially complex species with large colonies and behaviorally distinct dimorphic workers, and Formica subsericea, a socially basic species with small colonies containing monomorphic workers. We quantified volumes of functionally distinct brain compartments in newly eclosed and mature workers and measured the effects of visual experience on synaptic complex (microglomeruli) organization in the mushroom bodies-regions of higher-order sensory integration-to determine the extent of experience-dependent neuroplasticity. We demonstrate that, contrary to our hypothesis, O. smaragdina workers have significant age-related volume increases and synaptic reorganization in the mushroom bodies, whereas F. subsericea workers have reduced age-related neuroplasticity. We also found no visual experience-dependent synaptic reorganization in either species. Our findings thus suggest that changes in the mushroom body with age are associated with division of labor, and therefore social complexity, in ants. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 77: 1072-1085, 2017.

Social complexity influences brain investment and neural operation costs in ants.

The metabolic expense of producing and operating neural tissue required for adaptive behaviour is considered a significant selective force in brain evolution. In primates, brain size correlates positively with group size, presumably owing to the greater cognitive demands of complex social relationships in large societies. Social complexity in eusocial insects is also associated with large groups, as well as collective intelligence and division of labour among sterile workers. However, superorganism phenotypes may lower cognitive demands on behaviourally specialized workers resulting in selection for decreased brain size and/or energetic costs of brain metabolism. To test this hypothesis, we compared brain investment patterns and cytochrome oxidase (COX) activity, a proxy for ATP usage, in two ant species contrasting in social organization. Socially complex Oecophylla smaragdina workers had larger brain size and relative investment in the mushroom bodies (MBs)-higher order sensory processing compartments-than the more socially basic Formica subsericea workers. Oecophylla smaragdina workers, however, had reduced COX activity in the MBs. Our results suggest that as in primates, ant group size is associated with large brain size. The elevated costs of investment in metabolically expensive brain tissue in the socially complex O. smaragdina, however, appear to be offset by decreased energetic costs.

Lifespan behavioural and neural resilience in a social insect.

Analyses of senescence in social species are important to understanding how group living influences the evolution of ageing in society members. Social insects exhibit remarkable lifespan polyphenisms and division of labour, presenting excellent opportunities to test hypotheses concerning ageing and behaviour. Senescence patterns in other taxa suggest that behavioural performance in ageing workers would decrease in association with declining brain functions. Using the ant Pheidole dentata as a model, we found that 120-day-old minor workers, having completed 86% of their laboratory lifespan, showed no decrease in sensorimotor functions underscoring complex tasks such as alloparenting and foraging. Collaterally, we found no age-associated increases in apoptosis in functionally specialized brain compartments or decreases in synaptic densities in the mushroom bodies, regions associated with integrative processing. Furthermore, brain titres of serotonin and dopamine--neuromodulators that could negatively impact behaviour through age-related declines--increased in old workers. Unimpaired task performance appears to be based on the maintenance of brain functions supporting olfaction and motor coordination independent of age. Our study is the first to comprehensively assess lifespan task performance and its neurobiological correlates and identify constancy in behavioural performance and the absence of significant age-related neural declines.