Journal of Experimental Biology

Despite breathing water using their tidally ventilated rectal gills, dragonfly nymphs show a surprising ability to maintain oxygen (O2) extraction from the water during hypoxia. However, an increase in convective O2 transfer is insufficient to sustain aerobic demands by itself, which suggests that diffusive mechanisms must also be involved. This study examines the contributions of changing the O2 partial pressure gradient (PO2) and/or O2 conductance across the rectal gill in maintaining O2 extraction efficiency (OEE) of dragonfly nymphs during hypoxia. Data were collected using the same custom-designed respiro-spirometer described in a previous study with the addition of an implanted O2 sensor to measure hemolymph PO2. Results show that the implantation of the O2 sensor does not affect the respiratory and ventilatory response of nymphs to hypoxia. Hemolymph PO2 fell from 6.3 {+/-} 1.6 kPa at normoxia to 2.5 {+/-} 0.6 kPa at 16.0 kPa, which resulted in the PO2 diffusion gradient remaining statistically constant at these two water PO2s (17.5 {+/-} 1.7 and 15.4 {+/-} 0.7 kPa during normoxia and 16.0 kPa respectively). Beyond 16.0 kPa, a progressive reduction in hemolymph PO2 was unable to sustain the diffusion gradient. Mathematical modeling revealed that while the addition of hemolymph PO2 in tandem with ventilation frequency was able to elevate OEE during 16.0 kPa to that of normoxia, both were still insufficient during severe hypoxia and required an increase in O2 conductance. Estimating the change in whole-gill conductance showed that nymphs are indeed increasing their conductance as the water becomes hypoxic, demonstrating a reliance on both diffusion gradient and O2 conductance to enhance diffusive O2 transfer in conjunction with convective mechanisms to maintain O2 extraction during hypoxia.

In many animals, ultraviolet (UV) vision guides navigation, foraging, and communication, but few studies have addressed the contribution of UV vision to color discrimination, or behaviorally assessed UV discrimination thresholds. Here, we tested UV-color vision in an anemonefish (Amphiprion ocellaris) using a novel five-channel (RGB-V-UV) LED display designed to test UV perception. We first determined that the maximal sensitivity of the A. ocellaris UV cone was at [~]386 nm using microspectrophotometry. Three additional cone spectral sensitivities had maxima at [~]497, 515, and [~]535 nm, which together informed the modelling of the fishs color vision. Anemonefish behavioral discrimination thresholds for nine sets of colors were determined from their ability to distinguish a colored target pixel from grey distractor pixels of varying intensity. We found that A. ocellaris used all four cones to process color information and is therefore tetrachromatic, and fish were better at discriminating colors (i.e., color discrimination thresholds were lower, or more acute) when targets had UV chromatic contrast elicited by greater stimulation of the UV cone relative to other cone types. These findings imply that a UV component of color signals and cues improves their detectability, that likely increases the salience of anemonefish body patterns used in communication and the silhouette of zooplankton prey.

We present evidence that the faint polarised moonlight pattern of the sky can be detected and used for navigation in the diurnal bull ant Myrmecia tarsata, despite this species lacking the highly refined low-light visual specialisations of the nocturnal bull ant, Myrmecia midas. The position of celestial bodies such as the sun and moon can provide navigating animals directional information, yet direct observation can often be occluded. Animals can estimate positional information of solar and lunar cues via their polarised light pattern, present across the sky. The suns polarisation pattern is widely used in animals and a similar, yet much fainter pattern is produced by the moon, yet it is unknown how widespread moonlights use is in navigating animals. Here, we demonstrate that the bull ant Myrmecia tarsata, which forages throughout the day, returning home at sunset, can use both solar and lunar polarised light patterns to navigate. We compare these findings to the closely related M. midas navigating under identical light conditions, as this nocturnal bull ant is known to rely on these polarised light patterns as part of their celestial compass. While M. midas and M. tarsata can clearly use both the solar and lunar polarised light patterns to navigate, M. tarsata showed degraded performance under polarised moonlight as a function of lunar phase, decreasing performance as illumination decreased. M. midas in contrast, exhibited impressive attendance to the overhead lunar polarisation pattern throughout the lunar month, illustrating the highly specialised low-light-detection adaptations of bull ants.

An integral component of musculoskeletal systems are elastic elements mechanically in-series with muscle. Although these in-series elastic elements--e. g. tendons in vertebrates, or apodemes in invertebrates--can neither generate force nor do work, they are thought to bring substantial benefits to musculoskeletal performance; the mechanical properties of tendons, crucial determinants of these benefits, have consequently been subject of a large body of work. In sharp contrast, scarce information exists on the mechanical properties of apodemes. The little data that do exist appear to suggest that apodemes differ so substantially from tendons that their functional significance may differ, too. To increase our understanding of apodeme function, we determined the mechanical properties of the unguitractor apodeme (UTA) of Sungaya aeta stick insects. We devised an experimental protocol that permits tensile testing with slippery and brittle apodemes; we derived and validated a mechanical model that extracts the Youngs modulus from tensile tests with specimen with varying cross-sectional area, without the need for explicit measurement of the stress or strain distribution; and we interpreted the magnitude of the UTA modulus, strength and spring constant through allometric comparison with data on vertebrate tendons. The UTA modulus exceeds that of vertebrate tendons by almost one order of magnitude, but the size-corrected spring constant is nevertheless comparable and if anything smaller, due to systematic differences in apodeme and tendon shape. This many-to-one mapping suggests that apodemes may well convey the same functional benefits as tendons, and should not be prematurely excluded from invertebrate musculoskeletal models.

Small cursorial birds display remarkable walking skills and can negotiate complex and unstructured terrains with ease. The neuromechanical control strategies necessary to adapt to these challenging terrains are still not well understood. Here, we analyzed the 2D- and 3D pelvic and leg kinematic strategies employed by the common quail to negotiate visible step-up and step-down perturbations of 1 cm, 2.5 cm, and 5 cm. We used biplanar fluoroscopy to accurately describe joint positions in three dimensions and performed semi-automatic landmark localization using deep learning. Quails negotiated vertical perturbations without major problems and rapidly regained steady-state locomotion. When coping with step-up perturbations, the quail mostly adapted the trailing limb to permit the leading leg to step on the elevated substrate in a similar way as it did during level locomotion. When the quail negotiated step-down perturbations, both legs showed significant adaptations. For small and moderate perturbations (not inducing aerial running) the quail kept the function of the distal joints (i.e., their kinematic pattern) largely unchanged during uneven locomotion, and most changes occurred in proximal joints. The hip regulated leg length, while the distal joints maintained the spring-damped limb patterns. However, to negotiate the largest visible step perturbations, more dramatic kinematic alterations were observed. For these large perturbations, all joints contributed to leg lengthening/ shortening in the trailing leg and both the trailing and leading legs stepped more vertically and less abducted. This indicates a shift from a dynamic walking program to strategies that are focused on maximizing safety.

Research on birds suggests that extreme weather events during development may have long-lasting consequences on form and function. The underlying cellular mechanisms mediating such phenotypic effects are poorly studied. We raised Japanese quail in warm (30{degrees}C) or cold (10{degrees}C) temperatures from hatching until adulthood, and then measured mitochondrial metabolism in intact blood cells at representative normothermic body temperature (41{degrees}C) and a hyperthermic temperature (45{degrees}C) that quail commonly attain when heat stressed. To investigate whether any developmental effects were reversible, half of the cold- and warm-acclimated birds were assigned to a common garden (20{degrees}C) 3 weeks before the measurements. Across groups, hyperthermia was associated with increased proton leak, but decreases in both phosphorylating respiration (where ATP is produced) and working capacity of the mitochondria. Cold-acclimated birds were more strongly affected by heat stress: the increase in proton leak was 1.6-fold higher, and the decrease in phosphorylating capacity during endogenous respiration was 1.7-fold greater, compared to warm-acclimated birds. These differences did not remain in the common-garden birds. Our study suggests that developmental cold acclimation is traded off against heat tolerance at the level of cellular metabolism, with implications for our understanding of avian responses to climate change.

Although mitochondrial respiration is believed to explain a substantial part of the variation in whole-animal basal (BMR) or resting metabolic rate (RMR), few studies have addressed the relationship between organismal and cellular metabolism and how this may vary in environments where individual demands for energy differ. We investigated the relationship between whole-individual metabolic rate, measured in temperatures ranging thermoneutrality to far below thermoneutrality, and mitochondrial respiration of intact or permeabilized blood cells in two separate studies on wild great tits (Parus major L.). Our results show that, in permeabilized cells, there are significant positive relationships between BMR or RMR and several mitochondrial traits, including phosphorylating respiration rate through both complexes I and II (i.e., OXPHOS respiration). However, surprisingly, the LEAK respiration (i.e., basal respiration that mainly counteract for proton leakage) was not related to BMR or RMR. When measurements were performed using intact blood cells, BMR was positively related to ROUTINE respiration (i.e., mitochondrial respiration on endogenous substrates) in one of the two studies, but no other mitochondrial traits could explain variation in BMR or RMR in any thermal environment. These studies seem to show that the level of activation of mitochondrial metabolism as well as the permeabilization status of blood cells play a primary role on the extent to which blood metabolism might explain variations in the whole-individual metabolic rate.

Insects use rapid acclimation to enhance their tolerance of abiotic stresses within minutes to hours. These responses are critical adaptations for insects and other small ectotherms to tolerate drastic changes in temperature, hydration, or other factors that can fluctuate precipitously with ambient conditions or as a result of behavior. Rapid cold-hardening, where insects use brief exposure to modest chilling as a cue to enhance their cold tolerance, is the most thoroughly-studied of these responses and relatively little is known about rapid acclimation that is either triggered by or enhances tolerance of other abiotic stresses. Here, we used larvae of the Antarctic midge, Belgica antarctica, a polar extremophile that routinely experiences numerous stresses in nature, to investigate how 2 h exposure to modest environmental stresses affect stress tolerance in insects. Brief pretreatment by various stresses, including hyperosmotic challenge, hypoosmotic challenge, acidity, basicity, and UV irradiation enhanced stress tolerance in B. antarctica larvae relative to untreated controls. These results indicate that numerous environmental cues can trigger rapid acclimation in insects and that these responses can enhance tolerance of multiple stresses.

The Earths magnetic field is used as a navigational cue by many animals. For mammals, however, there is little data to show that navigation ability relies on sensing the natural magnetic field. In migratory bats, however, the calibration of a magnetic compass became plausible following experiments demonstrating a role for the solar azimuth at sunset in their orientation system. Here, we investigated how an altered magnetic field at sunset changes the nocturnal orientation of the bat Pipistrellus pygmaeus. We exposed bats to either the natural magnetic field, a horizontally shifted field (120{degrees}), or the same shifted field combined with a reversal of the natural value of inclination (70{degrees} to -70{degrees}). We later released the bats and found that the take-off orientation differed between all treatments. Bats that were exposed to the 120{degrees} shift were unimodally oriented northwards, in contrast to controls which exhibited a North-South distribution. Surprisingly, the orientation of bats exposed to both a 120{degrees}-shift and reverse inclination was indistinguishable from a uniform distribution. These results provide the missing link that these migratory bats calibrate a magnetic compass at sunset, and for the first time, they show that bats are sensitive to the angle of magnetic inclination.

Aerobic scope defines the energetic margin available for activity beyond maintenance and plays a central role in ecological performance. In many organisms, increasing body size or environmental stress can reduce this margin, a pattern often described as having "little left in the tank". However, species that rely on episodic but intense activity may require sufficient aerobic capacity from early life stages onward, particularly when individuals are relatively large at hatching or birth. Here, we examined the ontogenetic scaling of resting metabolic rate (RMR) and active metabolic rate (AMR) in the giant deep-sea scavenger Bathynomus doederleini across a broad size range (1.7-48.4 g) using intermittent-flow respirometry at 10 {degrees}C. RMR and AMR increased with body mass and exhibited nearly identical scaling exponents, resulting in a size-invariant factorial aerobic scope (median = 2.83). This pattern suggests that aerobic capacity is established early and maintained proportionally with maintenance costs, supporting locomotion and scavenging throughout ontogeny in an energy-limited environment.

Bumblebees navigate complex environments where collisions with obstacles can impair flight performance. While bees possess innate collision avoidance reflexes, they may benefit from learning to identify and avoid high-risk collision areas using environmental cues. However, the temporal dynamics of how bees form associations between visual cues and collision experiences remain unclear. We investigated whether bumblebees associate visual cues perceived before or after collision events with a movement direction. Individual foragers were trained to navigate through a flight tunnel containing a transparent barrier and an LED panel that switched colours immediately after the bees first collision. Following training, we tested bees responses to each colour cue without the barrier. Bees demonstrated significant preferences for avoiding the previously blocked side when presented with either the pre-collision colour (81% correct responses) or post-collision colour (71% correct responses), while showing no preference when no colour cue was presented. Individual analysis revealed that 61% of bees responded to both cue types, while others showed selective responses to specific temporal windows. These results demonstrate that bees can form associations with visual cues encountered in different time windows relative to negative experiences, revealing temporal flexibility in associative learning that contributes to successful navigation in cluttered environments.

Animals must tune their physical performance to changing environmental conditions, and the breadth of environmental tolerance may contribute to delineating the species geographic range. A common environmental challenge that flying animals face is the reduction of air density at high elevation and a reduction in the effectiveness of lift production that accompanies it. Turkey vultures (Cathartes aura) inhabit a >3000 m elevation range, and fly considerably higher, necessitating that they compensate for air density differences through behavior, physiology, or biomechanics. We predicted that birds flying at high elevation would demonstrate higher median flight speeds while maintaining similar glide angles. We used 3-dimensional videography to track Turkey vultures flying at three elevations and found a negative relationship between median airspeed and air density that matched our prediction. Additionally, neither the ratio of horizontal speed to sinking speed nor flapping behavior varied with air density. These results were robust to varying flight behavior (climbing vs. level flight). Finally, we derived a glide polar from the free-flying vultures and showed that they are proficient at tuning their flight speed to minimize their cost of transport during straight-line flight, but transition to a minimum power strategy during gliding turns.

The lateral line is the primary modality fish use to create a hydrodynamic image of their environment. These images contribute to a variety of behaviors, from rheotaxis to escape responses. Here we discern the contributions of visual and lateral line modalities in hunting behavior of larvae that have developed under different photic conditions. In particular, cave animals have a hypertrophied sense of mechanosensation, and we studied the common animal model cavefish Astyanax mexicanus and its closest related surface relative. We raised larvae in a diurnal light-dark regimen and in complete darkness. We then examined the distribution of neuromasts in their lateral lines, and their hunting performance in light and dark conditions, with and without the contribution of the lateral line. We report that all larva depend on the lateral line for success in hunting and that surface fish raised in the dark have a greater dependency on the lateral line,

Maintaining positional estimates of goal locations is a fundamental task for navigating animals. Diverse animal groups, including both vertebrates and invertebrates, can accomplish this through path integration (PI). During PI, navigators integrate movement changes, tracking both distance and direction, to generate a spatial estimate of their start location, or global vector, allowing efficient direct return travel without retracing the outbound route. In ants, PI is accomplished through the coupling of pedometer and celestial compass estimates. Within the PI system, it has been theorized navigators may segment the global vector into local-vectors for way-pointing. However, this is controversial, as these navigators may instead be homing via the view alignment. Here, we present evidence trail-following ants can attend to segments of their global vector to retrace their non-straight pheromone trails, without the confound of familiar views. Veromessor pergandei foragers navigate via directionally distinct segments of their PI by orienting along separate legs of their inbound route at unfamiliar locations, indicating these changes are not triggered by familiar external cues, but by the PI state. These findings contrast with the view of path integration as a singular memory estimate and underscore the systems ability to way-point to intermediate goals along the inbound route. We discuss how the foraging ecology of ant species that rely on non-straight pheromone-marked trails may support attending vector segments to remain on the pheromone rather than attempting straight-line shortcuts back to the nest.

Overhead motion in humans often leads to shoulder injuries, a consequence of the evolutionary trade-off in glenohumeral joint anatomy that balances stability with mobility. Bats consistently engage in overhead motion during flight, subjecting their shoulders to substantial loading throughout their relatively long lifespan. Remarkably, despite the demands placed on a bats shoulder, instability and rotator cuff tears, which could be fatal to bats in short order, are not observed in nature. We were thus inspired to study functional adaptations in the shoulders of bats that enable this overhead motion. Comparative anatomical studies of the shoulders of bats and mice, similarly-sized quadrupeds, were performed and interpreted using a mathematical model. Scapular anatomy indicated a more prominent role for the infraspinatus muscle in the bat compared to the mouse. Measurements of bat and mice shoulders revealed that the bat glenoid had a larger curvature and arc length than that of mice, providing a larger articulating surface area with and deeper enclosing surface of the humeral head. Modeling results predicted that the bat shoulder is stable over a dramatically larger range of angles compared to the mouse shoulder. These results suggested that adaptations to constraints imposed by the bony anatomy and rotator cuff tendons of the shoulder may contribute to the ability of bats to sustain overhead motion in a high stress, repeated loading environment without injury. Results suggest that bats have evolved unique adaptations in their glenohumeral bony anatomy that reduce stress on the supraspinatus, enhance joint stability, and optimize strength across a broad range of motion.

Centipedes coordinate body and limb flexion to generate propulsion. On flat solid surfaces, the limb-stepping patterns can be characterized according to the direction in which limbaggregates propagate, opposite to (retrograde) or with the direction of motion (direct). It is unknown how limb and body dynamics are modified in terrain with terradynamic complexity more representative of their natural heterogeneous environments. Here, we investigated how centipedes that use retrograde and direct limp-stepping patterns, S. polymorpha and S. sexspinosus, respectively, coordinate their body and limbs to navigate laboratory environments which present footstep challenges and terrain rugosity. We recorded the kinematics and measured the locomotive performance of these animals traversing two rough terrains with randomly distributed step heights and compared the kinematics to those on a flat frictional surface. S. polymorpha exhibited similar body and limb dynamics across all terrains and a decrease in speed with increased terrain roughness. Unexpectedly, when placed in a rough terrain, S. sexspinosus changed the limb-stepping pattern from direct to retrograde. Further, for both species, traversal of rough terrains was facilitated by hypothesized passive mechanics: upon horizontal collision of a limb with a block, the limb passively bent and later continued the stepping pattern. While centipedes have many degrees of freedom. our results suggest these animals negotiate limb-substrate interactions and navigate complex terrains, by offloading complex control and leveraging the innate flexibility of their limbs.

Path integration is a computational strategy that allows an animal to maintain an internal estimate of its position relative to a point of origin. Many species use path integration to navigate back to specific locations, typically their homes, after lengthy and convoluted excursions. Hymenopteran insects are impressive path integrators, directly returning to their hives after hundreds of meters of outward travel. Recent neurobiological insights have established hypotheses for how path integration may be mediated by the brains of bees, but clear ways to test these hypotheses in the laboratory are currently unavailable. Here we report that the bumblebee, Bombus terrestris, uses path integration while walking over short distances in an indoor arena. They estimate accurate vector distances after displacement and orient by artificial celestial cues. Walking bumblebees also exhibited systematic search patterns when home vectors failed to lead them accurately back to the nest, closely resembling searches performed by other species in natural conditions. We thus provide a robust experimental system to test navigation behavior in the laboratory that reflects most aspects of natural path integration. Importantly, we established this assay in an animal that is both readily available and resilient to invasive manipulations. In the future, our behavioral assay therefore can be combined with current electrophysiological techniques, opening a path towards directly probing the neural basis of the sophisticated vector navigation abilities of bees.

Force-length (F-L) and force-velocity (F-V) properties characterize skeletal muscles intrinsic properties under controlled conditions, and it is thought that these properties can inform and predict in vivo muscle function. Here, we map dynamic in vivo operating range and mechanical function during walking and running, to the measured in situ F-L and F-V characteristics of guinea fowl (Numida meleagris) lateral gastrocnemius (LG), a primary ankle extensor. We use in vivo patterns of muscle tendon force, fascicle length, and activation to test the hypothesis that muscle fascicles operate at optimal lengths and velocities to maximize force or power production during walking and running. Our findings only partly support our hypothesis: in vivo LG velocities are consistent with optimizing power during work production, and economy of force at higher loads. However, LG does not operate at lengths on the force plateau ({+/-}5% Fmax) during force production. LG length was near L0 at the time of EMG onset but shortened rapidly such that force development during stance occurred almost entirely on the ascending limb of the F-L curve, at shorter than optimal lengths. These data suggest that muscle fascicles shorten across optimal lengths in late swing, to optimize the potential for rapid force development near the swing-stance transition. This may provide resistance against unexpected perturbations that require rapid force development at foot contact. We also found evidence of passive force rise (in absence of EMG activity) in late swing, at lengths where passive force is zero in situ, suggesting that dynamic history dependent and viscoelastic effects may contribute to in vivo force development. Direct comparison of in vivo work loops and physiological operating ranges to traditional measures of F-L and F-V properties suggests the need for new approaches to characterize dynamic muscle properties in controlled conditions that more closely resemble in vivo dynamics.

ABSTRACTFeathers act as aerodynamic cantilevers, and to withstand the prolonged cyclical loading that occurs during flight, feathers must be stiff, lightweight and strong. We experimentally tested the differences in feather structure, primarily stiffness and size, between (a) wild and captive Barnacle Geese Branta leucopsis, and (b) primary feathers dropped during the annual flight feather moult, and those feathers freshly regrown during the moult process. We found that, despite having undergone a 5,000km round-trip migration, flight feathers dropped during moult in the wild geese were stiffer than those measured in the captive geese, both for those dropped during moult and those re-grown. We propose that this may be related to diet or stress in the captive geese.Competing Interest StatementThe authors have declared no competing interest.View Full Text

The insect gut, which plays a role in ion and water balance, has been shown to leak solutes in the cold. Cold stress can also activate insect immune systems, but it is unknown if the leak of the gut microbiome is a possible immune trigger in the cold. We developed a novel feeding protocol to load the gut of locusts (Locusta migratoria) with fluorescent bacteria before exposing them to -2{degrees}C for up to 48 h. No bacteria were recovered from the hemolymph of cold-exposed locusts, regardless of exposure duration. To examine this further, we used an ex vivo gut sac preparation to re-test cold-induced fluorescent FITC-dextran leak across the gut and found no increased rate of leak. These results question not only the validity of FITC-dextran as a marker of paracellular barrier permeability in the gut, but also to what extent the insect gut becomes leaky in the cold.