Seeking temporal refugia to heat stress: increasing nocturnal activity despite predation risk.

Flexibility in activity timing may enable organisms to quickly adapt to environmental changes. Under global warming, diurnally adapted endotherms may achieve a better energy balance by shifting their activity towards cooler nocturnal hours. However, this shift may expose animals to new or increased environmental challenges (e.g. increased predation risk, reduced foraging efficiency). We analysed a large dataset of activity data from 47 ibex (Capra ibex) in two protected areas, characterized by varying levels of predation risk (presence versus absence of the wolf-Canis lupus). We found that ibex increased nocturnal activity following warmer days and during brighter nights. Despite the considerable sexual dimorphism typical of this species and the consequent different predation-risk perception, males and females demonstrated consistent responses to heat in both predator-present and predator-absent areas. This supports the hypothesis that shifting activity towards nighttime may be a common strategy adopted by diurnal endotherms in response to global warming. As nowadays different pressures are pushing mammals towards nocturnality, our findings emphasize the urgent need to integrate knowledge of temporal behavioural modifications into management and conservation planning.

Regulation of ddb2 expression in blind cavefish and zebrafish reveals plasticity in the control of sunlight-induced DNA damage repair.

We have gained considerable insight into the mechanisms which recognize and repair DNA damage, but how they adapt to extreme environmental challenges remains poorly understood. Cavefish have proven to be fascinating models for exploring the evolution of DNA repair in the complete absence of UV-induced DNA damage and light. We have previously revealed that the Somalian cavefish Phreatichthys andruzzii, lacks photoreactivation repair via the loss of light, UV and ROS-induced photolyase gene transcription mediated by D-box enhancer elements. Here, we explore whether other systems repairing UV-induced DNA damage have been similarly affected in this cavefish model. By performing a comparative study using P. andruzzii and the surface-dwelling zebrafish, we provide evidence for a conservation of sunlight-regulated Nucleotide Excision Repair (NER). Specifically, the expression of the ddb2 gene which encodes a key NER recognition factor is robustly induced following exposure to light, UV and oxidative stress in both species. As in the case of the photolyase genes, D-boxes in the ddb2 promoter are sufficient to induce transcription in zebrafish. Interestingly, despite the loss of D-box-regulated photolyase gene expression in P. andruzzii, the D-box is required for ddb2 induction by visible light and oxidative stress in cavefish. However, in the cavefish ddb2 gene this D-box-mediated induction requires cooperation with an adjacent, highly conserved E2F element. Furthermore, while in zebrafish UV-induced ddb2 expression results from transcriptional activation accompanied by stabilization of the ddb2 mRNA, in P. andruzzii UV induces ddb2 expression exclusively via an increase in mRNA stability. Thus, we reveal plasticity in the transcriptional and post transcriptional mechanisms regulating the repair of sunlight-induced DNA damage under long-term environmental challenges.

Adaptive phenotypic plasticity induces individual variability along a cognitive trade-off.

Animal species, including humans, display patterns of individual variability in cognition that are difficult to explain. For instance, some individuals perform well in certain cognitive tasks but show difficulties in others. We experimentally analysed the contribution of cognitive plasticity to such variability. Theory suggests that diametrically opposed cognitive phenotypes increase individuals' fitness in environments with different conditions such as resource predictability. Therefore, if selection has generated plasticity that matches individuals' cognitive phenotypes to the environment, this might produce remarkable cognitive variability. We found that guppies, Poecilia reticulata, exposed to an environment with high resource predictability (i.e. food available at the same time and in the same location) developed enhanced learning abilities. Conversely, guppies exposed to an environment with low resource predictability (i.e. food available at a random time and location) developed enhanced cognitive flexibility and inhibitory control. These cognitive differences align along a trade-off between functions that favour the acquisition of regularities such as learning and functions that adjust behaviour to changing conditions (cognitive flexibility and inhibitory control). Therefore, adaptive cognitive plasticity in response to resource predictability (and potentially similar factors) is a key determinant of cognitive individual differences.

Interspecific differences in developmental mode determine early cognitive abilities in teleost fish.

Most studies on developmental variation in cognition have suggested that individuals are born with reduced or absent cognitive abilities, and thereafter, cognitive performance increases with age during early development. However, these studies have been mainly performed in altricial species, such as humans, in which offspring are extremely immature at birth. In this work, we tested the hypothesis that species with other developmental modes might show different patterns of cognitive development. To this end, we analysed inhibitory control performance in two teleost species with different developmental modes, the zebrafish Danio rerio and the guppy Poecilia reticulata, exploiting a simple paradigm based on spontaneous behaviour and therefore applicable to subjects of different ages. Zebrafish hatch as larvae 3 days after fertilisation, and have an immature nervous system, a situation that mirrors extreme altriciality. We found that at the early stages of development, zebrafish displayed no evidence of inhibitory control, which only begun to emerge after one month of life. Conversely, guppies, which are born after approximately one month of gestation as fully developed and independent individuals, solved the inhibitory control task since their first days of life, although performance increased with sexual maturation. Our study suggests that the typical progression described during early ontogeny in humans and other species might not be the only developmental trend for animals' cognition and that a species' developmental mode might determine variation in cognition across subjects of different age.

Embryonic exposure to artificial light at night impairs learning abilities and their covariance with behavioural traits in teleost fish.

The natural light cycle has profound effects on animals' cognitive systems. Its alteration owing to human activities, such as artificial light at night (ALAN), affects the biodiversity of mammalian and avian species by impairing their cognitive functions. The impact of ALAN on cognition, however, has not been investigated in aquatic species, in spite of the common occurrence of this pollution along water bodies. We exposed eggs of a teleost fish (the zebrafish Danio rerio) to ALAN and, upon hatching, we measured larvae' cognitive abilities with a habituation learning paradigm. Both control and ALAN-exposed larvae showed habituation learning, but the latter learned significantly slower, suggesting that under ALAN conditions, fish require many more events to acquire ecologically relevant information. We also found that individuals' learning performance significantly covaried with two behavioural traits in the control zebrafish, but ALAN disrupted one of these relationships. Additionally, ALAN resulted in an average increase in larval activity. Our results showed that both fish's cognitive abilities and related individual differences are negatively impacted by light pollution, even after a short exposure in the embryonic stage.

Modeling Sarcoglycanopathy in Danio rerio.

Sarcoglycanopathies, also known as limb girdle muscular dystrophy 3-6, are rare muscular dystrophies characterized, although heterogeneous, by high disability, with patients often wheelchair-bound by late adolescence and frequently developing respiratory and cardiac problems. These diseases are currently incurable, emphasizing the importance of effective treatment strategies and the necessity of animal models for drug screening and therapeutic verification. Using the CRISPR/Cas9 genome editing technique, we generated and characterized δ-sarcoglycan and ß-sarcoglycan knockout zebrafish lines, which presented a progressive disease phenotype that worsened from a mild larval stage to distinct myopathic features in adulthood. By subjecting the knockout larvae to a viscous swimming medium, we were able to anticipate disease onset. The δ-SG knockout line was further exploited to demonstrate that a δ-SG missense mutant is a substrate for endoplasmic reticulum-associated degradation (ERAD), indicating premature degradation due to protein folding defects. In conclusion, our study underscores the utility of zebrafish in modeling sarcoglycanopathies through either gene knockout or future knock-in techniques. These novel zebrafish lines will not only enhance our understanding of the disease's pathogenic mechanisms, but will also serve as powerful tools for phenotype-based drug screening, ultimately contributing to the development of a cure for sarcoglycanopathies.

Individual differences and knockout in zebrafish reveal similar cognitive effects of BDNF between teleosts and mammals.

The remarkable similarities in cognitive performance between teleosts and mammals suggest that the underlying cognitive mechanisms might also be similar in these two groups. We tested this hypothesis by assessing the effects of the brain-derived neurotrophic factor (BDNF), which is critical for mammalian cognitive functioning, on fish's cognitive abilities. We found that individual differences in zebrafish's learning abilities were positively correlated with bdnf expression. Moreover, a CRISPR/Cas9 mutant zebrafish line that lacks the BDNF gene (bdnf-/-) showed remarkable learning deficits. Half of the mutants failed a colour discrimination task, whereas the remaining mutants learned the task slowly, taking three times longer than control bdnf+/+ zebrafish. The mutants also took twice as long to acquire a T-maze task compared to control zebrafish and showed difficulties exerting inhibitory control. An analysis of habituation learning revealed that cognitive impairment in mutants emerges early during development, but could be rescued with a synthetic BDNF agonist. Overall, our study indicates that BDNF has a similar activational effect on cognitive performance in zebrafish and in mammals, supporting the idea that its function is conserved in vertebrates.

Medaka as a model for seasonal plasticity: Photoperiod-mediated changes in behaviour, cognition, and hormones.

Teleosts display the highest level of brain plasticity of all vertebrates. Yet we still know little about how seasonality affects fish behaviour and the underlying cognitive mechanisms since the common neurobehavioral fish models are native to tropical environments where seasonal variation is absent or reduced. The medaka, Oryzias latipes, which inhabits temperate zone habitats, represents a promising model in this context given its large phenotypic changes associated with seasonality and the possibility to induce seasonal plasticity by only manipulating photoperiod. Here, we report the first extended investigation of seasonal plasticity in medaka behaviour and cognition, as well as the potential underlying molecular mechanisms. We compared medaka exposed to summer photoperiod (16 h light:8 h dark) with medaka exposed to winter photoperiod (8 h light:16 h dark), and detected substantial differences. Medaka were more active and less social in summer photoperiod conditions, two effects that emerged in the second half of an open-field and a sociability test, respectively, and might be at least in part related to habituation to the testing apparatus. Moreover, the cognitive phenotype was significantly affected: in the early response to a social stimulus, brain functional lateralisation shifted between the two hemispheres under the two photoperiod conditions, and inhibitory and discrimination learning performance were reduced in summer conditions. Finally, the expression of genes encoding key pituitary hormones, tshß and gh, and of the tshß regulatory transcription factor tef in the brain was increased in summer photoperiod conditions. This work reveals remarkable behavioural and cognitive phenotypic plasticity in response to photoperiod in medaka, and suggests a potential regulatory role for the same hormones involved in seasonal plasticity of other vertebrates.

Size-selective mortality induces evolutionary changes in group risk-taking behaviour and the circadian system in a fish.

Intensive and trait-selective mortality of fish and wildlife can cause evolutionary changes in a range of life-history and behavioural traits. These changes might in turn alter the circadian system due to co-evolutionary mechanisms or correlated selection responses both at behavioural and molecular levels, with knock-on effects on daily physiological processes and behavioural outputs. We examined the evolutionary impact of size-selective mortality on group risk-taking behaviour and the circadian system in a model fish species. We exposed zebrafish Danio rerio to either large or small size-selective harvesting relative to a control over five generations, followed by eight generations during which harvesting was halted to remove maternal effects. Size-selective mortality affected fine-scale timing of behaviours. In particular, small size-selective mortality, typical of specialized fisheries and gape-limited predators targeting smaller size classes, increased group risk-taking behaviuor during feeding and after simulated predator attacks. Moreover, small size-selective mortality increased early peaks of daily activity as well as extended self-feeding daily activity to the photophase compared to controls. By contrast large size-selective mortality, typical of most wild capture fisheries, only showed an almost significant effect of decreasing group risk-taking behaviour during the habituation phase and no clear changes in fine-scale timing of daily behavioural rhythms compared to controls. We also found changes in the molecular circadian core clockwork in response to both size-selective mortality treatments. These changes disappeared in the clock output pathway because both size-selected lines showed similar transcription profiles. This switch downstream to the molecular circadian core clockwork also resulted in similar overall behavioural rhythms (diurnal swimming and self-feeding in the last hours of darkness) independent of the underlying molecular clock. To conclude, our experimental harvest left an asymmetrical evolutionary legacy in group risk-taking behaviour and in fine-scale daily behavioural rhythms. Yet, the overall timing of activity showed evolutionary resistance probably maintained by a molecular switch. Our experimental findings suggest that size-selective mortality can have consequences for behaviour and physiological processes.

Behavioural effects of early-life exposure to parabens in zebrafish larvae.

Parabens are classified as endocrine disrupting chemicals due to their ability to activate several nuclear receptors causing changes in hormones-dependent signalling pathways. Central nervous system of developing organisms is particularly vulnerable to changes in hormonal pathways, which could lead to altered brain function, abnormal behaviour and even diseases later in life. The aim of the present study was to investigate the effects of exposure to butylparaben (BuP), ethylparaben (EtP) and methylparaben (MeP) during early development on nervous system using zebrafish larvae's behavioural models. Zebrafish were exposed until 4 days post fertilization (dpf) to three concentrations of each paraben chosen considering the environmentally realistic concentrations of human exposure and the benchmark-dose lower bound calculated for zebrafish larvae (BuP: 5, 50 and 500 µg/L; EtP: 50, 500 and 5000 µg/L; MeP: 100, 1000 and 10,000 µg/L). Activity in novel and in familiar environment, thigmotaxis, visual startle response and photic synchronization of the behavioural circadian rhythms were analysed at 4, 5 and 6 dpf. Zebrafish larvae exposed to BuP 500 µg/L and EtP 5000 µg/L revealed increased anxiety-like behaviour in novel environment. Larvae treated with 500 µg/L of BuP showed reduced activity in familiar and marginally in unfamiliar environment, and larvae exposed to 5000 µg/L of EtP exhibited hyperactivity in familiar environment. Parabens exposure did not influence the visual startle response and the photic synchronization of circadian rhythms in zebrafish larvae. This research highlighted as the exposure to parabens has the potential to interfere with behavioural development of zebrafish.

Guppies show sex and individual differences in the ability to inhibit behaviour.

In humans, individual and sex differences have been long reported for several cognitive tasks and are at least in part due to variability in the function that inhibits behaviour (i.e. inhibitory control). Similar evidence of individual and sex differences in inhibitory abilities is also present in other vertebrates, but is scarce outside primates. Experiments on reversal learning, which requires inhibiting behaviours, suggest that this variability may exist in a teleost fish, the guppy, Poecilia reticulata. We tested this hypothesis by observing guppies in an inhibitory task. Guppies were exposed to unreachable prey inside a transparent tube for six trials. Guppies showed a marked reduction in the number of attempts to catch the prey within the first trial and also over repeated trials. We found a striking sex difference in the capacity to inhibit foraging behaviour. Males attempted to attack the prey twice as often as females and showed negligible improvement over trials. Irrespective of sex, individuals remarkably differed in their performance, with some guppies being systematically more skilled than others across the repeated trials. These results confirm that individual and sex differences in the ability to inhibit behaviour are not restricted to humans and other primates, suggesting that they might be widespread among vertebrates. Variability in inhibitory ability provides an explanation for emerging records of variability in other cognitive tasks in fish.

Lateralization correlates with individual differences in inhibitory control in zebrafish.

Individual fitness often depends on the ability to inhibit behaviours not adapted to a given situation. However, inhibitory control can vary greatly between individuals of the same species. We investigated a mechanism that might maintain this variability in zebrafish (Danio rerio). We demonstrate that inhibitory control correlates with cerebral lateralization, the tendency to process information with one brain hemisphere or the other. Individuals that preferentially observed a social stimulus with the right eye and thus processed social information with the left brain hemisphere, inhibited foraging behaviour more efficiently. Therefore, selective pressures that maintain lateralization variability in populations might provide indirect selection for variability in inhibitory control. Our study suggests that individual cognitive differences may result from complex multi-trait selection mechanisms.

Photoperiod Manipulation Affects Transcriptional Profile of Genes Related to Lipid Metabolism and Apoptosis in Zebrafish (Danio rerio) Larvae: Potential Roles of Gut Microbiota.

Gut microbiota plays a fundamental role in maintaining host's health by controlling a wide range of physiological processes. Administration of probiotics and manipulation of photoperiod have been suggested as modulators of microbial composition and are currently undergoing an extensive research in aquaculture as a way to improve health and quality of harvested fish. However, our understanding regarding their effects on physiological processes is still limited. In the present study we investigated whether manipulation of photoperiod and/or probiotic administration was able to alter microbial composition in zebrafish larvae at hatching stage. Our findings show that probiotic does not elicit effects while photoperiod manipulation has a significant impact on microbiota composition. Moreover, we successfully predicted lipid biosynthesis and apoptosis to be modulated by microbial communities undergoing continuous darkness. Interestingly, expression levels of caspase 3 gene (casp3) and lipid-related genes (hnf4a, npc1l1, pparγ, srebf1, agpat4 and fitm2) were found to be significantly overexpressed in dark-exposed larvae, suggesting an increase in the occurrence of apoptotic processes and a lipid metabolism impairment, respectively (p < 0.05). Our results provide the evidence that microbial communities in zebrafish at early life stages are not modulated by a short administration of probiotics and highlight the significant effect that dark photoperiod elicits on zebrafish microbiota and potentially on health.

Inhibitory control in zebrafish, Danio rerio.

We assessed whether zebrafish, Danio rerio, display inhibitory control using a simple and rapid behavioural test. Zebrafish were exposed to a prey stimulus placed inside a transparent tube, which initially elicited attack behaviour. However, zebrafish showed a rapid reduction in the number of attacks towards the prey, which indicated the ability to inhibit their foraging behaviour. Zebrafish also exhibited mnemonic retention of foraging inhibition, as indicated by a reduced number of attacks in a subsequent exposure to the unreachable prey. The ability to inhibit the foraging behaviour varied across three genetically separated wild-type strains and across different individuals within strains, suggesting that zebrafish show heritable within-species differences in inhibitory control. Our behavioural test might be suitable for screening large zebrafish populations in mutational studies and assessing the effects of pharmacologically active substances on inhibitory control.

Cavefish eye loss in response to an early block in retinal differentiation progression.

The troglomorphic phenotype shared by diverse cave-dwelling animals is regarded as a classical example of convergent evolution. One unresolved question is whether the characteristic eye loss in diverse cave species is based on interference with the same genetic program. Phreatichthys andruzzii, a Somalian cavefish, has evolved under constant conditions in complete darkness and shows severe troglomorphic characteristics, such as complete loss of eyes, pigments and scales. During early embryonic development, a complete eye is formed but is subsequently lost. In Astyanax mexicanus, another blind cavefish, eye loss has been attributed to interference during eye field patterning. To address whether similar pathways have been targeted by evolution independently, we investigated the retinal development of P. andruzzii, studying the expression of marker genes involved in eye patterning, morphogenesis, differentiation and maintenance. In contrast to Astyanax, patterning of the eye field and evagination of the optic vesicles proceeds without obvious deviation. However, the subsequent differentiation of retinal cell types is arrested during generation of the first-born cell type, retinal ganglion cells, which also fail to project correctly to the optic tectum. Eye degeneration in both species is driven by progressive apoptosis. However, it is retinal apoptosis in Phreatichthys that progresses in a wave-like manner and eliminates progenitor cells that fail to differentiate, in contrast to Astyanax, where lens apoptosis appears to serve as a driving force. Thus, evolution has targeted late retinal differentiation events, indicating that there are several ways to discontinue the development and maintenance of an eye.

Normalization of disrupted clock gene expression in males with tetraplegia: a crossover randomized placebo-controlled trial of melatonin supplementation.

STUDY DESIGN: Crossover double blind, randomized placebo-controlled trial. OBJECTIVES: Circadian oscillators are located both in the brain and in peripheral organs. Melatonin, the main brain-derived hormone governing circadian variations, is highly associated with daylight patterns. However, in subjects with tetraplegia the melatonin levels are blunted. Here we studied peripheral oscillators in peripheral blood mononuclear cells (PBMCs) in males with tetraplegia by examining how exogenous melatonin may influence the expression of clock gene mRNAs. SETTING: Sunnaas Rehabilitation Hospital, Nesoddtangen, Norway. METHODS: Six males with tetraplegia received 2 mg of melatonin or placebo 4 days before the study period. We also included six able-bodied men sleeping or kept awake during the night. Plasma samples were collected four times during a 24-h period. The mRNA expression levels of the clock genes PER1, PER2, BMAL1, and REV-ERBα were quantified in PBMCs using quantitative RT-PCR. RESULTS: The mRNA expression levels of PER-1 and -2 and REV-ERBα were increased at 04:00 h compared with the able-bodied controls (p < 0.05). Melatonin supplementation changed mRNA peak-time toward the time of supplementation. CONCLUSIONS: Several peripheral clock genes displayed distorted expression levels in tetraplegia. Supplementation with melatonin changed the mRNA expression levels of these genes toward those observed among able-bodied. SPONSORSHIP: Financial support was provided from the Throne Holst Foundation, Sunnaas Rehabilitation hospital and the University of Ferrara (FAR2016).

Quantitative abilities in a reptile (Podarcis sicula).

The ability to identify the largest amount of prey available is fundamental for optimizing foraging behaviour in several species. To date, this cognitive skill has been observed in all vertebrate groups except reptiles. In this study we investigated the spontaneous ability of ruin lizards to select the larger amount of food items. In Experiment 1, lizards proved able to select the larger food item when presented with two alternatives differing in size (0.25, 0.50, 0.67 and 0.75 ratio). In Experiment 2 lizards presented with two groups of food items (1 versus 4, 2 versus 4, 2 versus 3 and 3 versus 4 items) were unable to select the larger group in any contrast. The lack of discrimination in the presence of multiple items represents an exception in numerical cognition studies, raising the question as to whether reptiles' quantitative abilities are different from those of other vertebrate groups.

Fly cryptochrome and the visual system.

Cryptochromes are flavoproteins, structurally and evolutionarily related to photolyases, that are involved in the development, magnetoreception, and temporal organization of a variety of organisms. Drosophila CRYPTOCHROME (dCRY) is involved in light synchronization of the master circadian clock, and its C terminus plays an important role in modulating light sensitivity and activity of the protein. The activation of dCRY by light requires a conformational change, but it has been suggested that activation could be mediated also by specific "regulators" that bind the C terminus of the protein. This C-terminal region harbors several protein-protein interaction motifs, likely relevant for signal transduction regulation. Here, we show that some functional linear motifs are evolutionarily conserved in the C terminus of cryptochromes and that class III PDZ-binding sites are selectively maintained in animals. A coimmunoprecipitation assay followed by mass spectrometry analysis revealed that dCRY interacts with Retinal Degeneration A (RDGA) and with Neither Inactivation Nor Afterpotential C (NINAC) proteins. Both proteins belong to a multiprotein complex (the Signalplex) that includes visual-signaling molecules. Using bioinformatic and molecular approaches, dCRY was found to interact with Neither Inactivation Nor Afterpotential C through Inactivation No Afterpotential D (INAD) in a light-dependent manner and that the CRY-Inactivation No Afterpotential D interaction is mediated by specific domains of the two proteins and involves the CRY C terminus. Moreover, an impairment of the visual behavior was observed in fly mutants for dCRY, indicative of a role, direct or indirect, for this photoreceptor in fly vision.

A blind circadian clock in cavefish reveals that opsins mediate peripheral clock photoreception.

The circadian clock is synchronized with the day-night cycle primarily by light. Fish represent fascinating models for deciphering the light input pathway to the vertebrate clock since fish cell clocks are regulated by direct light exposure. Here we have performed a comparative, functional analysis of the circadian clock involving the zebrafish that is normally exposed to the day-night cycle and a cavefish species that has evolved in perpetual darkness. Our results reveal that the cavefish retains a food-entrainable clock that oscillates with an infradian period. Importantly, however, this clock is not regulated by light. This comparative study pinpoints the two extra-retinal photoreceptors Melanopsin (Opn4m2) and TMT-opsin as essential upstream elements of the peripheral clock light input pathway.

Cryptochrome expression in the eye of migratory birds depends on their migratory status.

Most passerine birds are nocturnal migrants. When kept in captivity during the migratory periods, these species show a migratory restlessness, or Zugunruhe. Recent studies on Sylvia warblers have shown that Zugunruhe is an excellent proxy of migratory disposition. Passerine birds can use the Earth's geomagnetic field as a compass to keep their course during their migratory flight. Among the candidate magnetoreceptive mechanisms are the cryptochromes, flavoproteins located in the retina that are supposed to perceive the magnetic field through a light-mediated process. Previous work has suggested that expression of Cryptochrome 1 (Cry1) is increased in migratory birds compared with non-migratory species. Here we tested the hypothesis that Cry1 expression depends on migratory status. Blackcaps Sylvia atricapilla were caught before fall migration and held in registration cages. When the birds were showing robust Zugunruhe, we applied a food deprivation protocol that simulates a long migratory flight. When the birds were refed after 2 days, their Zugunruhe decreased substantially, as is expected from birds that would interrupt migration for a refuelling stopover. We found that Cry1 expression was higher at night than during daytime in birds showing Zugunruhe, whereas in birds that underwent the fasting-and-refeeding protocol and reduced their levels of Zugunruhe, night Cry1 expression decreased to daytime levels. Our work shows that Cry1 expression is dependent on the presence of Zugunruhe and not on species-specific or seasonal factors, or on the birds being active versus inactive. These results support the hypothesis that cryptochromes underlie magnetoreceptive mechanisms in birds.