Tameness and stress physiology in a predator-naive island species confronted with novel predation threat

Tame behaviour, i.e. low wariness, in terrestrial island species is often attributed to low predation pressure. However, we know little about its physiological control and its flexibility in the face of predator introductions. Marine iguanas (Amblyrhynchus cristatus) on the Galapagos Islands are a good model to study the physiological correlates of low wariness. They have lived virtually without predation for 5-15 Myr until some populations were first confronted with feral cats and dogs some 150 years ago. We tested whether and to what extent marine iguanas can adjust their behaviour and endocrine stress response to novel predation threats. Here, we show that a corticosterone stress response to experimental chasing is absent in naive animals, but is quickly restored with experience. Initially, low wariness also increases with experience, but remains an order of magnitude too low to allow successful escape from introduced predators. Our data suggest that the ability of marine iguanas to cope with predator introductions is limited by narrow reaction norms for behavioural wariness rather than by constraints in the underlying physiological stress system. In general, we predict that island endemics show flexible physiological stress responses but are restricted by narrow behavioural plasticity.     

Frequency of moulting by shore crabs Carcinus maenas (L.) changes their colour and their success in mating and physiological performance

Male shore crabs, Carcinus maenas [Linnaeus, 1758], compete aggressively for access to receptive females to mate. Size is the single most important factor for the outcome of these conflicts, large males with carapace width (CW) over 60 mm being much more likely to gain access to receptive females than smaller males. To compete aggressively, large male shore crabs decrease moulting frequencies and may potentially terminate moulting to enter a state of anecdysis, in which further growth is suspended to increase reproductive output. This change from a “growth” strategy to a “reproduction” strategy results in the creation of two morphs, which can roughly be separated by their colouration. As the new exoskeleton created during moult is always green in appearance, crabs tend to be green in appearance during periods where they grow rapidly and moult frequently. Green crabs are found in all size classes. However, as the exoskeleton becomes older, the colour gradually changes to a darker red colour, and large crabs that have spent an extended period in intermoult are therefore often red in appearance. Also, the exoskeleton of red crabs exhibits a higher incidence of epibionts and wear and tear. Red crabs can also be found in all size classes, but their relative proportion in the population increases dramatically in size classes above 60-mm CW. Size for size, the red morph has a thicker carapace and larger master chelae than the green morph. Also, the reproductive indices (RI) for red crabs are higher than for green crabs, and they experience higher mating success. However, this mating success appears to be achieved at the expense of a lower physiological tolerance, green crabs being better adapted to deal with changes in the surrounding environment. This increased tolerance is not only observed with regard to natural variations in the habitats where shore crabs live, but green crabs also appear to be more tolerant to variations caused by anthropogenic pollution. Consequently, the shift from growth to reproduction exerts a profound effect on the behaviour, physiology and ecotoxicology of male shore crabs. The present paper reviews the studies conducted so far, proposes a mechanism by which some of these differences between the two morphs are created and discusses their ecological and ecotoxicological significance.     

Sexual selection and the physiological consequences of habitat choice by a fiddler crab

In mid-Atlantic salt marshes, reproductively active male sand fiddler crabs, Uca pugilator, use a single greatly enlarged major claw as both a weapon to defend specialized breeding burrows from other males and an ornament to attract females for mating. During the summer breeding season, females strongly prefer to mate with males controlling burrows in open areas high on the shore. Food availability decreases while temperature and desiccation stress increase with increasing shore height, suggesting that the timing and location of fiddler crab mating activity may result in a potential trade-off between reproductive success and physiological condition for male crabs. We compared thermal preferences in laboratory choice experiments to body temperatures of models and living crabs in the field and found that from the perspective of a fiddler crab, the thermal environment of the mating area is quite harsh relative to other marsh microhabitats. High temperatures significantly constrained fiddler crab activity on the marsh surface, a disadvantage heightened by strongly reduced food availability in the breeding area. Nevertheless, when the chance of successfully acquiring a mate was high, males accepted a higher body temperature (and concomitantly higher metabolic and water loss rates) than when the chances of mating were low. Likewise, experimentally lowering costs by adding food and reducing thermal stress in situ increased fiddler crab waving display levels significantly. Our data suggest that fiddler crabs can mitigate potential life history trade-offs by tuning their behavior in response to the magnitude of both energetic and non-energetic costs and benefits.     

Stress reduces hemolymph ecdysteroid levels in the crab: mediation by the eyestalks

In decapod crustaceans, molt hormone (ecdysone) production by Y-organs is suppressed by an eyestalk neurosecretory product, molt-inhibiting hormone (MIH). Environmental stressors are known to delay or prevent molting in crabs. The present study assesses the function of the MIH-Y-organ neuroendocrine system in the crab Cancer antennarius under conditions of daily handling stress. After three days, stressed crabs showed significant suppression of hemolymph ecdysteroid levels, which continued to fall to 20% of controls by day 14. Ecdysteroid titers of stressed crabs returned to prestress levels seven days after stress termination. Ecdysteroid levels in de-eyestalked (DES) crabs rose 160% within 48 hr post-DES. Stressing DES crabs over 16 subsequent days did not significantly alter ecdysteroid levels compared with unstressed DES controls. Handling stress thus depresses hemolymph ecdysteroid levels in the crab, a response that is mediated by eyestalks and appears to result from stress-induced MIH release.     

Density affects mating mode and large male mating advantage in a fiddler crab

Fiddler crabs show two different mating modes: either females search and crabs mate underground in male burrows, or males search and crabs mate on the surface near female burrows. We explored the relationship between crab density, body size, the searching behavior of both sexes, and the occurrence of both mating modes in the fiddler crab Uca uruguayensis. We found that crabs change their mating mode depending on their size and crab density. Crabs mated mostly on the surface at low densities, and underground at high densities. The proportion of wandering receptive females but not courting males accounted for the variation in mating modes. This suggests that whether crabs mate underground (or on the surface) is determined by the presence (or absence) of searching females. We found that the change in the mating mode affected the level of assortative mating; males mating underground were bigger than those mating on the surface, suggesting active female choice. Given that fiddler crabs experience multiple reproductive cycles, they are prone to showing behavioral plasticity in their mating strategy whenever the payoffs of using different mating modes differ between reproductive events. Our results suggest that the incorporation of different levels of environmental variability may be important in theoretical models aimed at improving our understanding of the evolution of alternative mating tactics and strategies.     

Thermal biology of horseshoe crab embryos and larvae: A role for heat shock proteins

Eggs of the American horseshoe crab, Limulus polyphemus L., develop on sandy estuarine beaches during the spring and summer, and are potentially vulnerable to thermal stress during the 3-4 weeks of development to the first instar (trilobite) larval stage. In many marine taxa, heat shock (stress) proteins (Hsp's) help individuals acclimate to stresses by restoring the proper folding of cellular proteins whose shape has been altered by temperature shock or other forms of environmental stress. We examined the survival of embryos and first instar (trilobite) larvae following heat shock, and compared the levels of Hsp70 in heat shocked and control animals. Animals acclimated to 13 or 22 °C had close to 100% survival when heat shocked for 3 h at 35 or 40 °C, but exposure to 45 °C for 3 h was lethal. To study the effect of heat shock on Hsp70 production under environmentally realistic conditions, animals were acclimated to either 13 or 22 °C, heat-shocked at 35 °C for 3 h, and soluble proteins were extracted following 0, 2, 4, or 6 h recovery at 22 °C. The relative amounts of Hsp70 in horseshoe crab embryos and larvae were examined using SDS-PAGE and Western blotting. Relative to controls animals held at a constant temperature, there was a slight elevation of Hsp70 only among heat shocked trilobite larvae in the 6 h recovery treatment. Hsp70 levels did not differ significantly between control and heat shocked embryos. Horseshoe crabs have adapted to living in a thermally stressful environment by maintaining a high baseline (constitutive) level of cellular stress proteins such as Hsp70, rather than by synthesizing inducible Hsp's when stressful temperatures are encountered. This may be an effective strategy given that the heat shocks encountered by intertidal embryos and larvae occur regularly as a function of diurnal and tidal temperature changes.     

How does an increase in minimum daily temperatures during incubation influence reproduction in the great tit Parus major?

Temperature variation affects all life stages of organisms, especially early development, and considering global warming, it is urgent to understand precisely its consequences. In egg‐laying species, incubation behaviour can buffer embryo developmental temperature variation and influence offspring development. We experimentally investigated the effect of an increase in minimum daily nest temperature during incubation in the great tit Parus major, by placing a hand warming pad under the nest in the evenings. As compared to controls, the experimental treatment increased nest temperature at night by an average of 4°C, and this increase carried over to the following day. We measured the consequences of this mainly nocturnal temperature increase during incubation on 1) parental behaviour (incubation and nestling feeding), 2) parental health (quantified by body condition, immune status, physiological and oxidative stress) and 3) reproductive success (nestling body condition, growth, i.e. mass gain, hatching and fledging success, and nestling immune status, physiological and oxidative stress). This study yielded three major results. First, we found that heating the nest did not change the duration of incubation as compared to controls. Second, increasing nest temperature during incubation decreased nestling feeding behaviour but did not affect parental health in terms of body condition, immune status, physiological and oxidative stress. Third, nestling mass at hatching was greater but nestling mass gain was slower in heated nests than in control nests, resulting in similar fledging mass. The present study demonstrates that increased environmental temperatures during incubation influenced nestling development in the great tit and especially hatchling mass, which might produce long‐term life history consequences.     

Sex-specific mediation of foraging in the shore crab, Carcinus maenas

Experiments were conducted to investigate the sex-specific differences to feeding responses of the shore crab Carcinus maenas throughout the year. Results demonstrate that female shore crabs exhibit stronger feeding responses than males throughout the year with a significantly reduced feeding response in males during the summer months' reproductive season. We also studied the possible function(s) of the moulting hormone, 20-hydroxyecdysone (Crustecdysone) that has been described as a potential female-produced sex pheromone to initiate male reproductive behaviour in a number of crustaceans. We recently presented evidence that for shore crabs this is not the case and now show that the steroid is instead functioning as a sex-specific feeding deterrent protecting the moulting 'soft' female crabs. Whilst male shore crabs were deterred from prey (Mytilus edulis) and synthetic feeding stimulants glycine and taurine when these feeding stimulants were spiked with crustecdysone, intermoult female crabs were significantly less affected and rarely deterred from feeding. This sex specificity of the moulting hormone, in combination with the female sex pheromone, which has no anti-feeding properties, ensures that male crabs mate with soft-shelled, moulted females rather than engage in cannibalism, such as found frequently in cases when soft-shelled females are exposed to intermoult females.     

Heart rate and gill ventilation in ovigerous and non‐ovigerous edible crabs,Cancer pagurus: The effects of disturbance,substrate and starvation

This study compared the cardiac and ventilatory behaviour of disturbed and settled (48 h recovery) ovigerous Cancer pagurus with the behaviour of non‐ovigerous crabs. It also examined the effects of starvation and a sand substrate on ovigerous females. Ovigerous crabs had significantly lower heart rates than non‐ovigerous crabs. This implies that they have a reduced metabolic rate, confirmed by an earlier study which described lower rates of oxygen uptake in ovigerous crabs. Scaphognathite beat frequency was unaffected by either the presence of eggs, starvation or a sand substrate, but the amplitude of ventilatory pressure changes was higher in settled ovigerous females, implying greater force. The duration of periods of reversed ventilation was also substantially extended in ovigerous crabs, and disturbance caused a ten‐fold increase in the frequency of these reversals, resulting in ventilation being carried out in a predominantly reversed direction. It is suggested that these respiratory adaptations serve to ventilate the egg mass. Disturbance also led to a greater elevation of heart rate in ovigerous crabs, indicating that they are more prone to handling stress. Fed ovigerous crabs spent a higher percentage of time ventilating unilaterally; this response was lost following starvation. Starved ovigerous crabs had the lowest mean heart rate, highest mean ventilatory pressures and, when disturbed above a sand substrate, they showed the highest frequency of reversals of ventilation, possibly in association with their efforts to bury their abdomen bearing the egg mass.     

No evidence of morphine analgesia to noxious shock in the shore crab, Carcinus maenas

A number of criteria have been suggested for testing if pain occurs in animals, and these include an analgesic effect of opiates (Bateson, 1991). Morphine reduces responses to noxious stimuli in crustaceans but also reduces responsiveness in a non-pain context. Here we use a paradigm in which shore crabs receive a shock in a preferred dark shelter but not if they remain in an unpreferred light area. Analgesia should thus enhance movement to the preferred dark area because they should not experience ‘pain’. However, morphine inhibits rather than enhances this movement even when no shock is given. Morphine produces a general effect of non-responsiveness rather than a specific analgesic effect and this could also explain previous studies claiming analgesia. However, we question the utility of this criterion of pain and suggest instead that behavioural criteria be employed.     

Brittle-star bioluminescence functions as an aposematic signal to deter crustacean predators

The behaviour of three species of nocturnally active, visually orienting crabs (Portunus sebae, P. spinnimanus and P. ordwayi) was observed to determine whether the luminescent signals produced by Ophiopsila riisei (Echinodermata: Ophiuroidea) function as an aposematic deterrent against crustacean predators. In repeated experimental trials, crabs showed more rapid rejection of luminescent unpalatable ophiuroids than of non-luminescent controls. After five trials, unpalatable luminescent prey were rejected three times as quickly as either unpalatable or palatable non-luminescent controls. Over the course of the trials, crabs damaged fewer luminescent ophiuroids than non-luminescent controls. Furthermore, blind crabs (that could not perceive the luminescent flashes) caused significantly more damage to luminescent brittle-stars than did crabs with intact eyes (that could see the light signals). All three species of crab ophiuroids versus non-luminescent controls. Ophiopsila riisei survived the majority of crab attacks, and crabs can therefore learn to reject unpalatable luminescent prey without killing them. This suggests that individual selection may be an important mechanism for the evolution of aposematic luminescent signals in this, and probably other, luminescent species.     

Thermal tolerance of the crab Pachygrapsus marmoratus: intraspecific differences at a physiological (CTMax) and molecular level (Hsp70)

Temperature is one of the most important variables influencing organisms, especially in the intertidal zone. This work aimed to test physiological and molecular intraspecific differences in thermal tolerance of the crab Pachygrapsus marmoratus (Fabricius, 1787). The comparisons made focused on sex, size, and habitat (estuary and coast) differences. The physiological parameter was upper thermal limit, tested via the critical thermal maximum (CTMax) and the molecular parameter was total heat shock protein 70 (Hsp70 and Hsp70 plus Hsc70) production, quantified via an enzyme-linked imunosorbent assay. Results showed that CTMax values and Hsp70 production are higher in females probably due to different microhabitat use and potentially due to different hormonal regulation in males and females. Among females, non-reproducing ones showed a higher CTMax value, but no differences were found in Hsp70, even though reproducing females showed higher variability in Hsp70 amounts. As reproduction takes up a lot of energy, its allocation for other activities, including stress responses, is lower. Juveniles also showed higher CTMax and Hsp70 expression because they occur in greater shore heights and ageing leads to alterations in protein synthesis. Comparing estuarine and coastal crabs, no differences were found in CTMax but coastal crabs produce more Hsp70 than estuarine crabs because they occur in drier and hotter areas than estuarine ones, which occur in moister environments. This work shows the importance of addressing intraspecific differences in the stress response at different organizational levels. This study shows that these differences are key factors in stress research, climate research, and environmental monitoring.     

Individual quality and personality: bolder males are less fecund in the hermit crab Pagurus bernhardus

One explanation for animal personality is that different behavioural types derive from different life-history strategies. Highly productive individuals, with high growth rates and high fecundity, are assumed to live life at a fast pace showing high levels of boldness and risk taking, compared with less productive individuals. Here, we investigate among-individual differences in mean boldness (the inverse of the latency to recover from a startling stimulus) and in the consistency of boldness, in male hermit crabs in relation to two aspects of life-history investment. We assessed aerobic scope by measuring the concentration of the respiratory pigment haemocyanin, and we assessed fecundity by measuring spermatophore size. First, we found that individuals investing in large spermatophores also had high concentrations of haemocyanin. Using doubly hierarchical-generalized linear models to analyse longitudinal data on startle responses, we show that hermit crabs vary both in their mean response durations and in the consistency of their behaviour. Individual consistency was unrelated to haemocyanin concentration or spermatophore size, but mean startle response duration increased with spermatophore size. Thus, counter to expectations, it was the most risk-averse individuals, rather than the boldest and most risk prone, that were the most productive. We suggest that similar patterns should be present in other species, if the most productive individuals avoid risky behaviour.     

Cold tolerance of the invasive Carcinus maenas in the east Pacific: molecular mechanisms and implications for range expansion in a changing climate

Physiological studies have long been utilized to understand the role of environmental temperature in the distribution of native organisms within marine communities. For the invasive crab Carcinus maenas, temperature has been implicated as the main predictor of establishment success across temperate regions. Therefore, we determined whether the lower temperature tolerances of this non-native crab would restrict it from spreading farther poleward from a relatively new recipient environment. Cold tolerance capacity was determined in the laboratory by holding crabs sampled from Vancouver Island, British Columbia (BC)—near the present northern limit for the northeast Pacific metapopulation to an overwintering thermal profile generated from Sitka, Alaska, USA. These crabs were physiologically capable of overwintering north of their present range boundary. The cellular response to cold stress was investigated using two functional categories of the cellular stress response. We measured cyclin D1, a cell-cycle regulator, and Hsp70, a protein chaperone, after laboratory acclimation and acute cold stress on two populations of C. maenas from the west coast of North America that have disparate thermal histories (crabs sampled from CA or BC). We found site-specific differential expression of cyclin D1 after cold acclimation and cold shock, perhaps affecting invasion capacity in this species. Determining what physiological mechanisms are in place with respect to thermal tolerance and preference can give insight into what makes an invasive organism successful and aid in predicting probable distribution of such species within a new environment.     

Pre-natal stress amplifies the immediate behavioural responses to acute pain in piglets

Pre-natal stress (PNS) or undernutrition can have numerous effects on an individual''s biology throughout their lifetime. Some of these effects may be adaptive by allowing individuals to tailor their phenotype to environmental conditions. Here we investigated, in the domestic pig Sus scrofa, whether one possible consequence of a predicted adverse environment could be altered pain perception. The behavioural response of piglets to the surgical amputation (‘docking’) of their tail or a sham procedure was measured for 1 min in piglets born to mothers who either experienced mid-gestation social stress or were left undisturbed throughout pregnancy. A behavioural pain score was found to predict the docked status of piglets with high discriminant accuracy. Piglets exposed to PNS had a significantly higher pain score than controls, and for each litter of tail-docked piglets, the average pain score was correlated with mid-gestation maternal cortisol levels. The data presented here provide evidence that the experience of stress in utero can result in a heightened acute response to injury in early life. Speculatively, this may represent an adaptive alteration occurring as a consequence of a pre-natal ‘early warning’ of environmental adversity.     

Genomic approaches to detecting thermal stress in Calanus finmarchicus (Copepoda: Calanoida)

Zooplankton may be subjected to physiological stress as they encounter rapid and large changes in temperature through vertical migration or transfer into different water masses. Induction of one or more heat shock proteins (hsp) is a common protective response to thermal stress in organisms. We looked for evidence for such a response in Calanus finmarchicus. We compared hsp70 expression in copepods exposed to temperature stress with that for non-stressed controls. Partial sequences of the amplified cDNA product were obtained and aligned with known hsp70 sequences to establish the identity of the heat shock protein. In one experiment, animals were transferred from their collection temperature (8 °C) to 20 °C for 30 min, and then returned to 8 °C for 4 h before sampling for gene expression levels. In another, the animals were exposed to 18 °C over 48 h before sampling for molecular analysis. A four-fold induction of hsp70 was measured in both groups of heat shocked animals using quantitative real time polymerase chain reaction (PCR). The experimental temperatures, although high for C. finmarchicus, are within the range of temperatures experienced by this species in their habitat. In addition to confirming an hsp70-mediated response in C. finmarchicus, the findings suggest that a recent history of thermal stress may be assessed in natural populations through a routine molecular assay.     

Training dogs with help of the shock collar: short and long term behavioural effects

Behavioural effects of the use of a shock collar during guard dog training of German shepherd dogs were studied. Direct reactions of 32 dogs to 107 shocks showed reactions (lowering of body posture, high pitched yelps, barks and squeals, avoidance, redirection aggression, tongue flicking) that suggest stress or fear and pain. Most of these immediate reactions lasted only a fraction of a second. The behaviour of 16 dogs that had received shocks in the recent past (S-dogs) was compared with the behaviour of 15 control dogs that had received similar training but never had received shocks (C-dogs) in order to investigate possible effects of a longer duration. Only training sessions were used in which no shocks were delivered and the behaviour of the dogs (position of body, tail and ears, and stress-, pain- and aggression-related behaviours) was recorded in a way that enabled comparison between the groups. During free walking on the training grounds S-dogs showed a lower ear posture and more stress-related behaviours than C-dogs. During obedience training and during manwork (i.e. excercises with a would-be criminal) the same differences were found. Even a comparison between the behaviour of C-dogs with that of S-dogs during free walking and obedience exercises in a park showed similar differences. Differences between the two groups of dogs existed in spite of the fact that C-dogs also were trained in a fairly harsh way. A comparison between the behaviour during free walking with that during obedience exercises and manwork, showed that during training more stress signals were shown and ear positions were lower. The conclusions, therefore are, that being trained is stressful, that receiving shocks is a painful experience to dogs, and that the S-dogs evidently have learned that the presence of their owner (or his commands) announces reception of shocks, even outside of the normal training context. This suggests that the welfare of these shocked dogs is at stake, at least in the presence of their owner.     

Hydrostatic pressure is like high temperature and oxidative stress in the damage it causes to yeast

Abstract A comparison of barotolerance, thermotolerance and oxygen tolerance was made under different physiological conditions, such as heat shocked and recovered state, different growth phases and changes of physiological conditions by mutations. The three kinds of tolerance showed similar features under different physiological conditions. We suggest that the damage caused by hydrostatic pressure may be essentially the same as that due to high temperature and oxidative stress in yeast.     

Copper exposure reduces production of red carotenoids in a marine copepod

Sub-lethal exposure to copper has been shown to modulate both mitochondrial function and antioxidant gene expression in zooplankton. To date, however, researchers have not identified a quantifiable phenotypic trait that reliably indicates such physiological responses to copper exposure. Red ketocarotenoids are abundant in marine zooplankton serving both physiological and coloration roles, and their production is sensitive to environmental stress. In this study the expression of mitochondrial gene cytochrome c oxidase I (COI) and antioxidant gene glutathione reductase (GR), and the production of red ketocarotenoid, astaxanthin, was measured in response to sub-lethal copper exposure. We found that mRNA of COI and GR was more abundant in copper-exposed copepods than controls, suggesting there was a physiological response to copper exposure. At the same time, copper-exposed copepods produced less astaxanthin than controls. We suggest that ketocarotenoid content of zooplankton has the potential to be a sensitive bioindicator of marine environmental pollution. Understanding how cellular responses to environmental stressors manifest in the phenotypes of marine animals will greatly increase our capacity to monitor marine ecosystem health.