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Biological rhythms that are mediated by exogenous factors, such as light and temperature, drive the physiology of organisms and affect processes ranging from cellular to population levels. For elasmobranchs (i.e. sharks, rays, and skates), studies documenting diel activity and movement patterns indicate that many species are crepuscular or nocturnal in nature. However, few studies have investigated the rhythmicity of elasmobranch physiology to understand the mechanisms underpinning these distinct patterns. Here, we assess diel patterns of metabolic rates in a small meso-predator, the epaulette shark ( Hemiscyllium ocellatum), across ecologically relevant temperatures and upon acutely removing photoperiod cues. This species possibly demonstrates behavioral sleep during daytime hours, which is supported herein by low metabolic rates during the day and a 1.7-fold increase in metabolic rates at night. From spring to summer seasons, where average average water temperature temperatures for this species range 24.5 to 28.5 °C, time of day, and not temperature, had the strongest influence on metabolic rate. These results indicate that this species, and perhaps other similar species from tropical and coastal environments, may have physiological mechanisms in place to maintain metabolic rate on a seasonal time scale regardless of temperature fluctuations that are relevant to their native habitats.

Accelerative manoeuvres, such as fast-starts, are crucial for fish to avoid predation. Escape responses are fast-starts that include fundamental survival traits for prey that experience high predation pressure. However, no previous study has assessed escape performance in neonate tropical sharks. We quantitatively evaluated vulnerability traits of neonate tropical sharks by testing predictions on their fast-start escape performance. We predicted (1) high manoeuvrability, given their high flexibility, but (2) low propulsive locomotion owing to the drag costs associated with pectoral fin extension during escape responses. Further, based on previous work on dogfish, Squalus suckleyi, we predicted (3) long reaction times (as latencies longer than teleosts, >20 ms). We used two-dimensional, high-speed videography analysis of mechano-acoustically stimulated neonate blacktip reef shark, Carcharhinus melanopterus (n=12), and sicklefin lemon shark, Negaprion acutidens (n=8). Both species performed a characteristic C-start double-bend response (i.e. two body bends), but single-bend responses were only observed in N. acutidens. As predicted, neonate sharks showed high manoeuvrability with high turning rates and tight turning radii (3–11% of body length) but low propulsive performance (i.e. speed, acceleration and velocity) when compared with similar-sized teleosts and S. suckleyi. Contrary to expectations, escape latencies were

Ocean acidification (OA) is postulated to affect the physiology, behavior, and life-history of marine species, but potential for acclimation or adaptation to elevated pCO2 in wild populations remains largely untested. We measured brain transcriptomes of six coral reef fish species at a natural volcanic CO2 seep and an adjacent control reef in Papua New Guinea. We show that elevated pCO2 induced common molecular responses related to circadian rhythm and immune system but different magnitudes of molecular response across the six species. Notably, elevated transcriptional plasticity was associated with core circadian genes affecting the regulation of intracellular pH and neural activity in Acanthochromis polyacanthus. Gene expression patterns were reversible in this species as evidenced upon reduction of CO2 following a natural storm-event. Compared with other species, Ac. polyacanthus has a more rapid evolutionary rate and more positively selected genes in key functions under the influence of elevated CO2, thus fueling increased transcriptional plasticity. Our study reveals the basis to variable gene expression changes across species, with some species possessing evolved molecular toolkits to cope with future OA.

Highly variable thermal environments, such as coral reef flats, are challenging for marine ectotherms and are thought to invoke the use of behavioural strategies to avoid extreme temperatures and seek out thermal environments close to their preferred temperatures. Common to coral reef flats, the epaulette shark (Hemiscyllium ocellatum) possesses physiological adaptations to hypoxic and hypercapnic conditions, such as those experienced on reef flats, but little is known regarding the thermal strategies used by these sharks. We investigated whether H. ocellatum uses behavioural thermoregulation (i.e., movement to occupy thermally favourable microhabitats) or tolerates the broad range of temperatures experienced on the reef flat. Using an automated shuttlebox system, we determined the preferred temperature of H. ocellatum under controlled laboratory conditions and then compared this preferred temperature to 6 months of in situ environmental and body temperatures of individual H. ocellatum across the Heron Island reef flat. The preferred temperature of H. ocellatum under controlled conditions was 20.7 +/- 1.5 degrees C, but the body temperatures of individual H. ocellatum on the Heron Island reef flat mirrored environmental temperatures regardless of season or month. Despite substantial temporal variation in temperature on the Heron Island reef flat (15-34 degrees C during 2017), there was a lack of spatial variation in temperature across the reef flat between sites or microhabitats. This limited spatial variation in temperature creates a low-quality thermal habitat limiting the ability of H. ocellatum to behaviourally thermoregulate. Behavioural thermoregulation is assumed in many shark species, but it appears that H. ocellatum may utilize other physiological strategies to cope with extreme temperature fluctuations on coral reef flats. While H. ocellatum appears to be able to tolerate acute exposure to temperatures well outside of their preferred temperature, it is unclear how this, and other, species will cope as temperatures continue to rise and approach their critical thermal limits. Understanding how species will respond to continued warming and the strategies they may use will be key to predicting future populations and assemblages.

Many species with broad distributions are exposed to different thermal regimes which often select for varied phenotypes. This intraspecific variation is often overlooked but may be critical in dictating the vulnerability of different populations to environmental change. We reared Port Jackson shark (Heterodontus portusjacksoni) eggs from two thermally discrete populations (i.e. Jervis Bay and Adelaide) under each location's present-day mean temperatures, predicted end-of-century temperatures and under reciprocal-cross conditions to establish intraspecific thermal sensitivity. Rearing temperatures strongly influenced ṀO2 Max and critical thermal limits, regardless of population, indicative of acclimation processes. However, there were significant population-level effects, such that Jervis Bay sharks, regardless of rearing temperature, did not exhibit differences in ṀO2 Rest , but under elevated temperatures exhibited reduced maximum swimming activity with step-wise increases in temperature. In contrast, Adelaide sharks reared under elevated temperatures doubled their ṀO2 Rest , relative to their present-day temperature counterparts; however, maximum swimming activity was not influenced. With respect to reciprocal-cross comparisons, few differences were detected between Jervis Bay and Adelaide sharks reared under ambient Jervis Bay temperatures. Similarly, juveniles (from both populations) reared under Adelaide conditions had similar thermal limits and swimming activity (maximum volitional velocity and distance) to each other, indicative of conserved acclimation capacity. However, under Adelaide temperatures, the ṀO2 Rest of Jervis Bay sharks was greater than that of Adelaide sharks. This indicates that the energetics of cooler water population (Adelaide) is likely more thermally sensitive than that of the warmer population (Jervis Bay). While unique to elasmobranchs, these data provide further support that by treating species as static, homogeneous populations, we ignore the impacts of thermal history and intraspecific variation on thermal sensitivity. With climate change, intraspecific variation will manifest as populations move, demographics change or extirpations occur, starting with the most sensitive populations.

The gill is one of the most important organs for growth and survival of fishes. Early life stages in coral reef fishes often exhibit extreme physiological and demographic characteristics that are linked to well-established respiratory and ionoregulatory processes. However, gill development and function in coral reef fishes is not well understood. Therefore, we investigated gill morphology, oxygen uptake and ionoregulatory systems throughout embryogenesis in two coral reef damselfishes, Acanthochromis polyacanthus and Amphiprion melanopus (Pomacentridae). In both species, we found key gill structures to develop rapidly early in the embryonic phase. Ionoregulatory cells appear on gill filaments 3–4 days post-fertilization and increase in density, whilst disappearing or shrinking in cutaneous locations. Primary respiratory tissue (lamellae) appears 5–7 days post-fertilization, coinciding with a peak in oxygen uptake rates of the developing embryos. Oxygen uptake was unaffected by phenylhydrazine across all ages (pre-hatching), indicating that haemoglobin is not yet required for oxygen uptake. This suggests that gills have limited contribution to respiratory functions during embryonic development, at least until hatching. Rapid gill development in damselfishes, when compared with that in most previously investigated fishes, may reflect preparations for a high-performance, challenging lifestyle on tropical reefs, but may also make reef fishes more vulnerable to anthropogenic stressors.

Teleost fishes are diverse and successful, comprising almost half of all extant vertebrate species. It has been suggested that their success as a group is related, in part, to their unique O2 transport system, which includes pH-sensitive hemoglobin, a red blood cell β-adrenergic Na+/H+ exchanger (RBC β-NHE) that protects red blood cell pH, and plasma accessible carbonic anhydrase which is absent at the gills but present in some tissues, that short-circuits the β-NHE to enhance O2 unloading during periods of stress. However, direct support for this has only been examined in a few species of salmonids. Here, we expand the knowledge of this system to two warm-water, highly active marine percomorph fish, cobia (Rachycentron canadum) and mahi-mahi (Coryphaena hippurus). We show evidence for RBC β-NHE activity in both species, and characterize the Hb-O2 transport system in one of those species, cobia. We found significant RBC swelling following β-adrenergic stimulation in both species, providing evidence for the presence of a rapid, active RBC β-NHE in both cobia and mahi-mahi, with a time-course similar to that of salmonids. We generated oxygen equilibrium curves (OECs) for cobia blood and determined the P50, Hill, and Bohr coefficients, and used these data to model the potential for enhanced O2 unloading. We determined that there was potential for up to a 61% increase in O2 unloading associated with RBC β-NHE short-circuiting, assuming a - 0.2 ∆pHa-v in the blood. Thus, despite phylogenetic and life history differences between cobia and the salmonids, we found few differences between their Hb-O2 transport systems, suggesting conservation of this physiological trait across diverse teleost taxa.

In teleost fishes, catecholamine-induced increases in the activity of cation exchangers compensate for decreases in hemoglobin oxygen affinity and maximum blood oxygen carrying capacity caused by decreases in plasma pH (i.e., metabolic acidosis). The resultant red blood cell (RBC) swelling has been documented in sandbar (Carcharhinus plumbeus) and epaulette (Hemiscyllium ocellatum) sharks following capture by rod-and-reel or after a 1.5 h exposure to anoxia (respectively), although the underlying mechanisms remain unknown. To determine if RBC swelling could be documented in other elasmobranch fishes, we collected blood samples from clearnose skate (Rostroraja eglanteria), blacktip reef shark (Carcharhinus melanopterus), and sicklefin lemon shark (Negaprion acutidens) subjected to exhaustive exercise or air exposure (or both) and measured hematocrit, hemoglobin concentration, RBC count, RBC volume, and mean corpuscular hemoglobin content. We did likewise with sandbar and epaulette sharks to further explore the mechanisms driving swelling when present. We could not document RBC swelling in any species; although hematocrit increased in all species (presumably due to RBC ejection from the spleen or fluid shifts out of the vascular compartment) except epaulette shark. Our results indicate RBC swelling and associated ion shifts in elasmobranch fishes is not inducible by exercise or hypoxia, thus implying this response maybe of lesser importance for maintaining oxygen delivery during acute acidosis than in teleost fishes.

Temporal environmental variability causes behavioural and physiological responses in organisms that can affect their spatial location in time, and ultimately drive changes in population and community dynamics. Linking ecological changes with underlying environmental drivers is a complex task that can however be facilitated through the integration of physiology. Our overarching aim was to investigate the association between physiological performance and habitat utilisation patterns modulated by short temporal fluctuations in environmental factors. We used in situ monitoring data from a system experiencing extreme environmental fluctuations over a few hours and we selected four fish species with different habitat utilisation patterns across dissolved oxygen (DO) fluctuations: two commonly observed species (Siganus lineatus and Acanthopagrus pacificus), including at low DO (40 and 50% saturation, respectively), and two reef species (Heniochus acuminatus and Chaetodon vagabundus) never recorded below 70% saturation. We hypothesised that these patterns were associated to species' physiological performance in hypoxia. Therefore, we measured different metabolic variables (O2crit, incipient lethal oxygen (ILO), time to ILO, index of cumulative ambient oxygen deficit (O2deficit), maximum oxygen supply capacity (α)) using respirometry. Physiological performance differed among species and was intrinsically associated to habitat use patterns. S. lineatus had a lower O2crit than H. acuminatus, A. pacificus and C. vagabundus (13, 18.7, 20 and 20.2% saturation respectively). Additionally, S. lineatus and A. pacificus displayed better capacity for survival below O2crit than C. vagabundus and H. acuminatus (lower ILO, higher O2deficit and longer time to ILO) and higher α. Field monitoring data revealed that DO temporarily falls below species’ O2crit and even ILO on most days, suggesting that short temporal variability in DO likely forces species to temporarily avoid harmful conditions, driving important changes in ecosystem structure over a few hours. Our results support the hypothesis that organismal physiology can provide insights into ecological changes occurring over a few hours as a result of environmental variability. Consequently, integrating physiology with ecological data at relevant temporal scales may help predict temporal shifts in ecosystems structure and functions to account for ecological patterns often overlooked and difficult to identify.

Temperature has a profound effect on all life and a particularly influential effect on ectotherms, such as fishes. Amphibious fishes have a variety of strategies, both physiological and/or behavioural, to cope with a broad range of thermal conditions. This study examined the relationship between prolonged (5 weeks) exposure to a range of temperatures (22, 25, 28, or 32 degrees C) on oxygen uptake rate and movement behaviours (i.e., thermoregulation and emergence) in a common amphibious fish, the barred mudskipper (Periophthalmus argentilneatuis). At the highest temperature examined (32 degrees C, approximately 5 degrees C above their summer average temperatures), barred mudskippers exhibited 33.7-97.7% greater oxygen uptake rates at rest ((M) over dotO(2Rest)), emerged at a higher temperature (CTe; i.e., a modified critical thermal maxima (CTMax) methodology) of 41.3 +/- 0.3 degrees C relative to those maintained at 28, 25, or 22 degrees C. The 32 degrees C-maintained fish also ceased movement activity at the highest holding temperature suggesting that prolonged submergence at elevated temperatures is physiologically and energetically stressful to the individual. Using exhaustive exercise protocols with and without air exposure to simulate a predatory chase, the time to recovery was examined for all individuals. When submerged, mudskippers required 2.5x longer recovery time to return to resting oxygen uptake from exhaustive exercise than those fully emerged in air. Oxygen uptake data revealed that air exposure did not accrue oxygen debt, thereby allowing faster return to resting oxygen consumption rates. If the option to emerge was not available, mudskippers preferentially sought more benign water temperatures (26.7 +/- 2.1 degrees C), resembling those experienced by these fish during the Austral autumn, regardless of prolonged exposure higher or lower temperatures. These results add to our understanding of the strategies that amphibious fishes may use to mitigate extra costs associated with living in warm waters, and could be the key to understanding how such species will cope with increasing temperatures in the future.

Climate change is affecting thermal regimes globally, and organisms relying on their environment to regulate biological processes face unknown consequences. In ectotherms, temperature affects development rates, body condition, and performance. Embryonic stages may be the most vulnerable life history stages, especially for oviparous species already living at the warm edge of their distribution, as embryos cannot relocate during this developmental window. We reared 27 epaulette shark (Hemiscyllium ocellatum) embryos under average summer conditions (27 °C) or temperatures predicted for the middle and end of the twenty-first century with climate change (i.e., 29 and 31 °C) and tracked growth, development, and metabolic costs both in ovo and upon hatch. Rearing sharks at 31 °C impacted embryonic growth, yolk consumption, and metabolic rates. Upon hatch, 31 °C-reared sharks weighed significantly less than their 27 °C-reared counterparts and exhibited reduced metabolic performance. Many important growth and development traits in this species may peak after 27 °C and start to become negatively impacted nearing 31 °C. We hypothesize that 31 °C approximates the pejus temperature (i.e., temperatures at which performance of a trait begin to decline) for this species, which is alarming, given that this temperature range is well within ocean warming scenarios predicted for this species’ distribution over the next century.

As climate-driven heat waves become more frequent and intense, there is increasing urgency to understand how thermally sensitive species are responding. Acute heating events lasting days to months may elicit acclimation responses to improve performance and survival. However, the coordination of acclimation responses remains largely unknown for most stenothermal species. We documented the chronology of 18 metabolic and cardiorespiratory changes that occur in the gills, blood, spleen, and muscles when tropical coral reef fishes are thermally stressed (+3.0°C above ambient). Using representative coral reef fishes (Caesio cuning and Cheilodipterus quinquelineatus) separated by >100 million years of evolution and with stark differences in major life-history characteristics (i.e. lifespan, habitat use, mobility, etc.), we show that exposure duration illicited coordinated responses in 13 tissue and organ systems over 5 weeks. The onset and duration of biomarker responses differed between species, with C. cuning – an active, mobile species – initiating acclimation responses to unavoidable thermal stress within the first week of heat exposure; conversely, C. quinquelineatus – a sessile, territorial species – exhibited comparatively reduced acclimation responses that were delayed through time. Seven biomarkers, including red muscle citrate synthase and lactate dehydrogenase activities, blood glucose and hemoglobin concentrations, spleen somatic index, and gill lamellar perimeter and width, proved critical in evaluating acclimation progression and completion, as these provided consistent evaluation of thermal responses across species.

Temperature and oxygen limit the distribution of marine ectotherms. Haematological traits underlying blood-oxygen carrying capacity are thought to be correlated with thermal tolerance in certain fishes, and this relationship is hypothesised to be explained by oxygen supply capacity. We tested this hypothesis using reef shark neonates as experimental models because they live near their upper thermal limits and are physiologically sensitive to low oxygen conditions. We first described in situ associations between temperature and oxygen at the study site (Moorea, French Polynesia) and found that the habitats for reef shark neonates (Carcharhinus melanopterus and Negaprion acutidens) were hyperoxic at the maximum recorded temperatures. Next, we tested for in situ associations between thermal habitat characteristics and haematological traits of neonates. Contrary to predictions, we only demonstrated a negative association between haemoglobin concentration and maximum habitat temperatures in C. melanopterus. Next, we tested for ex situ associations between critical thermal maximum (CTMax) and haematological traits, but only demonstrated a negative association between haematocrit and CTMax in C. melanopterus. Finally, we measured critical oxygen tension (pcrit) ex situ and estimated its temperature sensitivity to predict oxygen-dependent values of CTMax. Estimated temperature sensitivity of pcrit was similar to reported values for sharks and skates, and predicted values for CTMax equalled maximum habitat temperatures. These data demonstrate unique associations between haematological traits and thermal tolerance in a reef shark that are likely not explained by oxygen supply capacity. However, a relationship between oxygen supply capacity and thermal tolerance remains to be demonstrated empirically.

Ocean warming and acidification act concurrently on marine ectotherms with the potential for detrimental, synergistic effects; yet, effects of these stressors remain understudied in large predatory fishes, including sharks. We tested for behavioural and physiological responses of blacktip reef shark (Carcharhinus melanopterus) neonates to climate change relevant changes in temperature (28 and 31 °C) and carbon dioxide partial pressures (pCO2; 650 and 1050 µatm) using a fully factorial design. Behavioural assays (lateralisation, activity level) were conducted upon 7–13 days of acclimation, and physiological assays (hypoxia tolerance, oxygen uptake rates, acid–base and haematological status) were conducted upon 14–17 days of acclimation. Haematocrit was higher in sharks acclimated to 31 °C than to 28 °C. Significant treatment effects were also detected for blood lactate and minimum oxygen uptake rate; although, these observations were not supported by adequate statistical power. Inter-individual variability was considerable for all measured traits, except for haematocrit. Moving forward, studies on similarly ‘hard-to-study’ species may account for large inter-individual variability by increasing replication, testing larger, yet ecologically relevant, differences in temperature and pCO2, and reducing measurement error. Robust experimental studies on elasmobranchs are critical to meaningfully assess the threat of global change stressors in these data-deficient species.

Ocean acidification, resulting from increasing atmospheric carbon dioxide (CO

Molecular responses of fishes to a heatwave are species specific and do not concur with measures of oxygen uptake in captivity., The marine heatwave of 2016 was one of the longest and hottest thermal anomalies recorded on the Great Barrier Reef, influencing multiple species of marine ectotherms, including coral reef fishes. There is a gap in our understanding of what the physiological consequences of heatwaves in wild fish populations are. Thus, in this study, we used liver transcriptomes to understand the molecular response of five species to the 2016 heatwave conditions. Gene expression was species specific, yet we detected overlap in functional responses associated with thermal stress previously reported in experimental setups. The molecular response was also influenced by the duration of exposure to elevated temperatures. This study highlights the importance of considering the effects of extreme warming events when evaluating the consequences of climate change on fish communities.

Increasing ocean temperatures and the resulting poleward range shifts of species has highlighted the importance of a species preferred temperature and thermal range in shaping ecological communities. Understanding the temperatures preferred and avoided by individual species, and how these are influenced by species interactions is critical in predicting the future trajectories of populations, assemblages, and ecosystems. Using an automated shuttlebox system, we established the preferred temperature and upper and lower threshold temperatures (i.e., avoided temperatures) of a common coral reef fish, the black-axil chromis, Chromis atripectoralis. We then investigated how the presence of conspecifics, heterospecifics (Neopomacentrus bankieri), or a predator (Cephalopholis spiloparaea) influenced the selection of these temperatures. Control C. atripectoralis preferred 27.5 ± 1.0 °C, with individuals avoiding temperatures below 23.5 ± 0.9 °C and above 29.7 ± 0.7 °C. When associating with either conspecifics or heterospecifics, C. atripectoralis selected significantly lower temperatures (conspecifics: preferred = 21.2 ± 1.4 °C, lower threshold = 18.1 ± 0.8 °C; heterospecifics: preferred = 21.1 ± 1.1 °C, lower threshold = 19.2 ± 0.9 °C), but not higher temperatures (conspecifics: preferred = 28.9 ± 1.2 °C, upper threshold = 30.8 ± 0.9 °C; heterospecifics: preferred = 29.7 ± 1.1 °C, upper threshold = 31.4 ± 0.8 °C). The presence of the predator, however, had a significant effect on both lower and upper thresholds. Individual C. atripectoralis exposed themselves to temperatures ~ 5.5 °C cooler or warmer (lower threshold: 18.6 ± 0.5 °C, upper threshold: 35.2 ± 0.5 °C) than control fish before moving into the chamber containing the predator. These findings demonstrate how behavioural responses due to species interactions influence the thermal ecology of a tropical reef fish; however, there appears to be limited scope for individuals to tolerate higher temperatures unless faced with the risk of predation.

Marine ectotherms are often sensitive to thermal stress, and certain life stages can be particularly vulnerable (e.g., larvae or spawners). In this study, we investigated the critical thermal maxima (CTmax) of larval and early juvenile life stages of three tropical marine fishes (Acanthochromis polyacanthus, Amphiprion melanopus, and Lates calcarifer). We tested for potential effects of developmental acclimation, life stage, and experimental heating rates, and we measured metabolic enzyme activities from aerobic (citrate synthase, CS) and anaerobic pathways (lactate dehydrogenase, LDH). A slightly elevated rearing temperature neither influenced CTmax nor CS activity, which otherwise could have indicated thermal acclimation. However, we found CTmax to either remain stable (Acanthrochromis polyacanthus) or increase with body mass during early ontogeny (Amphiprion melanopus and Lates calcarifer). In all three species, faster heating rates lead to higher CTmax. Acute temperature stress did not change CS or LDH activities, suggesting that overall aerobic and anaerobic metabolism remained stable. Lates calcarifer, a catadromous species that migrates from oceanic to riverine habitats upon metamorphosis, had higher CTmax than the two coral reef fish species. We highlight that, for obtaining conservative estimates of a fish species’ upper thermal limits, several developmental stages and body mass ranges should be examined. Moreover, upper thermal limits should be assessed using standardized heating rates. This will not only benefit comparative approaches but also aid in assessing geographic (re-) distributions and climate change sensitivity of marine fishes.

Thermal dependence of growth and metabolism can influence thermal preference and tolerance in marine ectotherms, including threatened and data-deficient species. Here, we quantified the thermal dependence of physiological performance in neonates of a tropical shark species (blacktip reef shark

Parasites can account for a substantial proportion of the biomass in marine communities. As such, parasites play a significant ecological role in ecosystem functioning via host interactions. Unlike macropredators, such as large piscivores, micropredators rarely cause direct mortality. Rather, micropredators impose an energetic tax, thus significantly affecting host physiology and behaviour via such sublethal effects. Recent research suggests that infection by gnathiid isopods (Crustacea) causes significant physiological stress and increased mortality rates. However, it is unclear whether infection causes changes in the behaviours that underpin escape responses or changes in routine activity levels. Moreover, it is poorly understood whether the cost of gnathiid infection manifests as an increase in cortisol. To investigate this, we examined the effect of experimental gnathiid infection on the swimming and escape performance of a newly settled coral reef fish and whether infection would lead to increased cortisol levels. We found that micropredation by a single gnathiid caused fast-start escape performance and swimming behaviour to significantly decrease and cortisol levels to double. Fast-start escape performance is an important predictor of recruit survival in the wild. As such, altered fitness related traits and short-term stress, perhaps especially during early life stages, may result in large scale changes in the number of fish that successfully recruit to adult populations.

Elevated CO

The impacts of ocean acidification will depend on the ability of marine organisms to tolerate, acclimate and eventually adapt to changes in ocean chemistry. Here, we use a unique transgenerational experiment to determine the molecular response of a coral reef fish to short-term, developmental and transgenerational exposure to elevated CO2, and to test how these responses are influenced by variations in tolerance to elevated CO2 exhibited by the parents. Within-generation responses in gene expression to end-of-century predicted CO2 levels indicate that a self-amplifying cycle in GABAergic neurotransmission is triggered, explaining previously reported neurological and behavioural impairments. Furthermore, epigenetic regulator genes exhibited a within-generation specific response, but with some divergence due to parental phenotype. Importantly, we find that altered gene expression for the majority of within-generation responses returns to baseline levels following parental exposure to elevated CO2 conditions. Our results show that both parental variation in tolerance and cross-generation exposure to elevated CO2 are crucial factors in determining the response of reef fish to changing ocean chemistry.

Acquiring and storing energy is vital to sharks of all age-classes. Viviparous shark embryos receive endogenous maternal energy reserves to sustain the first weeks after birth. Then, in order to maintain body condition, sharks must start foraging. Our goal was to understand whether maternal energy investments vary between blacktip reef sharks (Carcharhinus melanopterus) from two populations and to what extent body condition and the initiation of foraging might be affected by presumably variable maternal investments. A total of 546 young sharks were captured at St. Joseph atoll (Seychelles) and Moorea (French Polynesia) between 2014 and 2018, and indices of body condition and percentage of stomachs containing prey were measured. Maternal investment was found to be site-specific, with significantly larger, heavier, and better conditioned individuals in Moorea. Despite these advantages, as time progressed, Moorea sharks exhibited significant decreases in body condition and were slower to initiate foraging. We suggest that the young sharks’ foraging success is independent of the quality of maternal energy resources, and that other factors, such as prey availability, prey quality, and/or anthropogenic stressors are likely responsible for the observed differences across sites. Insights into intraspecific variations in early life-stages may further support site-specific management strategies for young sharks from nearshore habitats.

Many studies have examined the average effects of ocean acidification and warming on phenotypic traits of reef fishes, finding variable, but often negative effects on behavioural and physiological performance. Yet the presence and nature of a relationship between these traits is unknown. A negative relationship between phenotypic traits could limit individual performance and even the capacity of populations to adapt to climate change. Here, we examined the relationship between behavioural and physiological performance of a juvenile reef fish under elevated CO2 and temperature in a full factorial design. Behaviourally, the response to an alarm odour was negatively affected by elevated CO2, but not elevated temperature. Physiologically, aerobic scope was significantly diminished under elevated temperature, but not under elevated CO2. At the individual level, there was no relationship between behavioural and physiological traits in the control and single-stressor treatments. However, a statistically significant negative relationship was detected between the traits in the combined elevated CO2 and temperature treatment. Our results demonstrate that trade-offs in performance between behavioural and physiological traits may only be evident when multiple climate change stressors are considered, and suggest that this negative relationship could limit adaptive potential to climate change.

Increasing temperatures are expected to significantly affect the physiological performance of ectotherms, particularly in tropical locations. The shape of an organism's thermal reaction norm can provide important information on its capacity to persist under climate change scenarios; however, difficulty lies in choosing a measurable trait that best depicts physiological performance. This study investigated the effects of elevated temperatures on processes related to oxygen uptake and delivery, including oxygen consumption, haematology, and tissue health for a low-latitude population of coral reef damselfish. Acanthochromis polyacanthus were collected from the Torres Strait (10°31–46′S, 142°20–35′E) and maintained at current average ocean temperatures (+0 °C; seasonally cycling), + 1.5 °C and + 3 °C higher than present day temperatures for 10 months. Aerobic performance indicated a limit to metabolic function at + 3 °C (33 °C), following an increase in aerobic capacity at + 1.5 °C (31.5 °C). Neither haematological parameters nor gill morphology showed the same improvement in performance at + 1.5 °C. Gill histopathology provided the first indicator of a decline in organism health, which corresponded with mortality observations from previous research. Findings from this study suggest thermal specialisation in this low-latitude population as well as variation in thermal sensitivity, depending on the physiological trait.

Sharks have maintained a key role in marine food webs for 400 million years and across varying physicochemical contexts, suggesting plasticity to environmental change. In this study, we investigated the biochemical effects of ocean acidification (OA) levels predicted for 2100 (pCO(2) ~ 900 μatm) on newly hatched tropical whitespotted bamboo sharks (Chiloscyllium plagiosum). Specifically, we measured lipid, protein, and DNA damage levels, as well as changes in the activity of antioxidant enzymes and non-enzymatic ROS scavengers in juvenile sharks exposed to elevated CO(2) for 50 days following hatching. Moreover, we also assessed the secondary oxidative stress response, i.e., heat shock response and ubiquitin levels. Newly hatched sharks appear to cope with OA-related stress through a range of tissue-specific biochemical strategies, specifically through the action of antioxidant enzymatic compounds. Our findings suggest that ROS-scavenging molecules, rather than complex enzymatic proteins, provide an effective defense mechanism in dealing with OA-elicited ROS formation. We argue that sharks’ ancient antioxidant system, strongly based on non-enzymatic antioxidants (e.g., urea), may provide them with resilience towards OA, potentially beyond the tolerance of more recently evolved species, i.e., teleosts. Nevertheless, previous research has provided evidence of detrimental effects of OA (interacting with other climate-related stressors) on some aspects of shark biology. Moreover, given that long-term acclimation and adaptive potential to rapid environmental changes are yet experimentally unaccounted for, future research is warranted to accurately predict shark physiological performance under future ocean conditions. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12192-018-0892-3) contains supplementary material, which is available to authorized users.

Reduced water quality, in particular increases in suspended sediments, has been linked to declines in fish abundance on coral reefs. Changes in gill structure induced by suspended sediments have been hypothesized to impair gill function and may provide a mechanistic basis for the observed declines; yet, evidence for this is lacking. We exposed juveniles of three reef fish species ( Amphiprion melanopus , Amphiprion percula and Acanthochromis polyacanthus ) to suspended sediments (0–180 mg l −1 ) for 7 days and examined changes in gill structure and metabolic performance (i.e. oxygen consumption). Exposure to suspended sediments led to shorter gill lamellae in A. melanopus and A. polyacanthus and reduced oxygen diffusion distances in all three species. While A. melanopus exhibited impaired oxygen uptake after suspended sediment exposure, i.e. decreased maximum and increased resting oxygen consumption rates resulting in decreased aerobic scope, the oxygen consumption rates of the other two species remained unaffected. These findings imply that species sensitive to changes in gill structure such as A. melanopus may decline in abundance as reefs become more turbid, whereas species that are able to maintain metabolic performance despite suspended sediment exposure, such as A. polyacanthus or A. percula , may be able to persist or gain a competitive advantage.

Equatorial populations of marine species are predicted to be most impacted by global warming because they could be adapted to a narrow range of temperatures in their local environment. We investigated the thermal range at which aerobic metabolic performance is optimum in equatorial populations of coral reef fish in northern Papua New Guinea. Four species of damselfishes and two species of cardinal fishes were held for 14 days at 29, 31, 33, and 34 °C, which incorporated their existing thermal range (29-31 °C) as well as projected increases in ocean surface temperatures of up to 3 °C by the end of this century. Resting and maximum oxygen consumption rates were measured for each species at each temperature and used to calculate the thermal reaction norm of aerobic scope. Our results indicate that one of the six species, Chromis atripectoralis, is already living above its thermal optimum of 29 °C. The other five species appeared to be living close to their thermal optima (ca. 31 °C). Aerobic scope was significantly reduced in all species, and approached zero for two species at 3 °C above current-day temperatures. One species was unable to survive even short-term exposure to 34 °C. Our results indicate that low-latitude reef fish populations are living close to their thermal optima and may be more sensitive to ocean warming than higher-latitude populations. Even relatively small temperature increases (2-3 °C) could result in population declines and potentially redistribution of equatorial species to higher latitudes if adaptation cannot keep pace.

Ocean surface CO2 levels are increasing in line with rising atmospheric CO2 and could exceed 900μatm by year 2100, with extremes above 2000μatm in some coastal habitats. The imminent increase in ocean pCO2 is predicted to have negative consequences for marine fishes, including reduced aerobic performance, but variability among species could be expected. Understanding interspecific responses to ocean acidification is important for predicting the consequences of ocean acidification on communities and ecosystems. In the present study, the effects of exposure to near-future seawater CO2 (860μatm) on resting (M˙ O2rest) and maximum (M˙O2max) oxygen consumption rates were determined for three tropical coral reef fish species interlinked through predator-prey relationships: juvenile Pomacentrus moluccensis and Pomacentrus amboinensis, and one of their predators: adult Pseudochromis fuscus. Contrary to predictions, one of the prey species, P. amboinensis, displayed a 28-39% increase in M˙O2max after both an acute and four-day exposure to near-future CO2 seawater, while maintaining M˙O2rest. By contrast, the same treatment had no significant effects on M˙O2rest or M˙O2max of the other two species. However, acute exposure of P. amboinensis to 1400 and 2400μatm CO2 resulted in M˙O2max returning to control values. Overall, the findings suggest that: (1) the metabolic costs of living in a near-future CO2 seawater environment were insignificant for the species examined at rest; (2) the M˙O2max response of tropical reef species to near-future CO2 seawater can be dependent on the severity of external hypercapnia; and (3) near-future ocean pCO2 may not be detrimental to aerobic scope of all fish species and it may even augment aerobic scope of some species. The present results also highlight that close phylogenetic relatedness and living in the same environment, does not necessarily imply similar physiological responses to near-future CO2.

Neurosensory and behavioural disruptions are some of the most consistently reported responses upon exposure to ocean acidification-relevant CO2 levels, especially in coral reef fishes. The underlying cause of these disruptions is thought to be altered current across the GABAA receptor in neuronal cells due to changes in ion gradients (HCO3− and/or Cl−) that occur in the body following compensation for elevated ambient CO2. Despite these widely-documented behavioural disruptions, the present study is the first to pair a behavioural assay with measurements of relevant intracellular and extracellular acid-base parameters in a coral reef fish exposed to elevated CO2. Spiny damselfish (Acanthochromis polyacanthus) exposed to 1900 μatm CO2 for 4 days exhibited significantly increased intracellular and extracellular HCO3− concentrations and elevated brain pHi compared to control fish, providing evidence of CO2 compensation. As expected, high CO2 exposed damselfish spent significantly more time in a chemical alarm cue (CAC) than control fish, supporting a potential link between behavioural disruption and CO2 compensation. Using HCO3− measurements from the damselfish, the reversal potential for GABAA (EGABA) was calculated, illustrating that biophysical properties of the brain during CO2 compensation could change GABAA receptor function and account for the behavioural disturbances noted during exposure to elevated CO2.

Previous studies hailed thermal tolerance and the capacity for organisms to acclimate and adapt as the primary pathways for species survival under climate change. Here we challenge this theory. Over the past decade more than 365 tropical stenothermal fish species have been documented moving pole-ward, away from ocean warming hotspots where temperatures 2-3 °C above long-term annual means can compromise critical physiological processes. We examined the capacity of a model species - a thermally-sensitive coral reef fish, Chromis viridis (Pomacentridae) – to use preference behaviour to regulate its body temperature. Movement could potentially circumvent the physiological stress response associated with elevated temperatures and may be a strategy relied upon before genetic adaptation can be effectuated. Individuals were maintained at one of six temperatures (23, 25, 27, 29, 31 and 33 °C) for at least six weeks. We compared the relative importance of acclimation temperature to changes in upper critical thermal limits, aerobic metabolic scope, and thermal preference. While acclimation temperature positively affected the upper critical thermal limit, neither aerobic metabolic scope nor thermal preference exhibited such plasticity. Importantly, when given the choice to stay in a habitat reflecting their acclimation temperatures or relocate, fish acclimated to end-of-century predicted temperatures (i.e., 31 or 33 °C) preferentially sought out cooler temperatures, those equivalent to long-term summer averages in their natural habitats (~29 °C). This was also the temperature providing the greatest aerobic metabolic scope and body condition across all treatments. Consequently, acclimation can confer plasticity in some performance traits, but may be an unreliable indicator of the ultimate survival and distribution of mobile stenothermal species under global warming. Conversely, thermal preference can arise long before, and remain long after, the harmful effects of elevated ocean temperatures take hold and may be the primary driver of the escalating pole-ward migration of species.

SUMMARYLike most teleosts, sablefish (Anoplopoma fimbria Pallas 1814) blood exhibits a moderate Root effect (~35% maximal desaturation), where a reduction in blood pH dramatically reduces O2 carrying capacity, a mechanism important for oxygenating the eye and filling the swim bladder (SB) in teleosts. Although sablefish lack a SB, we observed a well-defined choroid rete at the eye. The adrenergically mediated cell swelling typically associated with a functional red blood cell (RBC) β-adrenergic Na+/H+ exchanger (βNHE), which would normally protect RBC pH, and thus O2 transport, during a generalized acidosis, was not observed in sablefish blood. Neither isoproterenol (a β-agonist) nor 8-bromo cAMP could elicit this response. Furthermore, RBC osmotic shrinkage, known to stimulate NHEs in general and βNHE in other teleosts such as trout and flounder, resulted in no significant regulatory volume increase (RVI), further supporting the absence of a functional RBC βNHE. The onset of the Root effect occurs at a much lower RBC pH (6.83–6.92) than in other teleosts, and thus RBC βNHE may not be required to protect O2 transport during a generalized acidosis in vivo. Phylogenetically, sablefish may represent a fifth group of teleosts exhibiting a secondary reduction or loss of βNHE activity. However, sablefish have not lost the choroid rete at the eye (unlike in the other four groups), which may still function with the Root effect to oxygenate the retina, but the low pH onset of the Root effect may ensure haemoglobin (Hb)-O2 binding is not compromised at the respiratory surface during a general acidosis in the absence of RBC βNHE. The sablefish may represent an anomaly within the framework of Root effect evolution, in that they possess a moderate Root effect and a choroid rete at the eye, but lack the RBC βNHE and the SB system.

Worldwide, increasing coastal development has played a major role in shaping coral reef species assemblages, but the mechanisms underpinning distribution patterns remain poorly understood. Recent research demonstrated delayed development in larval fishes exposed to suspended sediment, highlighting the need to further understand the interaction between suspended sediment as a stressor and energetically costly activities such as growth and development that are essential to support biological fitness. We examined the gill morphology and the gill microbiome in clownfish larvae (Amphiprion percula) exposed to suspended sediment concentrations (using Australian bentonite) commonly found on the inshore Great Barrier Reef. The gills of larvae exposed to 45 mg L−1 of suspended sediment had excessive mucous discharge and growth of protective cell layers, resulting in a 56% thicker gill epithelium compared to fish from the control group. Further, we found a shift from ‘healthy’ to pathogenic bacterial communities on the gills, which could increase the disease susceptibility of larvae. The impact of suspended sediments on larval gills may represent an underlying mechanism behind the distribution patterns of fish assemblages. Our findings underscore the necessity for future coastal development to consider adverse effects of suspended sediments on fish recruitment and consequently fish populations and ecosystem health.

SUMMARY Erythropoietin (EPO) is a glycoprotein hormone that regulates the proliferation and differentiation of erythroid progenitor cells in mammals. Although EPO has been identified in fish, the specific function and effects of hypoxia have not been investigated previously. In this study, we have demonstrated a relationship between increases in renal EPO levels and decreases in spleen EPO levels and spleen-somatic index (SSI), with increases in haemoglobin (Hb) concentration in the blood during hypoxia exposure in rainbow trout. Splenic contraction and the subsequent red blood cell release accounts for the initial increase in Hb concentration in the blood, whereas EPO action probably accounts for the later increases in hemoglobin concentration in the blood. Our data indicate that fish and mammalian erythropoietic systems are similar in response to hypoxia, in that erythropoiesis in fish is influenced by EPO.

Many teleostean fish, including rainbow trout, regulate red blood cell (RBC) pH (pH(i)) in the presence of a stress-induced acidosis such as hypoxia, hypercapnia, or exhaustive exercise. This is accomplished through activation of RBC Na+/H+ exchange (beta-NHE), ultimately minimizing impairment to oxygen transport. Presence and characterization of the RBC beta-NHE in fish is best tested in blood from cannulated, resting animals; however, several studies have used blood from stressed animals drawn from the caudal vein and stored prior to use. The effects of sampling procedures and storage on the beta-NHE response is not known and is the focus of this study. Whole blood drawn from cannulated, resting rainbow trout was compared with RBCs obtained from the caudal vein rinsed and stored at 4 degrees C for 0, 6, 24, 48, 96 or 144 h. Isoproterenol (10(-5) M), a beta-adrenergic agonist, was added to hypoxia/hypercapnia incubated RBCs in vitro. In all treatments, isoproterenol induced a large beta-NHE response, and storage duration (or =96 h) had a minimal affect, indicating that rinsing and storing is an easy and viable means by which to obtain RBCs and investigate function. Storage for 144 h still resulted in a significant RBC beta-NHE response; however, viability of RBCs may be compromised.

Rising ocean temperatures are predicted to cause a poleward shift in the distribution of marine fishes occupying the extent of latitudes tolerable within their thermal range boundaries. A prevailing theory suggests that the upper thermal limits of fishes are constrained by hypoxia and ocean acidification. However, some eurythermal fish species do not conform to this theory, and maintain their upper thermal limits in hypoxia. Here we determine if the same is true for stenothermal species. In three coral reef fish species we tested the effect of hypoxia on upper thermal limits, measured as critical thermal maximum (CT max ). In one of these species we also quantified the effect of hypoxia on oxygen supply capacity, measured as aerobic scope (AS). In this species we also tested the effect of elevated CO 2 (simulated ocean acidification) on the hypoxia sensitivity of CT max . We found that CT max was unaffected by progressive hypoxia down to approximately 35 mmHg, despite a substantial hypoxia-induced reduction in AS. Below approximately 35 mmHg, CT max declined sharply with water oxygen tension ( P w O 2 ). Furthermore, the hypoxia sensitivity of CT max was unaffected by elevated CO 2 . Our findings show that moderate hypoxia and ocean acidification do not constrain the upper thermal limits of these tropical, stenothermal fishes.

Ocean warming and acidification are serious threats to marine life. While each stressor alone has been studied in detail, their combined effects on the outcome of ecological interactions are poorly understood. We measured predation rates and predator selectivity of two closely related species of damselfish exposed to a predatory dottyback. We found temperature and CO2 interacted synergistically on overall predation rate, but antagonistically on predator selectivity. Notably, elevated CO2 or temperature alone reversed predator selectivity, but the interaction between the two stressors cancelled selectivity. Routine metabolic rates of the two prey showed strong species differences in tolerance to CO2 and not temperature, but these differences did not correlate with recorded mortality. This highlights the difficulty of linking species-level physiological tolerance to resulting ecological outcomes. This study is the first to document both synergistic and antagonistic effects of elevated CO2 and temperature on a crucial ecological process like predator-prey dynamics.

Tropical regions are expected to be some of the most affected by rising sea surface temperatures (SSTs) because seasonal temperature variations are minimal. As temperatures rise, less oxygen dissolves in water, but metabolic requirements of fish and thus, the demand for effective oxygen uptake, increases. Gill remodelling is an acclimation strategy well documented in freshwater cyprinids experiencing large seasonal variations in temperature and oxygen as well as an amphibious killifish upon air exposure. However, no study has investigated whether tropical reef fishes remodel their gills to allow for increased oxygen demands at elevated temperatures. We tested for gill remodelling in five coral reef species (Acanthochromis polyacanthus, Chromis atripectoralis, Pomacentrus moluccensis, Dascyllus melanurus and Cheilodipterus quinquelineatus) from populations in northern Papua New Guinea (2° 35.765′ S; 150° 46.193′ E). Fishes were acclimated for 12-14 days to 29 and 31 °C, encompassing their seasonal range (29-31 °C), and 33 and 34 °C to account for end-of-century predicted temperatures. We measured lamellar perimeter, cross-sectional area, base thickness, and length for five filaments on the 2nd gill arches and qualitatively assessed 3rd gill arches via scanning electron microscopy (SEM). All species exhibited significant differences in the quantitative measurements made on the lamellae, but no consistent trends with temperature were observed. SEM only revealed alterations in gill morphology in P. moluccensis. The overall lack of changes in gill morphology with increasing temperature suggests that these near-equatorial reef fishes may fail to maintain adequate O2 uptake under future climate scenarios unless other adaptive mechanisms are employed.

Teleost fishes constitute 95% of extant aquatic vertebrates, and we suggest that this is related in part to their unique mode of tissue oxygenation. We propose the following sequence of events in the evolution of their oxygen delivery system. First, loss of plasma-accessible carbonic anhydrase (CA) in the gill and venous circulations slowed the Jacobs–Stewart cycle and the transfer of acid between the plasma and the red blood cells (RBCs). This ameliorated the effects of a generalised acidosis (associated with an increased capacity for burst swimming) on haemoglobin (Hb)–O2 binding. Because RBC pH was uncoupled from plasma pH, the importance of Hb as a buffer was reduced. The decrease in buffering was mediated by a reduction in the number of histidine residues on the Hb molecule and resulted in enhanced coupling of O2 and CO2 transfer through the RBCs. In the absence of plasma CA, nearly all plasma bicarbonate ultimately dehydrated to CO2 occurred via the RBCs, and chloride/bicarbonate exchange was the rate-limiting step in CO2 excretion. This pattern of CO2 excretion across the gills resulted in disequilibrium states for CO2 hydration/dehydration reactions and thus elevated arterial and venous plasma bicarbonate levels. Plasma-accessible CA embedded in arterial endothelia was retained, which eliminated the localized bicarbonate disequilibrium forming CO2 that then moved into the RBCs. Consequently, RBC pH decreased which, in conjunction with pH-sensitive Bohr/Root Hbs, elevated arterial oxygen tensions and thus enhanced tissue oxygenation. Counter-current arrangement of capillaries (retia) at the eye and later the swim bladder evolved along with the gas gland at the swim bladder. Both arrangements enhanced and magnified CO2 and acid production and, therefore, oxygen secretion to those specialised tissues. The evolution of β-adrenergically stimulated RBC Na+/H+ exchange protected gill O2 uptake during stress and further augmented plasma disequilibrium states for CO2 hydration/dehydration. Finally, RBC organophosphates (e.g. NTP) could be reduced during hypoxia to further increase Hb–O2 affinity without compromising tissue O2 delivery because high-affinity Hbs could still adequately deliver O2 to the tissues via Bohr/Root shifts. We suggest that the evolution of this unique mode of tissue O2 transfer evolved in the Triassic/Jurassic Period, when O2 levels were low, ultimately giving rise to the most extensive adaptive radiation of extant vertebrates, the teleost fishes.

Summary Metabolic rates of aquatic organisms are estimated from measurements of oxygen consumption rates (ṀO2) through swimming and resting respirometry. These distinct approaches are increasingly used in eco- and conservation physiology studies; however, few studies have tested whether they yield comparable results. We examined whether two fundamental ṀO2 measures, standard metabolic rate (SMR) and maximum metabolic rate (MMR), vary based on the method employed. Ten bridled monocle bream (Scolopsis bilineatus) were exercised using (1) a critical swimming speed (Ucrit) protocol, (2) a 15 min exhaustive chase protocol and (3) a 3 min exhaustive chase protocol followed by brief air exposure. Protocol (1) was performed in a swimming respirometer whereas protocols (2) and (3) were followed by resting respirometry. SMR estimates in swimming respirometry were similar to those in resting respirometry when a three-parameter exponential or power function was used to extrapolate the swimming speed-ṀO2 relationship to zero swimming speed. In contrast, MMR using the Ucrit protocol was 36% higher than MMR derived from the 15 min chase protocol and 23% higher than MMR using the 3 min chase 1 min air exposure protocol. For strong steady (endurance) swimmers, such as S. bilineatus, swimming respirometry can produce more accurate MMR estimates than exhaustive chase protocols because oxygen consumption is measured during exertion. However, when swimming respirometry is impractical, exhaustive chase protocols should be supplemented with brief air exposure to improve measurement accuracy. Caution is warranted when comparing MMR estimates obtained with different respirometry methods unless they are cross-validated on a species-specific basis.

Summary Teleost fishes and mammalian lineages diverged 400 million years ago, and environmental requirements (water vs. air) have resulted in marked differences in cardiovascular function between fish and mammals. Suggestions that the fish secondary vascular system (SVS) could be used as a model for the mammalian lymphatic system should be taken with caution. Despite molecular markers indicating similar genetic origin, functions of the SVS in teleost fish are probably different from those of the mammalian lymphatic system. We determined that, in resting glass catfish, (Kryptopterus bicirrhis), plasma moves from the primary vascular system (PVS) to the SVS through small connecting vessels less than 10 μm in diameter, smaller than the red blood cells (RBCs). During and following hypoxia or exercise, flow increases, and RBCs enter the SVS, possibly via β-adrenoreceptor-mediated dilation of the connecting vessels. The volume of the SVS can be large and, as RBCs flow into the SVS, the haematocrit of the PVS falls by as much as 50% of the resting value. Possible functions of the SVS, including skin respiration, ionic and osmotic buffering, and reductions in heart work and RBC turnover, are discussed.

SUMMARYDuring a generalized acidosis in rainbow trout, catecholamines are released into the blood, activating red blood cell (RBC) Na+/H+ exchange (βNHE), thus protecting RBC intracellular pH (pHi) and subsequent O2 binding at the gill. Because of the presence of a Root effect (a reduction in oxygen carrying capacity of the blood with a reduction in pH), the latter could otherwise be impaired. However, plasma-accessible carbonic anhydrase (CA) at the tissues (and absence at the gills) may result in selective short-circuiting of RBC βNHE pH regulation. This would acidify the RBCs and greatly enhance O2 delivery by exploitation of the combined Bohr-Root effect, a mechanism not previously proposed. As proof-of-principle, an in vitro closed system was developed to continuously monitor extracellular pH (pHe) and O2 tension (PO2) of rainbow trout blood. In this closed system, adding CA to acidified, adrenergically stimulated RBCs short-circuited βNHE pH regulation, resulting in an increase in PO2 by >30 mmHg, depending on the starting Hb-O2 saturation and degree of initial acidification. Interestingly, in the absence of adrenergic stimulation, addition of CA still elevated PO2, albeit to a lesser extent, a response that was absent during general NHE inhibition. If plasma-accessible CA-mediated short-circuiting is operational in vivo, the combined Bohr-Root effect system unique to teleost fishes could markedly enhance tissue O2 delivery far in excess of that in vertebrates possessing a Bohr effect alone and may lead to insights about the early evolution of the Root effect.

Tunas (family Scombridae) are exceptional among most teleost fishes in that they possess vascular heat exchangers which allow heat retention in specific regions of the body (termed 'regional heterothermy'). Seemingly exclusive to heterothermic fishes is a markedly reduced temperature dependence of blood-oxygen (blood-O(2)) binding, or even a reversed temperature dependence where increasing temperature increases blood-O(2) affinity. These unusual binding properties have been documented in whole blood and in haemoglobin (Hb) solutions, and they are hypothesised to prevent oxygen loss from arteries to veins within the vascular heat exchangers and/or to prevent excessive oxygen unloading to the warm tissues and ensure an adequate supply of oxygen to tissues positioned efferent to the heat exchangers. The temperature sensitivity of blood-O(2) binding has not been characterised in an ectothermic scombrid (mackerels and bonitos), but the existence of the unusual binding properties in these fishes would have clear implications for their proposed association with regional heterothermy. Accordingly, the present study examined oxygenation of whole blood of the chub mackerel (Scomber japonicus) at 10, 20 and 30 degrees C and at 0.5, 1 and 2% CO(2). Oxygen affinity was generally highest at 20 degrees C for all levels of CO(2). Temperature-independent binding was observed at low (0.5%) CO(2), where the PO(2) at 50% blood-O(2) saturation (P (50)) was not statistically different at 10 and 30 degrees C (2.58 vs. 2.78 kPa, respectively) with an apparent heat of oxygenation (H degrees ) close to zero (-6 kJ mol(-1)). The most significant temperature-mediated difference occurred at high (2%) CO(2), where the P (50) at 10 degrees C was twofold higher than that at 20 degrees C with a corresponding H degrees of +43 kJ mol(-1). These results provide clear evidence of independent and reversed open-system temperature effects on blood oxygenation in S. japonicus, and it is therefore speculated that these unusual blood-O(2) binding characteristics may have preceded the evolution of vascular heat exchangers and regional heterothermy in fishes.

We have investigated whether mild heat shock, and resulting Hsp70 expression, can confer cross-protection against the stress associated with transfer from freshwater (FW) to seawater (SW) in juvenile rainbow trout (Oncorhynchus mykiss). In experimental Series I, juvenile trout reared in FW were transferred from 13.5 degrees C to 25.5 degrees C in FW, held for 2 h, returned to 13.5 degrees C for 12 h, and then transferred to 32 ppt SW at 13.5 degrees C. Branchial Hsp70 increased approximately 10-fold in the heat-shocked fish relative to the control by the end of recovery and remained high 2, 8, and 24 h post-salinity transfer. However, no clear differences could be detected in blood parameters (blood hemoglobin, hematocrit, MCHC, plasma Na(+) and plasma osmolarity) or muscle water content between heat-shocked and sham-shocked fish in SW at any sampling interval (0, 2, 8, 24, 48, 120, 240 and 360 h post-SW transfer). In experimental Series II, trout acclimated to 8 degrees C were heat-shocked at 22 degrees C for 2 h, allowed to recover 18 h, and exposed to a more severe salinity transfer (either 36 or 45 ppt) than in Series I. Branchial Hsp70 levels increased approximately 6-fold in heat-shocked fish, but had declined to baseline after 120 h in SW. Plasma osmolarity and chloride increased in both groups upon transfer to 36 ppt; however, the increase was significantly less in heat-shocked fish when compared to the increase observed in sham-shocked fish at 24 h. No significant differences could be detected in branchial Na(+)/K(+)-ATPase activity or Na(+)/K(+)-ATPase alpha1a and alpha1b mRNA expression between the two groups. Our data indicate that a mild temperature shock has only modest effects on the ability of rainbow trout to resist osmotic stress during FW to SW transfer.

The teleost fishes represent over half of all extant vertebrates; they occupy nearly every body of water and in doing so, occupy a diverse array of environmental conditions. We propose that their success is related to a unique oxygen (O2) transport system involving their extremely pH-sensitive haemoglobin (Hb). A reduction in pH reduces both Hb-O2 affinity (Bohr effect) and carrying capacity (Root effect). This, combined with a large arterial-venous pH change (ΔpHa-v) relative to other vertebrates, may greatly enhance tissue oxygen delivery in teleosts (e.g., rainbow trout) during stress, beyond that in mammals (e.g., human). We generated oxygen equilibrium curves (OECs) at five different CO2 tensions for rainbow trout and determined that, when Hb-O2 saturation is 50% or greater, the change in oxygen partial pressure (ΔPO2) associated with ΔpHa-v can exceed that of the mammalian Bohr effect by at least 3-fold, but as much as 21-fold. Using known ΔpHa-v and assuming a constant arterial-venous PO2 difference (Pa-vO2), Root effect Hbs can enhance O2 release to the tissues by 73.5% in trout; whereas, the Bohr effect alone is responsible for enhancing O2 release by only 1.3% in humans. Disequilibrium states are likely operational in teleosts in vivo, and therefore the ΔpHa-v, and thus enhancement of O2 delivery, could be even larger. Modeling with known Pa-vO2 in fish during exercise and hypoxia indicates that O2 release from the Hb and therefore potentially tissue O2 delivery may double during exercise and triple during some levels of hypoxia. These characteristics may be central to performance of athletic fish species such as salmonids, but may indicate that general tissue oxygen delivery may have been the incipient function of Root effect Hbs in fish, a trait strongly associated with the adaptive radiation of teleosts.