Getting to the heart of anatomical diversity and phenotypic plasticity: fish hearts are an optimal organ model in need of greater mechanistic study.

Natural selection has produced many vertebrate 'solutions' for the cardiac life-support system, especially among the approximately 30,000 species of fishes. For example, across species, fish have the greatest range for central arterial blood pressure and relative ventricular mass of any vertebrate group. This enormous cardiac diversity is excellent ground material for mechanistic explorations. Added to this species diversity is the emerging field of population-specific diversity, which is revealing that cardiac design and function can be tailored to a fish population's local environmental conditions. Such information is important to conservation biologists and ecologists, as well as physiologists. Furthermore, the cardiac structure and function of an individual adult fish are extremely pliable (through phenotypic plasticity), which is typically beneficial to the heart's function when environmental conditions are variable. Consequently, exploring factors that trigger cardiac remodelling with acclimation to new environments represents a marvellous opportunity for performing mechanistic studies that minimize the genetic differences that accompany cross-species comparisons. What makes the heart an especially good system for the investigation of phenotypic plasticity and species diversity is that its function can be readily evaluated at the organ level using established methodologies, unlike most other organ systems. Although the fish heart has many merits as an organ-level model to provide a mechanistic understanding of phenotypic plasticity and species diversity, bringing this potential to fruition will require productive research collaborations among physiologists, geneticists, developmental biologists and ecologists.

Relationship between capillaries, mitochondria and maximum power of the heart: a meta-study from shrew to elephant.

This meta-study uses phylogenetic scaling models across more than 30 species, spanning five orders of magnitude in body mass, to show that cardiac capillary numerical density and mitochondrial volume density decrease with body mass raised to the -0.07 ± 0.03 and -0.04 ± 0.01 exponents, respectively. Thus, while an average 10 g mammal has a cardiac capillary density of approximately 4150 mm-2 and a mitochondrial density of 33%, a 1 t mammal has considerably lower corresponding values of 1850 mm-2 and 21%. These similar scaling trajectories suggest quantitative matching for the primary oxygen supply and oxygen consuming structures of the heart, supporting economic design at the cellular level of the oxygen cascade in this aerobic organ. These scaling trajectories are nonetheless somewhat shallower than the exponent of -0.11 calculated for the maximum external mechanical power of the cardiac tissue, under conditions of heavy exercise, when oxygen flow between capillaries and mitochondria is probably fully exploited. This mismatch, if substantiated, implies a declining external mechanical efficiency of the heart with increasing body mass, whereby larger individuals put more energy in but get less energy out, a scenario with implications for cardiovascular design, aerobic capacity and limits of body size.

Host-pathogen-environment interactions predict survival outcomes of adult sockeye salmon (Oncorhynchus nerka) released from fisheries.

Incorporating host-pathogen(s)-environment axes into management and conservation planning is critical to preserving species in a warming climate. However, the role pathogens play in host stress resilience remains largely unexplored in wild animal populations. We experimentally characterized how independent and cumulative stressors (fisheries handling, high water temperature) and natural infections affected the health and longevity of released wild adult sockeye salmon (Oncorhynchus nerka) in British Columbia, Canada. Returning adults were collected before and after entering the Fraser River, yielding marine- and river-collected groups, respectively (N = 185). Fish were exposed to a mild (seine) or severe (gill net) fishery treatment at collection, and then held in flow-through freshwater tanks for up to four weeks at historical (14°C) or projected migration temperatures (18°C). Using weekly nonlethal gill biopsies and high-throughput qPCR, we quantified loads of up to 46 pathogens with host stress and immune gene expression. Marine-collected fish had less severe infections than river-collected fish, a short migration distance (100 km, 5-7 days) that produced profound infection differences. At 14°C, river-collected fish survived 1-2 weeks less than marine-collected fish. All fish held at 18°C died within 4 weeks unless they experienced minimal handling. Gene expression correlated with infections in river-collected fish, while marine-collected fish were more stressor-responsive. Cumulative stressors were detrimental regardless of infections or collection location, probably due to extreme physiological disturbance. Because river-derived infections correlated with single stressor responses, river entry probably decreases stressor resilience of adult salmon by altering both physiology and pathogen burdens, which redirect host responses toward disease resistance.

Characterizing the hypoxic performance of a fish using a new metric: PAAS-50.

The hypoxic constraint on peak oxygen uptake (MO2,peak) was characterized in rainbow trout over a range of ambient oxygen tensions with different testing protocols and statistical models. The best-fit model was selected using both statistical criteria (R2 and AIC) and the model's prediction of three anchor points for hypoxic performance: critical PO2 (Pcrit), maximum MO2 and a new metric, the minimum PO2 that supports 50% of absolute aerobic scope (PAAS-50). The best-fitting model was curvilinear using five strategically selected PO2 values. This model predicted PAAS-50 as 70 mmHg (coefficient of variation, CV=9%) for rainbow trout. Thus, while a five-point hypoxic performance curve can characterize the limiting effects of hypoxia in fish, as envisaged by Fry over 75 years ago, PAAS-50 is a promising metric to compare hypoxic constraints on performance in a standardized manner both within and across fish species.

Rapid cardiac thermal acclimation in wild anadromous Arctic char (Salvelinus alpinus).

Migratory fishes commonly encounter large and rapid thermal variation, which has the potential to disrupt essential physiological functions. Thus, we acclimated wild, migratory Arctic char to 13°C (∼7°C above a summer average) for an ecologically relevant period (3 days) and measured maximum heart rate (ƒH,max) during acute warming to determine their ability to rapidly improve cardiac function at high temperatures. Arctic char exhibited rapid compensatory cardiac plasticity similar to past observations following prolonged warm acclimation: they reduced ƒH,max over intermediate temperatures (-8%), improved their ability to increase ƒH,max during warming (+10%), and increased (+1.3°C) the temperature at the onset of an arrhythmic heartbeat, a sign of cardiac failure. This rapid cardiac plasticity may help migrating fishes such as Arctic char mitigate short-term thermal challenges. Furthermore, by using mobile Arctic research infrastructure in a remote field location, the present study illustrates the potential for field-based, experimental physiology in such locations.

Warm, but not hypoxic acclimation, prolongs ventricular diastole and decreases the protein level of Na+/Ca2+ exchanger to enhance cardiac thermal tolerance in European sea bass.

One of the physiological mechanisms that can limit the fish's ability to face hypoxia or elevated temperature, is maximal cardiac performance. Yet, few studies have measured how cardiac electrical activity and associated calcium cycling proteins change with acclimation to those environmental stressors. To examine this, we acclimated European sea bass for 6 weeks to three experimental conditions: a seasonal average temperature in normoxia (16 °C; 100% air sat.), an elevated temperature in normoxia (25 °C; 100% air sat.) and a seasonal average temperature in hypoxia (16 °C; 50% air sat.). Following each acclimation, the electrocardiogram was measured to assess how acclimation affected the different phases of cardiac cycle, the maximal heart rate (fHmax) and cardiac thermal performance during an acute increase of temperature. Whereas warm acclimation prolonged especially the diastolic phase of the ventricular contraction, reduced the fHmax and increased the cardiac arrhythmia temperature (TARR), hypoxic acclimation was without effect on these functional indices. We measured the level of two key proteins involved with cellular relaxation of cardiomyocytes, i.e. sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) and Na+/Ca2+ exchanger (NCX). Warm acclimation reduced protein level of both NCX and SERCA and hypoxic acclimation reduced SERCA protein levels without affecting NCX. The changes in ventricular NCX level correlated with the observed changes in diastole duration and fHmax as well as TARR. Our results shed new light on mechanisms of cardiac plasticity to environmental stressors and suggest that NCX might be involved with the observed functional changes, yet future studies should also measure its electrophysiological activity.

Innate antiviral defense demonstrates high energetic efficiency in a bony fish.

BACKGROUND: Viruses can impose energetic demands on organisms they infect, in part by hosts mounting resistance. Recognizing that oxygen uptake reliably indicates steady-state energy consumption in all vertebrates, we comprehensively evaluated oxygen uptake and select transcriptomic messaging in sockeye salmon challenged with either a virulent rhabdovirus (IHNV) or a low-virulent reovirus (PRV). We tested three hypotheses relating to the energetic costs of viral resistance and tolerance in this vertebrate system: (1) mounting resistance incurs a metabolic cost or limitation, (2) induction of the innate antiviral interferon system compromises homeostasis, and (3) antiviral defenses are weakened by acute stress. RESULTS: IHNV infections either produced mortality within 1-4 weeks or the survivors cleared infections within 1-9 weeks. Transcription of three interferon-stimulated genes (ISGs) was strongly correlated with IHNV load but not respiratory performance. Instead, early IHNV resistance was associated with a mean 19% (95% CI = 7-31%; p = 0.003) reduction in standard metabolic rate. The stress of exhaustive exercise did not increase IHNV transcript loads, but elevated host inflammatory transcriptional signaling up to sevenfold. For PRV, sockeye tolerated high-load systemic PRV blood infections. ISG transcription was transiently induced at peak PRV loads without associated morbidity, microscopic lesions, or major changes in aerobic or anaerobic respiratory performance, but some individuals with high-load blood infections experienced a transient, minor reduction in hemoglobin concentration and increased duration of excess post-exercise oxygen consumption. CONCLUSIONS: Contrary to our first hypothesis, effective resistance against life-threatening rhabdovirus infections or tolerance to high-load reovirus infections incurred minimal metabolic costs to salmon. Even robust systemic activation of the interferon system did not levy an allostatic load sufficient to compromise host homeostasis or respiratory performance, rejecting our second hypothesis that this ancient innate vertebrate antiviral defense is itself energetically expensive. Lastly, an acute stress experienced during testing did not weaken host antiviral defenses sufficiently to promote viral replication; however, a possibility for disease intensification contingent upon underlying inflammation was indicated. These data cumulatively demonstrate that fundamental innate vertebrate defense strategies against potentially life-threatening viral exposure impose limited putative costs on concurrent aerobic or energetic demands of the organism.

Resilience of cardiac performance in Antarctic notothenioid fishes in a warming climate.

Warming in the region of the Western Antarctic Peninsula is occurring at an unprecedented rate, which may threaten the survival of Antarctic notothenioid fishes. Herein, we review studies characterizing thermal tolerance and cardiac performance in notothenioids - a group that includes both red-blooded species and the white-blooded, haemoglobinless icefishes - as well as the relevant biochemistry associated with cardiac failure during an acute temperature ramp. Because icefishes do not feed in captivity, making long-term acclimation studies unfeasible, we focus only on the responses of red-blooded notothenioids to warm acclimation. With acute warming, hearts of the white-blooded icefish Chaenocephalus aceratus display persistent arrhythmia at a lower temperature (8°C) compared with those of the red-blooded Notothenia coriiceps (14°C). When compared with the icefish, the enhanced cardiac performance of N. coriiceps during warming is associated with greater aerobic capacity, higher ATP levels, less oxidative damage and enhanced membrane integrity. Cardiac performance can be improved in N. coriiceps with warm acclimation to 5°C for 6-9 weeks, accompanied by an increase in the temperature at which cardiac failure occurs. Also, both cardiac mitochondrial and microsomal membranes are remodelled in response to warm acclimation in N. coriiceps, displaying homeoviscous adaptation. Overall, cardiac performance in N. coriiceps is malleable and resilient to warming, yet thermal tolerance and plasticity vary among different species of notothenioid fishes; disruptions to the Antarctic ecosystem driven by climate warming and other anthropogenic activities endanger the survival of notothenioids, warranting greater protection afforded by an expansion of marine protected areas.

The thermal acclimation potential of maximum heart rate and cardiac heat tolerance in Arctic char (Salvelinus alpinus), a northern cold-water specialist.

Increasing heart rate (ƒH) is a central, if not primary mechanism used by fishes to support their elevated tissue oxygen consumption during acute warming. Thermal acclimation can adjust this acute response to improve cardiac performance and heat tolerance under the prevailing temperatures. We predict that such acclimation will be particularly important in regions undergoing rapid environmental change such as the Arctic. Therefore, we acclimated Arctic char (Salvelinus alpinus), a high latitude, cold-adapted salmonid, to ecologically relevant temperatures (2, 6, 10, 14 and 18 °C) and examined how thermal acclimation influenced their cardiac heat tolerance by measuring the maximum heart rate (ƒHmax) response to acute warming. As expected, acute warming increased ƒHmax in all Arctic char before ƒHmax reached a peak and then became arrhythmic. The peak ƒHmax, and the temperature at which peak ƒHmax (Tpeak) and that at which arrhythmia first occurred (Tarr) all increased progressively (+33%, 49% and 35%, respectively) with acclimation temperature from 2 to 14 °C. When compared at the same test temperature ƒHmax also decreased by as much as 29% with increasing acclimation temperature, indicating significant thermal compensation. The upper temperature at which fish first lost their equilibrium (critical thermal maximum: CTmax) also increased with acclimation temperature, albeit to a lesser extent (+11%). Importantly, Arctic char experienced mortality after several weeks of acclimation at 18 °C and survivors did not have elevated cardiac thermal tolerance. Collectively, these findings suggest that if wild Arctic char have access to suitable temperatures (<18 °C) for a sufficient duration, warm acclimation can potentially mitigate some of the cardiorespiratory impairments previously documented during acute heat exposure.

Valid oxygen uptake measurements: using high r2 values with good intentions can bias upward the determination of standard metabolic rate.

This analysis shows good intentions in the selection of valid and precise oxygen uptake ( M ˙ O2 ) measurements by retaining only slopes of declining dissolved oxygen level in a respirometer that have very high values of the coefficient of determination, r2 , are not always successful at excluding nonlinear slopes. Much worse, by potentially removing linear slopes that have low r2 only because of a low signal-to-noise ratio, this procedure can overestimate the calculation of standard metabolic rate (SMR) of the fish. To remedy this possibility, a few simple diagnostic tools are demonstrated to assess the appropriateness of a given minimum acceptable r2 , such as calculating the proportion of rejected M ˙ O2 determinations, producing a histogram of the r2 values and a plot of r2 as a function of M ˙ O2 . The authors offer solutions for cases when many linear slopes have low r2 . The least satisfactory but easiest to implement is lowering the minimum acceptable r2 . More satisfactory solutions involve processing (smoothing) the raw signal of dissolved oxygen as a function of time to improve the signal-to-noise ratio and the r2 s.

Intraspecific variation in tolerance of warming in fishes.

Intraspecific variation in key traits such as tolerance of warming can have profound effects on ecological and evolutionary processes, notably responses to climate change. The empirical evidence for three primary elements of intraspecific variation in tolerance of warming in fishes is reviewed. The first is purely mechanistic that tolerance varies across life stages and as fishes become mature. The limited evidence indicates strongly that this is the case, possibly because of universal physiological principles. The second is intraspecific variation that is because of phenotypic plasticity, also a mechanistic phenomenon that buffers individuals' sensitivity to negative impacts of global warming in their lifetime, or to some extent through epigenetic effects over successive generations. Although the evidence for plasticity in tolerance to warming is extensive, more work is required to understand underlying mechanisms and to reveal whether there are general patterns. The third element is intraspecific variation based on heritable genetic differences in tolerance, which underlies local adaptation and may define long-term adaptability of a species in the face of ongoing global change. There is clear evidence of local adaptation and some evidence of heritability of tolerance to warming, but the knowledge base is limited with detailed information for only a few model or emblematic species. There is also strong evidence of structured variation in tolerance of warming within species, which may have ecological and evolutionary significance irrespective of whether it reflects plasticity or adaptation. Although the overwhelming consensus is that having broader intraspecific variation in tolerance should reduce species vulnerability to impacts of global warming, there are no sufficient data on fishes to provide insights into particular mechanisms by which this may occur.

A rapid intrinsic heart rate resetting response with thermal acclimation in rainbow trout, Oncorhynchus mykiss.

We examined cardiac pacemaker rate resetting in rainbow trout following a reciprocal temperature transfer. In the original experiment, performed in winter, 4°C-acclimated fish transferred to 12°C reset intrinsic heart rate after just 1 h (from 56.8±1.2 to 50.8±1.5 beats min-1); 12°C-acclimated fish transferred to 4°C reset intrinsic heart rate after 8 h (from 33.4±0.7 to 37.7±1.2 beats min-1). However, in a replicate experiment, performed in the summer using a different brood year, intrinsic heart rate was not reset, even after 10 weeks at a new temperature. Using this serendipitous opportunity, we compared mRNA expression changes of a suite of proteins in sinoatrial node (SAN), atrial and ventricular tissues after both 1 h and longer than 3 weeks for both experimental acclimation groups to identify those changes only associated with pacemaker rate resetting. Of the changes in mRNA expression occurring after more than 3 weeks of warm acclimation and associated with pacemaker rate resetting, we observed downregulation of NKA α1c in the atrium and ventricle, and upregulation of HCN1 in the ventricle. However, in the SAN there were no mRNA expression changes unique to the fish with pacemaker rate resetting after either 1 h or 3 weeks of warm acclimation. Thus, despite identifying changes in mRNA expression of contractile cardiac tissues, there was an absence of changes in mRNA expression directly involved with the initial, rapid pacemaker rate resetting with warm acclimation. Importantly, pacemaker rate resetting with thermal acclimation does not always occur in rainbow trout.

Cardiovascular responses to progressive hypoxia in ducks native to high altitude in the Andes.

The cardiovascular system is critical for delivering O2 to tissues. Here we examine the cardiovascular responses to progressive hypoxia in four high-altitude Andean duck species compared to four related low-altitude populations in North America, tested at their native altitude. Ducks were exposed to stepwise decreases in inspired partial pressure of O2 while we monitored heart rate, O2 consumption rate, blood O2 saturation, haematocrit (Hct), and blood haemoglobin concentration [Hb]. We calculated O2 pulse (the product of stroke volume and the arterial-venous O2 content difference), blood O2 concentration, and heart rate variability. Regardless of altitude, all eight populations maintained O2 consumption rate with minimal change in heart rate or O2 pulse, indicating that O2 consumption was maintained by either a constant arterial-venous O2 content difference (an increase in the relative O2 extracted from arterial blood) or by a combination of changes in stroke volume and the arterial-venous O2 content difference. Three high-altitude taxa (yellow-billed pintails, cinnamon teal, and speckled teal) had higher Hct and [Hb], increasing the O2 content of arterial blood, and potentially providing a greater reserve for enhancing O2 delivery during hypoxia. Hct and [Hb] between low- and high-altitude populations of ruddy duck were similar, representing a potential adaptation to diving life. Heart rate variability was generally lower in high-altitude ducks, concurrent with similar or lower heart rates than low-altitude ducks, suggesting a reduction in vagal and sympathetic tone. These unique features of the Andean ducks differ from previous observations in both Andean geese and bar-headed geese, neither of which exhibit significant elevations in Hct or [Hb] compared to their low-altitude relatives, revealing yet another avian strategy for coping with high altitude.

Cardiovascular responses to progressive hypoxia in ducks native to high altitude in the Andes.

The cardiovascular system is critical for delivering O2 to tissues. Here, we examined the cardiovascular responses to progressive hypoxia in four high-altitude Andean duck species compared with four related low-altitude populations in North America, tested at their native altitude. Ducks were exposed to stepwise decreases in inspired partial pressure of O2 while we monitored heart rate, O2 consumption rate, blood O2 saturation, haematocrit (Hct) and blood haemoglobin (Hb) concentration. We calculated O2 pulse (the product of stroke volume and the arterial-venous O2 content difference), blood O2 concentration and heart rate variability. Regardless of altitude, all eight populations maintained O2 consumption rate with minimal change in heart rate or O2 pulse, indicating that O2 consumption was maintained by either a constant arterial-venous O2 content difference (an increase in the relative O2 extracted from arterial blood) or by a combination of changes in stroke volume and the arterial-venous O2 content difference. Three high-altitude taxa (yellow-billed pintails, cinnamon teal and speckled teal) had higher Hct and Hb concentration, increasing the O2 content of arterial blood, and potentially providing a greater reserve for enhancing O2 delivery during hypoxia. Hct and Hb concentration between low- and high-altitude populations of ruddy duck were similar, representing a potential adaptation to diving life. Heart rate variability was generally lower in high-altitude ducks, concurrent with similar or lower heart rates than low-altitude ducks, suggesting a reduction in vagal and sympathetic tone. These unique features of the Andean ducks differ from previous observations in both Andean geese and bar-headed geese, neither of which exhibit significant elevations in Hct or Hb concentration compared with their low-altitude relatives, revealing yet another avian strategy for coping with high altitude.

Measuring maximum oxygen uptake with an incremental swimming test and by chasing rainbow trout to exhaustion inside a respirometry chamber yields the same results.

This study hypothesized that oxygen uptake (MO2 ) measured with a novel protocol of chasing rainbow trout Oncorhynchus mykiss to exhaustion inside a static respirometer while simultaneously monitoring MO2 (MO2chase ) would generate the same and repeatable peak value as when peak active MO2 (MO2active ) is measured in a critical swimming speed protocol. To reliably determine peak MO2chase , and compare to the peak during recovery of MO2 after a conventional chase protocol outside the respirometer (MO2rec ), this study applied an iterative algorithm and a minimum sampling window duration (i.e., 1 min based on an analysis of the variance in background and exercise MO2 ) to account for MO2 dynamics. In support of this hypothesis, peak MO2active (707 ± 33 mg O2 h-1 kg-1 ) and peak MO2chase (663 ± 43 mg O2 h-1 kg-1 ) were similar (P = 0.49) and repeatable (Pearson's and Spearman's correlation test; r ≥ 0.77; P < 0.05) when measured in the same fish. Therefore, estimates of MO2max can be independent of whether a fish is exhaustively chased inside a respirometer or swum to fatigue in a swim tunnel, provided MO2 is analysed with an iterative algorithm and a minimum but reliable sampling window. The importance of using this analytical approach was illustrated by peak MO2chase being 23% higher (P < 0.05) when compared with a conventional sequential interval regression analysis, whereas using the conventional chase protocol (1-min window) outside the respirometer increased this difference to 31% (P < 0.01). Moreover, because peak MO2chase was 18% higher (P < 0.05) than peak MO2rec , chasing a fish inside a static respirometer may be a better protocol for obtaining maximum MO2 .

Scaling of cardiac morphology is interrupted by birth in the developing sheep Ovis aries.

Scaling of the heart across development can reveal the degree to which variation in cardiac morphology depends on body mass. In this study, we assessed the scaling of heart mass, left and right ventricular masses, and ventricular mass ratio, as a function of eviscerated body mass across fetal and postnatal development in Horro sheep Ovis aries (~50-fold body mass range; N = 21). Whole hearts were extracted from carcasses, cleaned, dissected into chambers and weighed. We found a biphasic relationship when heart mass was scaled against body mass, with a conspicuous 'breakpoint' around the time of birth, manifest not by a change in the scaling exponent (slope), but rather a jump in the elevation. Fetal heart mass (g) increased with eviscerated body mass (Mb , kg) according to the power equation 4.90 Mb0.88 ± 0.26 (± 95%CI) , whereas postnatal heart mass increased according to 10.0 Mb0.88 ± 0.10 . While the fetal and postnatal scaling exponents are identical (0.88) and reveal a clear dependence of heart mass on body mass, only the postnatal exponent is significantly less than 1.0, indicating the postnatal heart becomes a smaller component of body mass as the body grows, which is a pattern found frequently with postnatal cardiac development among mammals. The rapid doubling in heart mass around the time of birth is independent of any increase in body mass and is consistent with the normalization of wall stress in response to abrupt changes in volume loading and pressure loading at parturition. We discuss variation in scaling patterns of heart mass across development among mammals, and suggest that the variation results from a complex interplay between hard-wired genetics and epigenetic influences.

Finding the peak of dynamic oxygen uptake during fatiguing exercise in fish.

As fish approach fatigue at high water velocities in a critical swimming speed (Ucrit) test, their swimming mode and oxygen cascade typically move to an unsteady state because they adopt an unsteady, burst-and-glide swimming mode despite a constant, imposed workload. However, conventional rate of oxygen uptake (MO2 ) sampling intervals (5-20 min) tend to smooth any dynamic fluctuations in active MO2  (MO2active) and thus likely underestimate the peak MO2active Here, we used rainbow trout (Oncorhynchus mykiss) to explore the dynamic nature of MO2active near Ucrit using various sampling windows and an iterative algorithm. Compared with a conventional interval regression analysis of MO2active over a 10-min period, our new analytical approach generated a 23% higher peak MO2active Therefore, we suggest that accounting for such dynamics in MO2active with this new analytical approach may lead to more accurate estimates of maximum MO2  in fishes.

Autonomic cardiac regulation facilitates acute heat tolerance in rainbow trout: in situ and in vivo support.

Acute warming in fish increases heart rate (fH) and cardiac output to peak values, after which performance plateaus or declines and arrhythmia may occur. This cardiac response can place a convective limitation on systemic oxygen delivery at high temperatures. To test the hypothesis that autonomic cardiac regulation protects cardiac performance in rainbow trout during acute warming, we investigated adrenergic and cholinergic regulation during the onset and progression of cardiac limitations. We explored the direct effects of adrenergic stimulation by acutely warming an in situ working perfused heart until arrhythmia occurred, cooling the heart to restore rhythmicity and rewarming with increasing adrenergic stimulation. Adrenergic stimulation produced a clear, dose-dependent increase in the temperature and peak fH achieved prior to the onset of arrhythmia. To examine how this adrenergic protection functions in conjunction with cholinergic vagal inhibition in vivo, rainbow trout fitted with ECG electrodes were acutely warmed in a respirometer until they lost equilibrium (CTmax) with and without muscarinic (atropine) and ß-adrenergic (sotalol) antagonists. Trout exhibited roughly equal and opposing cholinergic and adrenergic tone on fH that persisted up to critical temperatures. ß-Adrenergic blockade significantly lowered peak fH by 14-17%, while muscarinic blockade significantly lowered the temperature for peak fH by 2.0°C. Moreover, muscarinic and ß-adrenergic blockers injected individually or together significantly reduced CTmax by up to 3°C, indicating for the first time that cardiac adrenergic stimulation and cholinergic inhibition can enhance acute heat tolerance in rainbow trout at the level of the heart and the whole animal.

Salmonid gene expression biomarkers indicative of physiological responses to changes in salinity and temperature, but not dissolved oxygen.

An organism's ability to respond effectively to environmental change is critical to its survival. Yet, life stage and overall condition can dictate tolerance thresholds to heightened environmental stressors, such that stress may not be equally felt across individuals and at all times. Also, the transcriptional responses induced by environmental changes can reflect both generalized responses as well as others that are highly specific to the type of change being experienced. Thus, if transcriptional biomarkers specific to a stressor, even under multi-stressor conditions, can be identified, the biomarkers could then be applied in natural environments to determine when and where an individual experiences such a stressor. Here, we experimentally challenged juvenile Chinook salmon (Oncorhynchus tshawytscha) to validate candidate gill gene expression biomarkers. A sophisticated experimental design manipulated salinity (freshwater, brackish water and seawater), temperature (10, 14 and 18°C) and dissolved oxygen (normoxia and hypoxia) in all 18 possible combinations for 6 days using separate trials for three smolt statuses (pre-smolt, smolt and de-smolt). In addition, changes in juvenile behaviour, plasma variables, gill Na+/K+-ATPase activity, body size, body morphology and skin pigmentation supplemented the gene expression responses. We identified biomarkers specific to salinity and temperature that transcended the multiple stressors, smolt status and mortality (live, dead and moribund). Similar biomarkers for dissolved oxygen were not identified. This work demonstrates the unique power of gene expression biomarkers to identify a specific stressor even under multi-stressor conditions, and we discuss our next steps for hypoxia biomarkers using an RNA-seq study.

Adrenergic and adenosinergic regulation of the cardiovascular system in an Antarctic icefish: Insight into central and peripheral determinants of cardiac output.

Icefishes characteristically lack the oxygen-binding protein haemoglobin and therefore are especially reliant on cardiovascular regulation to augment oxygen transport when oxygen demand increases, such as during activity and warming. Using both in vivo and in vitro experiments, we evaluated the roles for adrenaline and adenosine, two well-established cardio- and vasoactive molecules, in regulating the cardiovascular system of the blackfin icefish, Chaenocephalus aceratus. Despite increasing cardiac contractility (increasing twitch force and contraction kinetics in isometric myocardial strip preparations) and accelerating heart rate (ƒH), adrenaline (5 nmol kg-1 bolus intra-arterial injection) did not significantly increase cardiac output (Q̇) in vivo because it elicited a large decrease in vascular conductance (Gsys). In contrast, and despite preliminary data suggesting a direct negative inotropic effect of adenosine on isolated atria and little effect on isolated ventricle strips, adenosine (500 nmol kg-1) generated a large increase in Q̇ by increasing Gsys, a change reminiscent of that previously reported during both acute warming and invoked activity. Our data thus illustrate how Q̇ in C. aceratus may be much more dependent on peripheral control of vasomotor tone than direct regulation of the heart.