Environmental variation associated with overwintering elicits marked metabolic plasticity in a temperate salmonid, Salvelinus fontinalis.

Poleward winters commonly expose animals, including fish, to frigid temperatures and low food availability. Fishes that remain active over winter must therefore balance trade-offs between conserving energy and maintaining physiological performance in the cold, yet the extent and underlying mechanisms of these trade-offs are not well understood. We investigated the metabolic plasticity of brook char (Salvelinus fontinalis), a temperate salmonid, from the biochemical to whole-animal level in response to cold and food deprivation. Acute cooling (1°C day-1) from 14°C to 2°C had no effect on food consumption but reduced activity by 77%. We then assessed metabolic performance and demand over 90 days with exposure to warm (8°C) or cold winter (2°C) temperatures while fish were fed or starved. Resting metabolic rate (RMR) decreased substantially during initial cooling from 8°C to 2°C (Q10=4.2-4.5) but brook char exhibited remarkable thermal compensation during acclimation (Q10=1.4-1.6). Conversely, RMR was substantially lower (40-48%) in starved fish, conserving energy. Thus, the absolute magnitude of thermal plasticity may be masked or modified under food restriction. This reduction in RMR was associated with atrophy and decreases in in vivo protein synthesis rates, primarily in non-essential tissues. Remarkably, food deprivation had no effect on maximum oxygen uptake rates and thus aerobic capacity, supporting the notion that metabolic capacity can be decoupled from RMR in certain contexts. Overall, our study highlights the multi-faceted energetic flexibility of Salvelinus spp. that likely contributes to their success in harsh and variable environments and may be emblematic of winter-active fishes more broadly.

Coronary circulation enhances the aerobic performance of wild Pacific salmon.

Female Pacific salmon often experience higher mortality than males during their once-in-a-lifetime up-river spawning migration, particularly when exposed to secondary stressors (e.g. high temperatures). However, the underlying mechanisms remain unknown. One hypothesis is that female Pacific salmon hearts are more oxygen-limited than males and are less able to supply oxygen to the body's tissues during this demanding migration. Notably, female hearts have higher coronary blood flow, which could indicate a greater reliance on this oxygen source. Oxygen limitations can develop from naturally occurring coronary blockages (i.e., coronary arteriosclerosis) found in mature salmon hearts. If female hearts rely more heavily on coronary blood flow but experience similar arteriosclerosis levels as males, they will have disproportionately impaired aerobic performance. To test this hypothesis, we measured resting (RMR) and maximum metabolic rate (MMR), aerobic scope (AS) and acute upper thermal tolerance in coho salmon (Oncorhynchus kisutch) with an intact or artificially blocked coronary oxygen supply. We also assessed venous blood oxygen and chemistry (cortisol, ions, and metabolite concentrations) at different time intervals during recovery from exhaustive exercise. We found that coronary blockage impaired MMR, AS, and the partial pressure of oxygen in venous blood (PvO2) during exercise recovery but did not differ between sexes. Coronary ligation lowered acute upper thermal tolerance by 1.1°C. Though we did not find evidence of enhanced female reliance on coronary supply, our findings highlight the importance of coronary blood supply for mature wild salmon, where migration success may be linked to cardiac performance, particularly during warm water conditions.

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.

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.

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.

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 .

Anadromy and marine habitat use of Lake trout (Salvelinus namaycush) from the central Canadian Arctic.

Anadromy was documented in 16 lake trout, Salvelinus namaycush, from Canada's central Arctic using capture data and otolith microchemistry. For the first time, estuarine/marine habitat use was described for five individuals using acoustic telemetry. Age-at-first-migration to sea was variable (10-39 years) among individuals and most S. namaycush undertook multiple anadromous migrations within their lifetime. Telemetry data suggested that S. namaycush do not travel far into marine habitats and prefer surface waters (<2 m). These results further our collective understanding of the marine ecology of Arctic S. namaycush.

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.

Acute exposure to a common suspended sediment affects the swimming performance and physiology of juvenile salmonids.

To study the effects of an acute exposure to turbidity generated by suspended sediment, we examined swimming performance (Ucrit) and related metabolic parameters in individual and groups of juvenile trout at three different concentrations of calcium carbonate. To investigate differences among strains or provenience, we compared one strain of rainbow trout (Oncorhynchus mykiss; RBT) and one strain of brown trout (Salmo trutta; BNT) from a common hatchery and one RBT strain from a separate hatchery. In general, trout swum individually or in groups exhibited a decrease in Ucrit as turbidity increased. Both RBT strains were more similar to each other and were impaired to a larger extent in swimming performance than BNT, which was less impacted. For groups, indicators of aerobic metabolism were elevated while those of anaerobic metabolism were depressed. Specifically, citrate synthase activities and glucose levels tended to be greater while plasma lactate and LDH activities were reduced. Lactate and LDH levels in individually swum trout under sediment exposure suggest a greater similarity of fish from the same provenience. We suggest that acute exposures to environmentally relevant turbidities generated by fine suspended sediment may cause a reduced Ucrit, and that these changes may be related to changes in the utilization of aerobic and anaerobic pathways.

Microplastic and other anthropogenic microparticles in Arctic char (Salvelinus alpinus) and their coastal habitat: A first-look at a central Canadian Arctic commercial fishery.

In the recent monitoring guidelines released by the Arctic Monitoring and Assessment Program's Litter and Microplastic Expert Group, Arctic salmonids were recommended as an important species for monitoring plastics in Arctic ecosystems, with an emphasis on aligning microplastic sampling and analysis methods in Arctic fishes. This recommendation was based on the minimal documentation of microplastics in Northern fishes, especially Arctic salmonids. In response, we worked collaboratively with local partners to quantify and characterize microplastics in Arctic char, Salvelinus alpinus, and their habitats in a commercial fishery near Iqaluktuuttiaq (Cambridge Bay), Nunavut. We sampled Arctic char, surface water, and benthic sediments within their summer foraging habitat at Palik (Byron Bay). We found microplastics in 95 % of char with an average of 26 (SD ± 19) particles per individual. On average, surface water samples had 23 (SD ± 12) particles/L and benthic sediment <1 particles/gww. This is the first documentation of plastic pollution in Arctic char and their coastal habitats. Future work should evaluate seasonal, temporal and spatial trends for long-term monitoring of microplastics in Arctic fishes and their habitats.

Impairing cardiac oxygen supply in swimming coho salmon compromises their heart function and tolerance to acute warming.

Climatic warming elevates mortality for many salmonid populations during their physically challenging up-river spawning migrations, yet, the mechanisms underlying the increased mortality remain elusive. One hypothesis posits that a cardiac oxygen insufficiency impairs the heart's capacity to pump sufficient oxygen to body tissues to sustain up-river swimming, especially in warm water when oxygen availability declines and cardiac and whole-animal oxygen demand increases. We tested this hypothesis by measuring cardiac and metabolic (cardiorespiratory) performance, and assessing the upper thermal tolerance of coho salmon (Oncorhynchus kisutch) during sustained swimming and acute warming. By surgically ligating the coronary artery, which naturally accumulates arteriosclerotic lesions in migrating salmon, we partially impaired oxygen supply to the heart. Coronary ligation caused drastic cardiac impairment during swimming, even at benign temperatures, and substantially constrained cardiorespiratory performance during swimming and progressive warming compared to sham-operated control fish. Furthermore, upper thermal tolerance during swimming was markedly reduced (by 4.4 °C) following ligation. While the cardiorespiratory capacity of female salmon was generally lower at higher temperatures compared to males, upper thermal tolerance during swimming was similar between sexes within treatment groups. Cardiac oxygen supply is a crucial determinant for the migratory capacity of salmon facing climatic environmental warming.

A sudden change of heart: Warm acclimation can produce a rapid adjustment of maximum heart rate and cardiac thermal sensitivity in rainbow trout.

Warm acclimation in fish is often characterized by an increase in heat tolerance and a reduction in physiological rates to improve the scope to respond to additional challenges including further warming. The speed of these responses can determine their effectiveness. However, acclimation rates vary across levels of biological organization and are poorly understood in part because most research is conducted after an acclimation period of >3 weeks, when acclimation is presumed to be complete. Here we show that when rainbow trout were transferred from 10 to 18 °C, over 50% of the total reduction of maximum heart rate (ƒHmax) (i.e. the thermal compensation at moderate temperatures) occurred within 72 h, with further compensation occurring more gradually over the following 25 days. Also, the ability to increase ƒHmax with acute warming improved within 24 h resulting in a 30% rise in peak ƒHmax, but this ultimately declined again with prolonged (28 days) exposure to 18 °C. In contrast with some previous studies, upper critical temperatures for ƒHmax did not increase. Nonetheless, we demonstrate that rapid cardiac plasticity is possible in rainbow trout and likely blunts the impacts of thermal variation over relatively short timescales, such as that associated with heat waves and migration between water bodies.

The thermal limits of cardiorespiratory performance in anadromous Arctic char (Salvelinus alpinus): a field-based investigation using a remote mobile laboratory.

Despite immense concern over amplified warming in the Arctic, physiological research to address related conservation issues for valuable cold-adapted fish, such as the Arctic char (Salvelinus alpinus), is lacking. This crucial knowledge gap is largely attributable to the practical and logistical challenges of conducting sensitive physiological investigations in remote field settings. Here, we used an innovative, mobile aquatic-research laboratory to assess the effects of temperature on aerobic metabolism and maximum heart rate (f Hmax) of upriver migrating Arctic char in the Kitikmeot region of Nunavut in the central Canadian Arctic. Absolute aerobic scope was unchanged at temperatures from 4 to 16°C, while f Hmax increased with temperature (Q 10 = 2.1), as expected. However, f Hmax fell precipitously below 4°C and it began to plateau above ~ 16°C, reaching a maximum at ~ 19°C before declining and becoming arrhythmic at ~ 21°C. Furthermore, recovery from exhaustive exercise appeared to be critically impaired above 16°C. The broad thermal range (~4-16°C) for increasing f Hmax and maintaining absolute aerobic scope matches river temperatures commonly encountered by migrating Arctic char in this region. Nevertheless, river temperatures can exceed 20°C during warm events and our results confirm that such temperatures would limit exercise performance and thus impair migration in this species. Thus, unless Arctic char can rapidly acclimatize or alter its migration timing or location, which are both open questions, these impairments would likely impact population persistence and reduce lifetime fitness. As such, future conservation efforts should work towards quantifying and accounting for the impacts of warming, variable river temperatures on migration and reproductive success.

Zebrafish (Danio rerio) as a model for the study of aging and exercise: physical ability and trainability decrease with age.

A rapidly aging global population has motivated the development and use of models for human aging. Studies on aging have shown parallels between zebrafish and humans at the internal organization level; however, few parallels have been studied at the whole-organism level. Furthermore, the effectiveness of exercise as a method to mitigate the effects of aging has not been studied in zebrafish. We investigated the effects of aging and intermittent exercise on swimming performance, kinematics and behavior. Young, middle-aged and old zebrafish (20-29, 36-48 and 60-71% of average lifespan, respectively) were exercised to exhaustion in endurance and sprint swimming tests once a week for four weeks. Both endurance and sprint performance decreased with increased age. Swimming performance improved with exercise training in young and middle-aged zebrafish, but not in old zebrafish. Tail-beat amplitude, which is akin to stride length in humans, increased for all age groups with training. Zebrafish turning frequency, which is an indicator of routine activity, decreased with age but showed no change with exercise. In sum, our results show that zebrafish exhibit a decline in whole-organism performance and trainability with age. These findings closely resemble the senescence-related declines in physical ability experienced by humans and mammalian aging models and therefore support the use of zebrafish as a model for human exercise and aging.

Growth differentiation factor 6 as a putative risk factor in neuromuscular degeneration.

Mutation of Glass bottom boat, the Drosophila homologue of the bone morphogenetic protein or growth/differentiation factor (BMP/GDF) family of genes in vertebrates, has been shown to disrupt development of neuromuscular junctions (NMJ). Here we tested whether this same conclusion can be broadened to vertebrate BMP/GDF genes. This analysis was also extended to consider whether such genes are required for NMJ maintenance in post-larval stages, as this would argue that BMP genes are viable candidates for analysis in progressive neuromuscular disease. Zebrafish mutants harboring homozygous null mutations in the BMP-family gene gdf6a were raised to adulthood and assessed for neuromuscular deficits. Fish lacking gdf6a exhibited decreased endurance (∼ 50%, p = 0.005) compared to wild type, and this deficit progressively worsened with age. These fish also presented with significantly disrupted NMJ morphology (p = 0.009), and a lower abundance of spinal motor neurons (∼ 50%, p<0.001) compared to wild type. Noting the similarity of these symptoms to those of Amyotrophic Lateral Sclerosis (ALS) model mice and fish, we asked if mutations in gdf6a would enhance the phenotypes observed in the latter, i.e. in zebrafish over-expressing mutant Superoxide Dismutase 1 (SOD1). Amongst younger adult fish only bigenic fish harboring both the SOD1 transgene and gdf6a mutations, but not siblings with other combinations of these gene modifications, displayed significantly reduced endurance (75%, p<0.05) and strength/power (75%, p<0.05), as well as disrupted NMJ morphology (p<0.001) compared to wild type siblings. Bigenic fish also had lower survival rates compared to other genotypes. Thus conclusions regarding a role for BMP ligands in effecting NMJ can be extended to vertebrates, supporting conservation of mechanisms relevant to neuromuscular degenerative diseases. These conclusions synergize with past findings to argue for further analysis of GDF6 and other BMP genes as modifier loci, potentially affecting susceptibility to ALS and perhaps a broader suite of neurodegenerative diseases.