Anthony Farrell

Professor

Relevant Degree Programs

 
 

Graduate Student Supervision

Doctoral Student Supervision (Jan 2008 - May 2021)
Interpreting species, intraspecific and intra-individual variability by comprehensively characterizing a fish's respiratory phenotype with valid measures of oxygen uptake (2021)

Vertebrate life is sustained by aerobic metabolism and temporary bouts of anaerobic metabolism. Steady-state aerobic metabolism can be assessed by measuring whole-organism oxygen uptake (ṀO₂), the core of Fry’s paradigm, which becomes a part of a respiratory phenotype that also includes anaerobic capacity. In fishes, environment profoundly affects respiratory phenotype, perhaps even determining migratory and reproductive successes, partially driven by hypoxia, increased thermal and pathogen load. Understanding how such factors affect capacity, functional costs and mortality needs a more accurate, precise, and comprehensive characterization of a fish’s respiratory phenotype. Hence, I developed an Integrated Respiratory Assessment Paradigm (IRAP) to characterize a respiratory phenotype (its needs, capacities & patterns) for individual fish that responds to Fry’s paradigm (controlling, limiting, masking, directive, and lethal factors). Chapter 2 describes an IRAP protocol that measures 14 metrics in 8 fish simultaneously over 3 days, a high sampling throughput to better bridge physiology-&-ecology. Chapter 3 introduces and validates new analytical approaches that improved the resolution and accuracy of maximum ṀO₂ (ṀO₂max) estimations in aquatic respirometry. Chapter 4 illustrates the detection sensitivity of the refined IRAP by distinguishing respiratory phenotypes among three rainbow trout strains using differences in ṀO₂max, absolute aerobic capacity, critical oxygen saturation (O₂crit) and hypoxia tolerance. Moreover, high aerobic and anaerobic capacities correlated with high hypoxic and thermal tolerances among these strains. Chapter 5 explores how acclimation to hypoxia affects the respiratory phenotype, discovering hypoxia-acclimated European sea bass suppress standard metabolic rate (SMR) and O₂crit, a respiratory phenotype better suited for exploiting hypoxic habitats. Chapter 6 explores the diagnostic power of the refined IRAP by assessing the load-dependent changes to SMR, O₂crit, daily energy expenditure and routine ṀO₂ of a high-virulence infectious hematopoietic necrosis virus infection in sockeye salmon and a low-virulence piscine orthoreovirus infection in sockeye and Atlantic salmon. Chapter 7 assesses the repeatability of IRAP indices, showing four traits for aerobic capacity, routine ṀO₂ and time spend above 50% absolute aerobic scope were particularly repeatable. Chapter 8, therefore, synthesizes that high-resolution and reproducible metabolic phenotyping better poises ichthyologists to develop an individual-based metabolic model and integrate it with a Genomic-Physiology-Ecology axis.

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Mechanisms of cardiac pacemaking and temperature-dependent depression of cardiac electrical excitation in the zebrafish (danio rerio) (2021)

The physiology of ectotherms is profoundly affected by the environmental temperature which governs the rate of physiological processes. Cardiac function, an essential function of all vertebrates, is no exception: during warming heart rate tracks temperature before declining at temperatures beyond maximum optimal temperature, ultimately collapsing with further warming. In fishes, like other vertebrates, intrinsic heart rate is set by pacemaker cells located in the sino-atrial node that spontaneously generate action potentials. At the cellular level, temperature-dependent deterioration of pacemaking mechanisms may contribute to the decline of cardiac function. Nevertheless, the mechanisms of pacemaking and their temperature-dependent deterioration remain elusive. Hence, I explored cardiac pacemaking mechanisms in zebrafish, as well as their relative thermal performance and limits.I validated blebbistatin as an effective excitation-contraction uncoupling agent that did not modify the cardiac action potential properties, thus providing an essential methodology for future cardiac pacemaking research enabling the direct recording of intracellular electrical activity of pacemaker cells. Using electrocardiograms, I confirmed that cardiac pacemaking involves two major mechanisms. Pharmacological blockade of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels with zatebradine reduced heart rate by up to 60%, suggesting HCN channels play the major role in cardiac pacemaking. Likewise, sarcoplasmic reticulum (SR) calcium cycling was pharmacologically blocked using ryanodine and thapsigargin to block ryanodine receptors and SERCA pumps, respectively, which reduced heart rate by ~40%, suggesting the SR plays a secondarily important role in pacemaking. However, the combination of these pharmacological interventions did not completely stop the heartbeat, suggesting that either mammalian pharmacological agents are less effective in producing total Hcn block in zebrafish, perhaps due to isoform specificity.HCN4, the major HCN channel involved in mammalian pacemaking, was knocked out using CRISPR to explore its role in zebrafish cardiac pacemaking. Heart rate did not differ significantly between mutant and control fish at any test temperature, including fish treated with inhibitors of HCN channels or SR calcium cycling. Thus, alternative Hcn channels compensated for the knockout of Hcn4, presumably contributing to a higher thermal tolerance. In addition, mutant fish had a higher upper thermal tolerance than control fish when SR calcium cycling was inhibited.

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Thermal limits to the cardiorespiratory performance of Arctic char (Salvelinus alpinus) in a rapidly warming north (2020)

The Canadian Arctic is warming at nearly three times the global rate. Consequently, thermal regimes of native cold-adapted species like the Arctic char (Salvelinus alpinus) are being rapidly reshaped. The Arctic char is the most northerly-distributed freshwater fish on Earth and is essential to Inuit food security and culture. Anadromous Arctic char migrate between freshwater habitats and the Arctic Ocean many times throughout their lives, which can expose them to an already extreme range of temperatures (21°C). My thesis examined the ability of Arctic char to cope with thermal variation, focusing specifically on cardiorespiratory performance.I used a novel, mobile laboratory in the central Canadian Arctic to assess how acute temperature changes impact cardiac function and aerobic metabolism in migrating Arctic char. Arctic char maintained aerobic performance over an impressive temperature range (4-16°C), but could not recover from exhaustive exercise above 16°C. Furthermore, maximum heart rate, which should increase with acute warming, began to plateau at 16°C, declined at 19°C and became arrhythmic at 21°C. I conducted similar assessments on four other populations to determine how they differed. One population that undertakes physically and thermally challenging migrations had higher maximum aerobic capacity and heat tolerance than another with less harsh migrations. Heart mass was also higher in populations with less challenging migrations. Next, I acclimated hatchery-reared Arctic char to naturally encountered temperatures (2-18°C) to characterize their thermal plasticity. As before, Arctic char maintained high aerobic performance over a broad temperature range (2-14°C) and warm acclimation improved swimming performance and dramatically increased (+35-45%) cardiac heat tolerance. However, mortality occurred with chronic exposure to 18°C. My research indicates that Arctic char have broad thermal performance and acclimation potential that may help mitigate the negative impacts of rapid environmental warming. Nevertheless, I showed that Arctic char already encounter temperatures in the Canadian Arctic that constrain cardiorespiratory performance and impair recovery from exhaustive exercise, which would likely hinder their obligatory return migration. Furthermore, my results suggest that intraspecific diversity in cardiorespiratory physiology, thermal tolerance, and local migration conditions may be important for Arctic char conservation and shape population-specific responses to warming.

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Physiological, Transcriptomic and Genomic Mechanisms of Thermal Adaptation in Oncorhynchus mykiss (2017)

Given the rate and magnitude of the ongoing global warming, there is some urgency to understand the underlying mechanism of thermal adaptation to evaluate and predict the ecological consequences. This thesis used Oncorhynchus mykiss from different thermal regimes to examine thermal adaptation at physiological, transcriptomic and genomic levels. Thermal tolerance was examined in three redband trout (O. mykiss gairdneri) populations from warm desert and cool montane climates (Idaho, USA), as well as in a domesticated rainbow trout (O. mykiss) strain raised in a thermally challenging environment for over 19 generations (Western Australia, Australia). Acclimated to 15°C, the desert redband trout had the highest critical thermal maximum (CTMAX; 29.7°C) and maintained an almost constant absolute aerobic scope (AAS) across a broader range of test temperatures (12-24°C) than seen in other strains, but had the lowest peak AAS, suggesting a tradeoff between thermal performance and tolerance. Western Australian rainbow trout had the highest AAS, even when tested at 21°C, which may be a result of hatchery selection for thermal performance. Although the rate transition temperatures for maximum heart rate (Arrhenius breakpoint and arrhythmia temperature for fH,max) were similar among all populations, fH,max was the highest in the desert redband trout population at all temperatures. Cardiac RNA sequencing revealed different patterns of gene regulation among redband trout populations during acute warming. Many genes had different mRNA abundances between populations due to constitutive and induced expression, and the number of differentially expressed genes among populations was positively correlated to the genetic distance, suggesting intraspecific cellular regulatory strategies in response to acute warming. Population and quantitative genetic studies identified potential genomic markers for thermal adaptation. A total of twenty-one loci were putatively under positive thermal selection (“outliers”). In addition, genotypes of some outlier loci had significantly different CTMAX. Genome-wide association study identified twelve loci that were significantly associated with individual CTMAX. Altogether, results in my dissertation demonstrated the capacity of thermal adaptation in O. mykiss populations at multiple organismal levels. This data lays a foundation to improve our understanding on the potential impact of global warming on wild aquatic populations.

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Upper thermal limits and acclimation potential of Arctic cod (Boreogadus saida): a key food web species in the Arctic Ocean (2017)

The recent rapid and unprecedented changes to the physical and biogeochemical properties of the Arctic Ocean have gained worldwide attention. The greater than 50% reduction in sea ice volume below average is of great concern. My thesis investigates the potential effects of a warmer Arctic Ocean upon the indigenous Arctic cod, Boreogadus saida. This fish make up a significant proportion of the lower trophic energy reserve available in Arctic marine systems. Predators rely upon Arctic cod to provide them bite sized access to those critical energy reserves. Yet despite their key role in the Arctic food web, their upper thermal limits are not well studied. Thus, my three objectives were: a) quantify, for the first time in this species, their upper thermal limits b) determine the acclimation potential of the species at three acclimation temperatures and c) contrast the results generated by different methods that quantify thermal limits for declining physiological performance (rate transition temperatures) that I determined.Boreogadus saida upper thermal limits were tested under acute warming conditions using three different methods: loss of equilibrium (Tcmax), absolute aerobic scope (AAS) derived from oxygen uptake rates and the cardiac method, which uses maximum heart rate (ƒHmax) to detect change in whole animal performance. In conducting the thermal acclimation studies, I discovered foremost that 6.5°C-acclimated fish grew at that temperature but, to date, have only produced eggs at 3.5°C water temperatures. The Tcmax significantly increased with acclimation temperature (0.5, 3.5 and 6.5°C) from 14.4, 15.5, up to 17.1°C respectively, while the more ecologically relevant AAS transition temperature limits were found at lower temperatures from 1.0 to 5.5°C. The temperature for peak ƒHmax (Tmax) occurred between 11.0 to 12.0°C and the performance of ƒHmax for larval B. saida during acute warming was not significantly different from the adults until Tmax was reached. This novel study of the thermal physiology of this key Arctic marine food web species revealed a greater than expected thermal tolerance and a significant acclimation potential up to 6.5°C, suggesting that this species may be more resilient to rapid climate change than previously thought.

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Cardiorespiratory Responses to Hypoxia in High- and Low-Altitude Geese and Ducks (2016)

High-altitude (HA) life is challenging due to the reduced partial pressure of oxygen (hypoxia). Hence, HA vertebrates have evolved increased capacities in their oxygen transport cascade enhancing oxygen transfer. The extent of interspecies variation in these responses within waterfowl, a taxon prolific at HA, remains largely unknown. This thesis investigated 17 waterfowl groups at different altitudes to address the overarching hypotheses that waterfowl use multiple cardiorespiratory strategies to maintain oxygen supply during hypoxia, and that HA exposure alters the waterfowl hypoxic ventilatory and cardiovascular responses. A comprehensive analysis of metabolic, cardiovascular, and ventilatory responses to progressive decreases in equivalent fractional composition of inspired oxygen was made on resting low-altitude (LA) barnacle geese, LA bar-headed geese, HA bar-headed geese, Andean geese, and crested ducks. Andean geese and crested ducks, lifelong HA residents, exhibited fundamentally different mechanisms for maintaining oxygen supply during hypoxia than bar-headed geese, transient HA migrators. Bar-headed geese robustly increased ventilation and heart rate, whereas Andean species increased lung oxygen extraction and stroke volume. Also, HA-reared bar-headed geese exhibited reduced oxygen consumption during hypoxia compared to LA-reared bar-headed geese. Similar cardiovascular studies were performed on five HA duck species (yellow-billed pintail, cinnamon teal, ruddy duck, speckled teal, and Puna teal) in Peru and six related LA duck species (northern pintail, cinnamon teal, ruddy duck, green-winged teal, gadwall, and mallard duck) in the USA. Heart rate and oxygen pulse remained generally unchanged. Instead, most HA ducks exhibited higher blood-oxygen carrying capacity and lower heart rate variability than LA ducks. While heart rate, stroke volume, oxygen pulse, and blood-oxygen carrying capacity contributed to all 17 groups’ hypoxic cardiovascular responses, the predominant responses were increased stroke volume and, in HA taxa, blood-oxygen carrying capacity. Only bar-headed geese increased heart rate appreciably.This thesis identifies multiple cardiovascular and respiratory strategies by which waterfowl maintain oxygen supply during hypoxia, and provides insight into how HA rearing impacts these responses. This thesis also suggests that short-term HA performance utilizes primarily functional enhancements (e.g. rapid heart rate and ventilation increases), whereas lifelong HA residency is supported predominantly by structural changes (e.g. lung and cardiac morphology enhancements).

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The functional significance and evolution of the coronary circulation in sharks (2016)

The coronary circulation first appeared in the chordate lineage in cartilaginous fishes where it perfuses the entire myocardium, just like in birds and mammals but unlike in most teleost fishes. Yet, despite the pivotal position of elasmobranchs in the evolution of the coronary oxygen supply, the functional significance of their coronary circulation has never been investigated. Elasmobranchs are of special interest because of the morphological arrangement of their cardiomyocytes, which has resulted in the majority of the ventricular myocardium having access to oxygen from both a coronary supply and the venous blood returning to the heart. In order to determine the relative contribution of the coronary oxygen supply to cardiovascular function, I measured coronary blood flow (CBF) in the sandbar shark, Carcharhinus plumbeus, and leopard shark, Triakis semifasciata, while manipulating cardiovascular status using pharmacological approaches and in vivo temperature changes, respectively. By exploring inter- and intra-individual variation in cardiovascular variables I show that coronary blood flow is directly related to heart rate in both bradycardic (R²= 0.6, P
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Cardiac Control in the Pacific Hagfish (Eptatretus Stoutii) (2014)

The Pacific hagfish (Eptatretus stoutii), being an extant ancestral craniate, possesses the most ancestral craniate-type heart with valved chambers, a response to increased filling pressure with increased stroke volume (Frank-Starling mechanism), and myogenic contractions. Unlike all other known craniate hearts, this heart receives no direct neural stimulation. Despite this, heart rate can vary four-fold during a prolonged, 36-h anoxic challenge followed by a normoxic recovery period, with heart rate decreasing in anoxia, and increasing beyond routine rates during recovery, a remarkable feat for an aneural heart. This thesis is a study of how the hagfish can regulate heart rate without the assistance of neural stimulation.A major role of hyperpolarization-activated cyclic nucleotide-activated (HCN) channels in heartbeat initiation was indicated by pharmacological application of zatebradine to spontaneously contracting, isolated hearts, which stopped atrial contraction and vastly reduced ventricular contraction. Tetrodotoxin inhibition of voltage-gated Na⁺ channels induced an atrioventricular block suggesting these channels play a role in cardiac conduction.Partial cloning of HCN channel mRNA extracted from hagfish hearts revealed six HCN isoforms, two hagfish representatives of vertebrate HCN2 (HCN2a and HCN2b), three of HCN3 (HCN3a, HCN3b and HCN3c) and one HCN4. Two paralogs of HCN3b were discovered, however, HCN3a dominated the expression of HCN isoforms followed by HCN4. All HCN isoforms bar HCN3b were dominantly expressed in the atrium, likely to support greater atrial excitability ensuring synchronous contractions. Phylogenetic analysis suggested that HCN3 is the ancestral isoform supporting previous observations.Studies with β-adrenoreceptor agonists and antagonists in isolated, spontaneously beating hearts showed that the routine normoxic heart rate may involve maximal catecholamine stimulation of heart rate through cAMP stimulation of HCN channels via transmembrane adenylyl cyclase (tmAC). Loss of this tonic β-adrenorecptor cardiac stimulation during anoxia reduces heart rate, but restoring β-adrenoreceptor stimulation during normoxic recovery does not produce the previously observed increase above routine heart rate in vivo. Instead, bicarbonate-stimulated, soluble adenylyl cyclase (sAC) mediated cAMP production was found to produce this tachycardia in addition to the reinstated tmAC produced cAMP. This is the first time sAC has been implicated in heart rate control.

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Oxygen supply in rainbow trout (Oncorhynchus mykiss) and its ecological impacts: an investigation of poor triploid performance (2013)

Acquisition of environmental O₂ and its delivery throughout the body is essential for vertebrates and dictates habitat, life style, variation in anatomical form and function and even survival. Triploid (3N) rainbow trout (Oncorhynchus mykiss), which are important to the $0.5 billion British Columbia sport fishing industry, provide an informative model organism to study corporeal O₂ supply limitations and associated effects on survival in the wild. Triploid O₂ supply limitations likely stem from enlarged cells and contribute to poor 3N tolerance of sub-optimal conditions, which, in turn, may lead to high 3N population-level mortality rates in nature.Therefore, in order to test the hypotheses that corporeal O₂ supply limits aerobic performance of 3N rainbow trout and that aerobic performance, in turn, limits survival in the wild, I compared the cardiorespiratory physiology of diploid (2N) and 3N Blackwater River rainbow trout facing a high temperature challenge in the lab and survival in the wild. I then investigated the potential of a 3N cardiac O₂ supply deficiency, using a modified Krogh diffusion model, and discussed its significance to temperature tolerance, endurance swimming and survival in the wild. Both of my hypotheses were supported. A slower increase in 3N heart rate with warming suggested reduced O₂ convection through the body of 3N fish at high temperatures. Relating these results and endurance swimming rank with survival and habitat utilization in lakes revealed thermal tolerance and aerobic capacity as important variables influencing lake survival. The Krogh model showed 3N relative to 2N cardiac O₂ supply limitations that were primarily driven by reduced 3N arterial O₂ content, which I showed not to be caused by reduced 3N haemoglobin - O₂ affinity. In supporting the 2 main hypotheses of my thesis, this theoretically predicted 3N cardiac O₂ supply deficiency may explain reduced 3N aerobic swimming capacity and heart rate response to warming. Thus, my findings are consistent with corporeal O₂ supply limitations to high temperature tolerance and aerobic swimming capacity of 3N rainbow trout, both of which can limit survival in the wild, depending on the biotic and abiotic conditions and physiological state of the organism.

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Cardiorespiratory physiology and temperature tolerance among populations of sockeye salmon (Oncorhynchus nerka) (2011)

Elevated summer water temperature has been associated with high mortality in adult sockeye salmon (Oncorhynchus nerka) during their once-in-a-lifetime migration up the Fraser River (British Columbia, Canada) to their spawning grounds. There are over 100 genetically distinct populations of sockeye salmon in the Fraser River watershed, varying in migration distance, elevation gain, river temperature and river flow. This thesis studied the physiological basis for temperature tolerance in sockeye salmon and examined the overall hypothesis that each sockeye salmon population has physiologically adapted to meet their specific upriver migration conditions.Swimming and cardiorespiratory performance were compared over a range of temperatures across six wild, migrating adult sockeye salmon populations. All populations maintained maximum performance across the entire range of temperatures typically encountered during their upriver migration, with Chilko sockeye salmon emerging as the most high temperature-tolerant. In addition, populations with more challenging migrations had greater aerobic scope, larger hearts and improved coronary supply. These results suggest that sockeye salmon populations have physiologically adapted to cope with their local upriver migration conditions, despite never before having performed the upriver migration. Temperatures exceeding the population-specific thermal optimum resulted in severely impaired aerobic scope and swimming performance. This study suggests that population-specific thermal limits are set by physiological limitations in aerobic performance. Specifically, fish may be unable to swim at warm temperature due to insufficient oxygen supply to meet demand, triggered via a cardiac limitation due to reduced scope for heart rate.Given the key role of the heart in limiting thermal tolerance, the role of cardiac adrenergic stimulation was examined as a potential mechanism underlying the observed differences in thermal tolerance across sockeye salmon populations. Chilko sockeye salmon had a greater density of ventricular β-adrenoceptors, which may provide greater cardiac capacity and protection at temperature extremes, thereby expanding their breadth of thermal tolerance compared to other populations. This thesis suggests that sockeye salmon populations will be differentially affected by warming river temperatures, raising conservation concerns for biodiversity. This work provides important insight into local adaptation in sockeye salmon and identifies a possible cause for in-river mortality associated with warm temperatures in sockeye salmon.

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Master's Student Supervision (2010 - 2020)
Thermal acclimation potential of Australian rainbow trout, Oncorhynchus mykiss (2020)

With impending global warming predictions, surviving animals will need to either escape increasing temperatures in their current biogeography, acclimate or adapt. As temperature increases above the optimal range of an animal, physiological performance is negatively affected, which results in decreased growth, feeding, reproduction and aerobic scope. My thesis studied a hatchery-raised strain of rainbow trout (Oncorhynchus mykiss, H-strain) previously selected for upper thermal tolerance at the Pemberton Freshwater Research Centre (PFRC) in Australia to understand their acclimation potential. This H-strain of rainbow trout was acclimated to six experimental temperatures (15, 17, 19, 21, 23, 25˚C) for over one month before performing a range of tests at each acclimation temperature that determined the optimal temperatures, or acclimation potential, for growth, digestibility (specific dynamic action; SDA), feed conversion (feed conversion ratio; FCR), aerobic performance and the response of maximum heart rate (fHmax) to acute warming. Intermittent-flow respirometry was used to determine respiratory oxygen uptake for analysis of SDA, standard metabolic rate (SMR), maximum oxygen uptake (ṀO₂max), excess-post exercise oxygen consumption (EPOC) and hypoxia tolerance (ILOS). Growth, feed efficiency, SDA duration and fHmax had acclimation potential up to 23˚C. ṀO₂max was also maintained up to 23˚C, while SMR followed a typical exponential increase; the calculated difference between ṀO₂max and SMR is absolute aerobic scope (AAS), which had an acclimation potential up to 21˚C. With acute warming, the critical thermal maximum (CTmax) plateaued at an acclimation temperature of 23˚C (reaching 31.2˚C), and the temperature of fHmax and arrhythmia had acclimation potential up to 23˚C. This integrative approach in assessing physiological performance illustrates that this warm-tolerant strain of rainbow trout has a broad thermal range for performance and a large-scale consideration of the thermal tolerance of other strains of rainbow trout is warranted given that rainbow trout are typically considered a cold-water species.

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Beta-adrenergic function in juvenile sockeye salmon hearts (2019)

Adult Fraser River sockeye salmon (Oncorhynchus nerka) show population level tailoring of their cardiorespiratory system specific to upriver migration challenges. My thesis sought to expand our knowledge to juvenile sockeye salmon populations by studying stimulation of their cardiac performance via β-adrenergic receptors. I made measurements of myocardial cell-surface β2-adrenoceptor density in fish hearts 14-times smaller than previously accomplished by modifying and validating the tritiated ligand technique. For the Chilko sockeye salmon population, smolts had about half the receptor density of adults, but still twice that of adult hatchery O. mykiss. With my new technique, cardiac receptor density can now be investigated in a much wider range of fish species and life stages.I also investigated the effects of acclimation temperature on myocardial β-adrenergic stimulation (βAS) in juvenile sockeye by studying in vitro ventricular preparations that developed maximum isometric tension over a range of pacing frequencies (a force-frequency relationship – FFR) at both 5 °C (0.2 – 0.8 Hz) and 14°C (0.2 – 1.6 Hz). I compared juveniles from the Chilko River and Weaver Creek populations raised in a common-garden laboratory rearing environment (captive-reared) and wild Chilko juveniles captured and acclimated to the laboratory environment (wild-reared). Under tonic βAS (0.01 µM isoprenaline), active tension at 5 °C was either unchanged by pacing frequency (both Chilko populations), or modestly biphasic FFR (Weaver), whereas at 14 °C all three study groups had a negative FFR. Maximal βAS (32 µM isoprenaline) at 0.2 Hz doubled active tension in all study groups and at both temperatures, and all study groups had a negative FFR independent of temperature. However, only at 14 °C was active tension under maximal βAS greater than under tonic βAS for the highest and physiologically relevant pacing frequencies. Importantly, other than the modest biphasic FFR of Weaver Creek, the FFR did not differ appreciably between juveniles of two Fraser River sockeye populations.

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Intrinsic heart rate resetting and associated changes in cardiac mRNA expression with thermal acclimation in rainbow trout, oncorhynchus mykiss (2019)

The original aim of my thesis was to follow the timescale of intrinsic heart rate resetting in rainbow trout. Groups of rainbow trout acclimated to either 4°C or 12°C were reciprocally transferred to follow intrinsic heart rate resetting. However, while one group did reset intrinsic heart rate after 1 h at 12°C from 56.8 ± 1.2 to 50.8 ± 1.5 bpm, and after 8 h at 4°C from 33.4 ± 0.7 to 37.7 ± 1.2 bpm; another group did not reset intrinsic heart rate, even after 10 weeks of either warm or cold acclimation. Even though this variation in intrinsic heart rate resetting was unexpected, this serendipitous discovery created an opportunity to better associate changes in mRNA expression specifically with intrinsic heart rate resetting responses, as opposed to more general responses to warm acclimation. Therefore, I used a Fluidigm qRT-PCR system to compare mRNA expression of 28 cardiac function proteins after warm acclimation in three different cardiac tissues (sino-atrial node [SAN], atrium and ventricle) for fish with, and without, an intrinsic heart rate resetting response. When mRNA expression of three cardiac tissues was compared under control conditions at 4°C, the SAN had a uniquely higher HCN1, Cav1.3 and collagen 1α1 expression, while the ventricle had a uniquely higher RYR3 and NKA α3 expression. After > 3 weeks of warm acclimation, downregulation of NKA α1c in the atrium and ventricle, and upregulation of HCN1 in the ventricle were discovered in fish with an intrinsic heart rate resetting response, whereas upregulation of HCN3 in the SAN and atrium was discovered in fish without an intrinsic heart rate resetting response. However, no mRNA expression changes were observed after just 1 h of warm acclimation. In conclusion, while initial intrinsic heart rate resetting does not involve changes in mRNA expression, changes in mRNA expression are apparent over the longer term and may be associated with the different intrinsic heart rate resetting responses. Moreover, my results suggest that while intrinsic heart rate resetting can be fast, it cannot be assumed to always occur in rainbow trout.

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Effects of bicarbonate on cardiac function in fish (2018)

An entirely novel mechanism to modulate heart rate was recently discovered in the Pacific hagfish (Eptatretus stoutii): a soluble adenylyl cyclase (sAC)-mediated pathway that increases cyclic adenosine monophosphate (cAMP) production upon stimulation by HCO₃₋ to increase heart rate. However, still unknown is whether this cardiac control pathway is present in other species as well. The objective of my study was to determine the effects of increasing extracellular [HCO₃₋] on the in vitro cardiac function of other fish species and whether the sAC-mediated pathway is associated with recovery of cardiac function during debilitating conditions. Exposure to severe hypoxia (100% N₂) and hypercapnic acidosis (7.5% or 15% CO₂) significantly decreased the heart rate of isolated, freely beating hearts and reduced the isometric tension (contractility) of electrically paced ventricular strips from Pacific lamprey (Lampetra richardsoni), Pacific spiny dogfish (Squalus suckleyi), Asian swamp eel (Monopterus albus), white sturgeon (Acipenser transmontanus), zebrafish (Danio rerio), and starry flounder (Platichthys stellatus). Spontaneous recovery in heart rate or contractility was not observed during severe hypoxia or hypercapnic acidosis for any of the species tested. Addition of HCO₃₋ (up to 50 mM) was associated with a complete and dose-dependent recovery of control heart rate in lamprey, dogfish, and swamp eel hearts during severe hypoxia, and in dogfish, sturgeon, and swamp eel hearts during hypercapnic acidosis. A partial recovery of control heart rate was observed in lamprey and zebrafish hearts during hypercapnic acidosis. However, HCO₃₋ had no effect on the heart rate or contractility in flounder hearts and had little to no effect on restoring control contractility in dogfish, swamp eel, and flounder ventricular strips.The addition of KH7 (sAC blocker) abolished the HCO₃₋-induced recovery of heart rate during severe hypoxia only in the lamprey heart. Thus, the sAC-mediated pathway in cardiac control appears to be unique to the cyclostomes and not present in the other species tested. While the sAC-mediated pathway was associated with the recovery of heart rate in the lamprey heart, the specific mechanisms behind how HCO₃₋ was associated with the recovery of heart rate in the other species still needs to be determined.

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Estimating aerobic and anaerobic capacities using the respiratory assessment paradigm: a validation using Atlantic salmon (Salmo salar) and European sea bass (Dicentrarchus Labrax) (2016)

Assessing fish performance, whether for extrapolation to natural conditions, culture conditions or toxicology, has a long history of using swim performance, metabolic rates and performance in hypoxia. However, rarely are these metrics unified in a test. Hence, thesis’ objective was to test a respiratory assessment paradigm (RAP), which comprehensively evaluated aerobic and anaerobic metabolism by explicitly measuring aerobic capacity, recovery and performance in hypoxia. To provide high fish throughput with each test, RAP used multiple intermittent-flow respirometry systems. I tested RAP by examining two practical questions: do aquaculture practices compromise the respiratory robustness of domesticated Atlantic salmon (Salmo salar) and, if so, how might it be reversed? Second, does an acute sub-lethal exposure to chemically dispersed oil have a chronic residual respiratory effect on European sea bass (Dicentrarchus labrax).RAP revealed a domesticated (Bolaks) strain of Atlantic salmon had a lower aerobic capacity than a wild (Lærdal) strain. Incremental exercise training significantly increased aerobic and recovery capacities in only the Lærdal strain. Thus, the Bolaks strain appeared to be athletically less robust than Lærdal strain. As a conclusion, ten generations of growth-oriented breeding program of Bolaks strain in commercial aquaculture have reduced its athletic robustness and cardiorespiratory plasticity as compared to their wild conspecifics. Given the success in improving athletic robustness of the wild strain, it still remains to explore whether an exercise- training protocol can be developed that will provide benefits to the salmon aquaculture industry. RAP discovered no residual effect of oil on aerobic capacity for either hypoxia tolerant (HT) or hypoxia sensitive (HS) sea bass. Instead, RAP discovered a residual effect for only the HT phenotype on its ability to survive in hypoxia, which was evidenced by an impaired tolerance to hypoxia and an increased oxygen deficit in hypoxia. Therefore, RAP identified a chronic impairment from acute exposure to oil to the hypoxia-tolerant segment of the sea bass population, perhaps increasing their risk of perishing in hypoxic episodes. These successful applications illustrate that RAP is a reliable methodology that could find further application to comparative studies of thermal, hypoxic and anthropogenic effects on the ecological performances of fishes.

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Maximum Heart Rate as a Means of Rapidly Estimating Optimal Temperature for Aeorbic Scope in Salmon: Its Potential for Application (2012)

Knowing the optimal temperature (Topt) for aerobic scope of fishes may be useful for predicting responses to warming environmental temperature because Topt is when fish can allocate the most oxygen to activity. However, the broad application of Topt measurements is hampered by the time required to determine Topt of a species. This thesis sought to develop a rapid method of estimating Topt of Pacific salmon species (genus Oncorhynchus) based on evidence that suggests the decline in aerobic scope above Topt is triggered by a limitation on maximum heart rate (fH). Thus, maximum fH at elevated temperature is thought to limit oxygen convection through the circulatory system, and hence limits both maximum metabolic rate and aerobic scope. Measurements of metabolic rate and fH were taken over a range of temperatures at rest and following exhaustive exercise in juvenile coho salmon (O. kisutch) to confirm the association between Topt and maximum fH. In vivo measurements revealed a maximum fH limitation at the Topt for aerobic scope, supporting the use of fH to estimate Topt. The potential for expediting measurements of maximum fH during acute warming was investigated using anaesthetized coho salmon and pharmacological stimulation of fH. In coho salmon sedated with low doses of anaesthetic, pharmacologically stimulated fH was equivalent to the maximum fH measured in vivo. Breakpoint analysis of the relationship between maximum fH and temperature demonstrated a limitation on maximum fH that corresponded closely with the Topt for aerobic scope. Further, while Topt measurements took three weeks to complete, maximum fH measurements were completed in three days. Therefore, the novel maximum fH technique considerably reduced the time needed to estimate Topt and may be broadly suited to estimating Topt both within and outside of the Oncorhynchus genus. Potential application of this rapid Topt estimation method is discussed in relation to temperature data collected from two local coho salmon-bearing streams. Temperature data also allowed for the examination of stream warming and cooling dynamics and identification of habitat critical to buffering anthropogenic disturbances to stream temperature. These data highlight the importance of riparian areas for maintaining the thermal integrity of waterways.

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Anoxic Survival and cardiovascular responses of the pacific hagfish (2010)

To determine if anoxic survival in the Pacific hagfish (Eptatretus stoutii L) is aided by the suppression of metabolic rate, excess post-anoxic oxygen consumption (EPAOC) and key metabolites in the glycolytic pathway were analyzed following anoxic exposures of different durations. As the cardiovascular system reflects the needs of the tissue and thus whole animal metabolic rate, cardiac performance during prolonged anoxia was also examined to gain insight into the anoxic cardiac ATP turnover rate. Hagfish survived 36-h exposure of complete anoxia at 10°C but showed 50% mortality if exposed to anoxia for 48 h. In order to determine if there had been metabolic rate suppression, changes in tissue metabolites were measured during 36 h anoxia exposure and EPAOC was monitored using respirometry. Analysis of EPAOC measurements suggest that hagfish use metabolic rate suppression to enhance anoxia survival during bouts of anoxia greater than 24 h and that metabolic rate was halved during the final third of a 36-h anoxic period. However, analysis of tissue metabolites in the liver, heart, tongue and skeletal muscle showed that glycogen levels were rapidly depleted over the first 6 h, but then stabilized for the duration of the anoxic exposure. Taken together, the results of the respirometry study and metabolic analysis suggest that anoxia survival in E. stoutii is enhanced by metabolic suppression, but that this suppression may occur earlier in the anoxic period than revealed by EPAOC measurements alone.To gain a better understanding of the use of metabolic rate suppression as a means for surviving anoxic exposures, cardiovascular function was examined during a 36-h anoxic exposure. Cardiac output and ventral aortic blood pressure were measured for 36 h of anoxia and through full recovery. Anoxic bradycardia that halved heart rate within 3 h, which then remained stable at 5 bpm for 33 h of anoxia. Cardiac output, however, was reduced by only ~33%, suggesting metabolite, hormone and waste transport remain important during anoxia. Furthermore, cardiac power output remained unchanged during anoxia. Thus, cardiac metabolic rate is not suppressed and its routine cardiac ATP demand is met through glycolysis and circulating blood glucose.

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