Mechanisms of cardiac collapse at high temperature in a marine teleost (Girella nigrians).

Heat-induced mortality in ectotherms may be attributed to impaired cardiac performance, specifically a collapse in maximum heart rate (fHmax), although the physiological mechanisms driving this phenomenon are still unknown. Here, we tested two proposed factors which may restrict cardiac upper thermal limits: noxious venous blood conditions and oxygen limitation. We hypothesized elevated blood [K+] (hyperkalemia) and low oxygen (hypoxia) would reduce cardiac upper thermal limits in a marine teleost (Girella nigricans), while high oxygen (hyperoxia) would increase thermal limits. We also hypothesized higher acclimation temperatures would exacerbate the harmful effects of an oxygen limitation. Using the Arrhenius breakpoint temperature test, we measured fHmax in acutely warmed fish under control (saline injected) and hyperkalemic conditions (elevated plasma [K+]) while exposed to hyperoxia (200% air saturation), normoxia (100% air saturation), or hypoxia (20% air saturation). We also measured ventricle lactate content and venous blood oxygen partial pressure (PO2) to determine if there were universal thresholds in either metric driving cardiac collapse. Elevated [K+] was not significantly correlated with any cardiac thermal tolerance metric. Hypoxia significantly reduced cardiac upper thermal limits (Arrhenius breakpoint temperature [TAB], peak fHmax, temperature of peak heart rate [TPeak], and temperature at arrhythmia [TARR]). Hyperoxia did not alter cardiac thermal limits compared to normoxia. There was no evidence of a species-wide threshold in ventricular [lactate] or venous PO2. Here, we demonstrate that oxygen limits cardiac thermal tolerance only in instances of hypoxia, but that other physiological processes are responsible for causing temperature-induced heart failure when oxygen is not limited.

Temperature sensitivity differs between heart and red muscle mitochondria in mahi-mahi (Coryphaena hippurus).

Maintaining energy balance over a wide range of temperatures is critical for an active pelagic fish species such as the mahi-mahi (Coryphaena hippurus), which can experience rapid changes in temperature during vertical migrations. Due to the profound effect of temperature on mitochondrial function, this study was designed to investigate the effects of temperature on mitochondrial respiration in permeabilized heart and red skeletal muscle (RM) fibres isolated from mahi-mahi. As RM is thought to be more anatomically isolated from rapid ambient temperature changes compared to the myocardium, it was hypothesized that heart mitochondria would be more tolerant of temperature changes through a greater ability to match respiratory capacity to an increase in temperature and to maintain coupling, when compared to RM mitochondria. Results show that heart fibres were more temperature sensitive and increased respiration rate with temperature increases to a greater degree than RM. Respiratory coupling ratios at the three assay temperatures (20, 26, and 30 °C), revealed that heart mitochondria were less coupled at a lower temperature (26 °C) compared to RM mitochondria (30 °C). In response to an in vitro acute temperature challenge, both tissues showed irreversible effects, where both heart and RM increased uncoupling whether the assay temperature was acutely changed from 20 to 30 °C or 30 to 20 °C. The findings from this study indicate that mahi-mahi heart mitochondria were more temperature sensitive compared to those from RM.

Surviving anoxia: the maintenance of energy production and tissue integrity during anoxia and reoxygenation.

The development of anoxia within tissues represents a significant challenge to most animals because of the decreased capacity for aerobic ATP production, the associated loss of essential cellular functions and the potential for detrimental tissue oxidation upon reoxygenation. Despite these challenges, there are many animals from multiple phyla that routinely experience anoxia and can fully recover. In this Review, we integrate knowledge gained from studies of anoxia-tolerant species across many animal taxa. We primarily focus on strategies used to reduce energy requirements, minimize the consequences of anaerobic ATP production and reduce the adverse effects of reactive oxygen species, which are responsible for tissue damage with reoxygenation. We aim to identify common strategies, as well as novel solutions, to the challenges of anoxia exposure. This Review chronologically examines the challenges faced by animals as they enter anoxia, as they attempt to maintain physiological function during prolonged anoxic exposure and, finally, as they emerge from anoxia. The capacity of animals to survive anoxia is also considered in relation to the increasing prevalence of anoxic zones within marine and freshwater environments, and the need to understand what limits survival.

Impacts of Deepwater Horizon Crude Oil on Mahi-Mahi (Coryphaena hippurus) Heart Cell Function.

Deepwater Horizon crude oil is comprised of polycyclic aromatic hydrocarbons that cause a number of cardiotoxic effects in marine fishes across all levels of biological organization and at different life stages. Although cardiotoxic impacts have been widely reported, the mechanisms underlying these impairments in adult fish remain understudied. In this study, we examined the impacts of crude oil on cardiomyocyte contractility and electrophysiological parameters in freshly isolated ventricular cardiomyocytes from adult mahi-mahi (Coryphaena hippurus). Cardiomyocytes directly exposed to oil exhibited reduced contractility over a range of environmentally relevant concentrations (2.8-12.9 µg l-1∑PAH). This reduction in contractility was most pronounced at higher stimulation frequencies, corresponding to the upper limits of previously measured in situ mahi heart rates. To better understand the mechanisms underlying impaired contractile function, electrophysiological studies were performed, which revealed oil exposure prolonged cardiomyocyte action potentials and disrupted potassium cycling (9.9-30.4 µg l-1∑PAH). This study is the first to measure cellular contractility in oil-exposed cardiomyocytes from a pelagic fish. Results from this study contribute to previously observed impairments to heart function and whole-animal exercise performance in mahi, underscoring the advantages of using an integrative approach in examining mechanisms of oil-induced cardiotoxicity in marine fish.

Acute crude oil exposure alters mitochondrial function and ADP affinity in cardiac muscle fibers of young adult Mahi-mahi (Coryphaena hippurus).

Mitochondrial function is critical to support aerobic metabolism through the production of ATP, and deficiencies in mitochondrial bioenergetics will directly impact the performance capacity of highly aerobic tissues such as the myocardium. Cardiac function in fish has been shown to be negatively affected by crude oil exposure, however, the mechanism for this adverse response is largely unexplored. We hypothesized that lipophilic polycyclic aromatic hydrocarbons (PAHs) found in crude oil disrupt the electron transport system (ETS) ultimately leading to mitochondrial dysfunction. In this study, mitochondrial respiration and ADP affinity we measured using high resolution respirometery in permeabilized cardiac muscle fibers of young adult Mahi-mahi (Coryphaena hippurus) after an acute (24 h) whole animal crude oil exposure. Oil exposure reduced both complex I-fueled ADP stimulated respiration (OXPHOSCI) and complex I,II-fueled ADP stimulated respiration (OXPHOSCI, CII) by 33%,while complex II-fueled ADP stimulated respiration (OXPHOSCII) was reduced by 25%. These changes were found without changes in enzyme activity or mitochondrial density between control and oil exposed Mahi. Additionally, mitochondrial affinity for ADP was decreased three-fold after acute exposure to crude oil. We purpose that acute crude oil exposure selectively impairs mitochondrial complexes of the electron transport system and ATP supply to the cell. This limited ATP supply could present several challenges to a predatory animal like the mahi; including a reliance on anaerobic metabolism and ultimately cell or tissue death as metabolic substrates are rapidly depleted. However, the impact of this impairment may only be evident under periods of increased aerobic metabolic demand.

Oil Exposure Impairs In Situ Cardiac Function in Response to ß-Adrenergic Stimulation in Cobia (Rachycentron canadum).

Aqueous crude oil spills expose fish to varying concentrations of dissolved polycyclic aromatic hydrocarbons (PAHs), which can have lethal and sublethal effects. The heart is particularly vulnerable in early life stages, as PAH toxicity causes developmental cardiac abnormalities and impaired cardiovascular function. However, cardiac responses of juvenile and adult fish to acute oil exposure remain poorly understood. We sought to assess cardiac function in a pelagic fish species, the cobia (Rachycentron canadum), following acute (24 h) exposure to two ecologically relevant levels of dissolved PAHs. Cardiac power output (CPO) was used to quantify cardiovascular performance using an in situ heart preparation. Cardiovascular performance was varied using multiple concentrations of the ß-adrenoceptor agonist isoproterenol (ISO) and by varying afterload pressures. Oil exposure adversely affected CPO with control fish achieving maximum CPO's (4 mW g-1 Mv) greater than that of oil-exposed fish (1 mW g-1 Mv) at ISO concentrations of 1 × 10-6 M. However, the highest concentration of ISO (1 × 10-5 M) rescued cardiac function. This indicates an interactive effect between oil-exposure and ß-adrenergic stimulation and suggests if animals achieve very large increases in ß-adrenergic stimulation it could play a compensatory role that may mitigate some adverse effects of oil-exposure in vivo.

Cardio-respiratory function during exercise in the cobia, Rachycentron canadum: The impact of crude oil exposure.

Aerobic exercise capacity is dependent on the cardiorespiratory system's ability to supply oxygen at a rate that meets energetic demands. In teleost fish crude oil exposure, with the associated polycyclic aromatic hydrocarbons (PAH's), reduces exercise performance and this has been hypothesized to be due to compromised cardiovascular function. In this study, we test this hypothesis by simultaneously measuring cardiovascular performance, oxygen consumption, and swim performance in a pelagic teleost, the cobia (Rachycentron canadum). Metabolic rate increased over 300% in both groups during the swim trial but as the fish approached the critical swim speed (Ucrit) MO2 was 12% lower in the oil exposed fish. Further, stroke volume was initially 35% lower while heart rate was 15% higher in the oil exposed compared to control fish. Our findings suggested, while aspects of cardiovascular and metabolic function are altered by oil exposure, additional studies are needed to further understand the homeostatic mechanisms that may sustain cardiovascular function at higher exercise intensities in cobia.

Determinants of coronary blood flow in sandbar sharks, Carcharhinus plumbeus.

The coronary circulation first appeared in the chordate lineage in cartilaginous fishes where, as in birds and mammals but unlike most teleost fishes, it supplies arterial blood to the entire myocardium. Despite the pivotal position of elasmobranch fishes in the evolution of the coronary circulation, the determinants of coronary blood flow have never been investigated in this group. Elasmobranch fishes are of special interest because of the morphological arrangement of their cardiomyocytes. Unlike teleosts, the majority of the ventricular myocardium in elasmobranch fishes is distant to the venous blood returning to the heart (i.e., the luminal blood). Also, the majority of the myocardium is in close association with the coronary circulation. To determine the relative contribution of the coronary and luminal blood supplies to cardiovascular function in sandbar sharks, Carcharhinus plumbeus, we measured coronary blood flow while manipulating cardiovascular status using acetylcholine and adrenaline. By exploring inter- and intra-individual variation in cardiovascular variables, we show that coronary blood flow is directly related to heart rate (R 2 = 0.6; P < 0.001), as it is in mammalian hearts. Since coronary blood flow is inversely related to coronary resistance both in vivo and in vitro, we suggest that in elasmobranch fishes, changes in heart rate mediate changes in coronary vascular resistance, which adjust coronary blood flow appropriately.

Effects of crude oil on in situ cardiac function in young adult mahi-mahi (Coryphaena hippurus).

Exposure to polycyclic aromatic hydrocarbons (PAH) negatively impacts exercise performance in fish species but the physiological modifications that result in this phenotype are poorly understood. Prior studies have shown that embryonic and juvenile mahi-mahi (Coryphaeus hippurus) exposed to PAH exhibit morphological abnormalities, altered cardiac development and reduced swimming performance. It has been suggested that cardiovascular function inhibited by PAH exposure accounts for the compromised exercise performance in fish species. In this study we used in-situ techniques to measure hemodynamic responses of young adult mahi-mahi exposed to PAH for 24h. The data indicate that stroke volume was reduced 44% in mahi-mahi exposed to 9.6±2.7µgl-1 geometric mean PAH (∑PAH) and resulted in a 39% reduction in cardiac output and a 52% reduction in stroke work. Maximal change in pressure over change in time was 28% lower in mahi-mahi exposed to this level of ∑PAH. Mean intraventricular pressures and heart rate were not significantly changed. This study suggests exposure to environmentally relevant PAH concentrations impairs aspects of cardiovascular function in mahi-mahi.

Introducing a novel mechanism to control heart rate in the ancestral Pacific hagfish.

Although neural modulation of heart rate is well established among chordate animals, the Pacific hagfish (Eptatretus stoutii) lacks any cardiac innervation, yet it can increase its heart rate from the steady, depressed heart rate seen in prolonged anoxia to almost double its normal normoxic heart rate, an almost fourfold overall change during the 1-h recovery from anoxia. The present study sought mechanistic explanations for these regulatory changes in heart rate. We provide evidence for a bicarbonate-activated, soluble adenylyl cyclase (sAC)-dependent mechanism to control heart rate, a mechanism never previously implicated in chordate cardiac control.

Morphological arrangement of the coronary vasculature in a shark (Squalus sucklei) and a teleost (Oncorhynchus mykiss).

The coronary circulation is of great importance in maintaining cardiovascular function and consequently it has been extensively studied in many mammalian species. However, much less attention has been paid to the coronary circulation in other vertebrates. For example, while elasmobranch fishes are of special interest as they are the most ancient lineage of vertebrates to possess a coronary circulation, only qualitative studies exist on their coronary circulation and most concern the architecture of the large arteries. Our study tested the prediction that the coronary circulation of sharks is better developed than previously thought. However, to test this idea, a methodology was needed to quantify vascularity, vessel morphology and oxygen diffusion distances in a heart with predominantly spongy myocardium. Here, we describe this methodology using dogfish and rainbow trout and suggest that the dogfish spongy myocardium appears to rely predominantly on the coronary circulation for its oxygen supply, an arrangement that contrasts with the spongy myocardial tissue of rainbow trout. In support of this suggestion, the density of the microvasculature of the spongy myocardial tissue of dogfish exceeded that of their compact tissue. Although vascularity in the compact myocardium of dogfish was significantly lower than trout, intervascular distances were similar on account of a significantly larger vessel diameter in dogfish, which corresponds to a larger red blood cell size of the dogfish when compared to trout. J. Morphol. 277:896-905, 2016. © 2016 Wiley Periodicals, Inc.

Characterizing the metabolic capacity of the anoxic hagfish heart.

Pacific hagfish, Eptatretus stoutii, can recover from 36 h of anoxia at 10°C. Such anoxia tolerance demands the mobilization of anaerobic fuels and the removal of metabolic wastes--processes that require a functional heart. The purpose of this study was to measure the metabolic response of the excised, cannulated hagfish heart to anoxia using direct calorimetry. These experiments were coupled with measurements of cardiac pH and metabolite concentrations, at multiple time points, to monitor acid-base balance and anaerobic ATP production. We also exposed hagfish to anoxia to compare the in vitro responses of the excised hearts with the in vivo responses. The calorimetry results revealed a significant reduction in the rate of metabolic heat production over the first hour of anoxia exposure, and a recovery over the subsequent 6 h. This response is likely attributable to a rapid anoxia-induced depression of aerobic ATP-production pathways followed by an upregulation of anaerobic ATP-production pathways such that the ATP production rate was restored to that measured in normoxia. Glycogen-depletion measurements suggest that metabolic processes were initially supported by glycolysis but that an alternative fuel source was used to support the sustained rates of ATP production. The maintenance of intracellular pH during anoxia indicates a remarkable ability of the myocytes to buffer/regulate protons and thus protect cardiac function. Altogether, these results illustrate that the low metabolic demand of the hagfish heart allows for near-routine levels of cardiac metabolism to be supported anaerobically. This is probably a significant contributor to the hagfish's exceptional anoxia tolerance.

The beat goes on: Cardiac pacemaking in extreme conditions.

In order for an animal to survive, the heart beat must go on in all environmental conditions, or at least restart its beat. This review is about maintaining a rhythmic heartbeat under the extreme conditions of anoxia (or very severe hypoxia) and high temperatures. It starts by considering the primitive versions of the protein channels that are responsible for initiating the heartbeat, HCN channels, divulging recent findings from the ancestral craniate, the Pacific hagfish (Eptatretus stoutii). It then explores how a heartbeat can maintain a rhythm, albeit slower, for hours without any oxygen, and sometimes without autonomic innervation. It closes with a discussion of recent work on fishes, where the cardiac rhythm can become arrhythmic when a fish experiences extreme heat.

Anoxic survival of the Pacific hagfish (Eptatretus stoutii).

It is not known how the Pacific hagfish (Eptatretus stoutii) can survive extended periods of anoxia. The present study used two experimental approaches to examine energy use during and following anoxic exposure periods of different durations (6, 24 and 36 h). By measuring oxygen consumption prior to anoxic exposure, we detected a circadian rhythm, with hagfish being active during night and showing a minimum routine oxygen consumption (RMR) during the daytime. By measuring the excess post-anoxic oxygen consumption (EPAOC) after 6 and 24 h it was possible to mathematically account for RMR being maintained even though heme stores of oxygen would have been depleted by the animal's metabolism during the first hours of anoxia. However, EPAOC after 36 h of anoxia could not account for RMR being maintained. Measurements of tissue glycogen disappearance and lactate appearance during anoxia showed that the degree of glycolysis and the timing of its activation varied among tissues. Yet, neither measurement could account for the RMR being maintained during even the 6-h anoxic period. Therefore, two independent analyses of the metabolic responses of hagfish to anoxia exposure suggest that hagfish utilize metabolic rate suppression as part of the strategy for longer-term anoxia survival.

Cardiac responses to anoxia in the Pacific hagfish, Eptatretus stoutii.

In the absence of any previous study of the cardiac status of hagfishes during prolonged anoxia and because of their propensity for oxygen-depleted environments, the present study tested the hypothesis that the Pacific hagfish Eptatretus stoutii maintains cardiac performance during prolonged anoxia. Heart rate was halved from the routine value of 10.4±1.3 beats min⁻¹ by the sixth hour of an anoxic period and then remained stable for a further 30 h. Cardiac stroke volume increased from routine (1.3±0.1 ml kg⁻¹) to partially compensate the anoxic bradycardia, such that cardiac output decreased by only 33% from the routine value of 12.3±0.9 ml min⁻¹ kg⁻¹. Cardiac power output decreased by only 25% from the routine value of 0.26±0.02 mW g⁻¹. During recovery from prolonged anoxia, cardiac output and heart rate increased to peak values within 1.5 h. Thus, the Pacific hagfish should be acknowledged as hypoxic tolerant in terms of its ability to maintain around 70% of their normoxic cardiac performance during prolonged anoxia. This is only the second fish species to be so classified.

The role of venous capacitance, circulating catecholamines, and heart rate in the hemodynamic response to increased temperature and hypoxia in the dogfish.

Hypoxia and increased temperature alter venous blood pressures in teleosts through active changes in venous tone. Elasmobranchs possess a capacious venous system but have limited adrenergic vascular innervation and subambient central venous pressure (P(cv)). In this study, we explored venous hemodynamic responses to acute temperature increase and moderate (6.9 kPa) and severe (2.5 kPa) hypoxia in the dogfish (Squalus acanthias). Normoxic dogfish at 10 degrees C had a P(cv) between -0.08 and -0.04 kPa and a mean circulatory filling pressure (P(mcf)) of approximately 0.12 kPa. At 16 degrees C, heart rate (f(H)), cardiac output (Q), and P(mcf) increased but P(cv) and plasma epinephrine and norepinephrine levels were unchanged. In contrast, moderate and severe hypoxia increased P(cv) and decreased Q and stroke volume (V(S)). f(H) decreased in severe hypoxia, whereas P(mcf) was unaffected despite elevated catecholamine levels. Atropine abolished hypoxic reductions in Q, V(S), and f(H), but P(cv) still increased. In contrast to the response in teleosts, this study on dogfish suggests that venous capacitance changes associated with warming and hypoxia are minimal and likely not mediated by circulating catecholamines. Thus hemodynamic status of the capacious elasmobranch venous circulation is potentially regulated by blood volume shifts from passive flow-mediated events and possibly through myogenic mechanisms.

Circulatory limits to oxygen supply during an acute temperature increase in the Chinook salmon (Oncorhynchus tshawytscha).

This study was undertaken to provide a comprehensive set of data relevant to disclosing the physiological effects and possible oxygen transport limitations in the Chinook salmon (Oncorhynchus tshawytscha) during an acute temperature change. Fish were instrumented with a blood flow probe around the ventral aorta and catheters in the dorsal aorta and sinus venosus. Water temperature was progressively increased from 13 degrees C in steps of 4 degrees C up to 25 degrees C. Cardiac output increased from 29 to 56 ml.min(-1).kg(-1) between 13 and 25 degrees C through an increase in heart rate (58 to 105 beats/min). Systemic vascular resistance was reduced, causing a stable dorsal aortic blood pressure, yet central venous blood pressure increased significantly at 25 degrees C. Oxygen consumption rate increased from 3.4 to 8.7 mg.min(-1).kg(-1) during the temperature increase, although there were signs of anaerobic respiration at 25 degrees C in the form of increased blood lactate and decreased pH. Arterial oxygen partial pressure was maintained during the heat stress, although venous oxygen partial pressure (Pv(O(2))) and venous oxygen content were significantly reduced. Cardiac arrhythmias were prominent in three of the largest fish (>4 kg) at 25 degrees C. Given the switch to anaerobic metabolism and the observation of cardiac arrhythmias at 25 degrees C, we propose that the cascade of venous oxygen depletion results in a threshold value for Pv(O(2)) of around 1 kPa. At this point, the oxygen supply to systemic and cardiac tissues is compromised, such that the oxygen-deprived and acidotic myocardium becomes arrhythmic, and blood perfusion through the gills and to the tissues becomes compromised.