Cardiac GR Mediates the Diurnal Rhythm in Ventricular Arrhythmia Susceptibility.

BACKGROUND: Ventricular arrhythmias (VAs) demonstrate a prominent day-night rhythm, commonly presenting in the morning. Transcriptional rhythms in cardiac ion channels accompany this phenomenon, but their role in the morning vulnerability to VAs and the underlying mechanisms are not understood. We investigated the recruitment of transcription factors that underpins transcriptional rhythms in ion channels and assessed whether this mechanism was pertinent to the heart's intrinsic diurnal susceptibility to VA. METHODS AND RESULTS: Assay for transposase-accessible chromatin with sequencing performed in mouse ventricular myocyte nuclei at the beginning of the animals' inactive (ZT0) and active (ZT12) periods revealed differentially accessible chromatin sites annotating to rhythmically transcribed ion channels and distinct transcription factor binding motifs in these regions. Notably, motif enrichment for the glucocorticoid receptor (GR; transcriptional effector of corticosteroid signaling) in open chromatin profiles at ZT12 was observed, in line with the well-recognized ZT12 peak in circulating corticosteroids. Molecular, electrophysiological, and in silico biophysically-detailed modeling approaches demonstrated GR-mediated transcriptional control of ion channels (including Scn5a underlying the cardiac Na+ current, Kcnh2 underlying the rapid delayed rectifier K+ current, and Gja1 responsible for electrical coupling) and their contribution to the day-night rhythm in the vulnerability to VA. Strikingly, both pharmacological block of GR and cardiomyocyte-specific genetic knockout of GR blunted or abolished ion channel expression rhythms and abolished the ZT12 susceptibility to pacing-induced VA in isolated hearts. CONCLUSIONS: Our study registers a day-night rhythm in chromatin accessibility that accompanies diurnal cycles in ventricular myocytes. Our approaches directly implicate the cardiac GR in the myocyte excitability rhythm and mechanistically link the ZT12 surge in glucocorticoids to intrinsic VA propensity at this time.

The integrative biology of the heart: mechanisms enabling cardiac plasticity.

Cardiac phenotypic plasticity, the remodelling of heart structure and function, is a response to any sustained (or repeated) stimulus or stressor that results in a change in heart performance. Cardiac plasticity can be either adaptive (beneficial) or maladaptive (pathological), depending on the nature and intensity of the stimulus. Here, we draw on articles published in this Special Issue of Journal of Experimental Biology, and from the broader comparative physiology literature, to highlight the core components that enable cardiac plasticity, including structural remodelling, excitation-contraction coupling remodelling and metabolic rewiring. We discuss when and how these changes occur, with a focus on the underlying molecular mechanisms, from the regulation of gene transcription by epigenetic processes to post-translational modifications of cardiac proteins. Looking to the future, we anticipate that the growing use of -omics technologies in integration with traditional comparative physiology approaches will allow researchers to continue to uncover the vast scope for plasticity in cardiac function across animals.

Developmental programming of sarcoplasmic-reticulum function improves cardiac anoxia tolerance in turtles.

Oxygen deprivation during embryonic development can permanently remodel the vertebrate heart, often causing cardiovascular abnormalities in adulthood. While this phenomenon is mostly damaging, recent evidence suggests developmental hypoxia produces stress-tolerant phenotypes in some ectothermic vertebrates. Embryonic common snapping turtles (Chelydra serpentina) subjected to chronic hypoxia display improved cardiac anoxia tolerance after hatching, which is associated with altered Ca2+ homeostasis in heart cells (cardiomyocytes). Here we examined the possibility that changes in Ca2+ cycling, through the sarcoplasmic reticulum (SR), underlie the developmentally programmed cardiac phenotype of snapping turtles. We investigated this hypothesis by isolating cardiomyocytes from juvenile turtles that developed in either normoxia (21% O2; "N21") or chronic hypoxia (10% O2; "H10") and subjected the cells to anoxia/reoxygenation, either in the presence or absence of SR Ca2+-cycling inhibitors. We simultaneously measured cellular shortening, intracellular [Ca2+], and intracellular pH (pHi). Under normoxic conditions, N21 and H10 cardiomyocytes shortened equally, but H10 Ca2+ transients (Δ[Ca2+]i) were twofold smaller than N21 cells, and SR inhibition only decreased N21 shortening and Δ[Ca2+]i. Anoxia subsequently depressed shortening, Δ[Ca2+]i, and pHi in control N21 and H10 cardiomyocytes, yet H10 shortening and Δ[Ca2+]i recovered to pre-anoxic levels, partly due to enhanced myofilament Ca2+ sensitivity. SR blockade abolished the recovery of anoxic H10 cardiomyocytes and potentiated decreases in shortening, Δ[Ca2+]i, and pHi. Our novel results provide the first evidence of developmental programming of SR function and demonstrate that developmental hypoxia confers a long-lasting, superior anoxia-tolerant cardiac phenotype in snapping turtles, by enhancing myofilament Ca2+ sensitivity and modifying SR function.

Developmental plasticity of the cardiovascular system in oviparous vertebrates: effects of chronic hypoxia and interactive stressors in the context of climate change.

Animals at early life stages are generally more sensitive to environmental stress than adults. This is especially true of oviparous vertebrates that develop in variable environments with little or no parental care. These organisms regularly experience environmental fluctuations as part of their natural development, but climate change is increasing the frequency and intensity of these events. The developmental plasticity of oviparous vertebrates will therefore play a critical role in determining their future fitness and survival. In this Review, we discuss and compare the phenotypic consequences of chronic developmental hypoxia on the cardiovascular system of oviparous vertebrates. In particular, we focus on species-specific responses, critical windows, thresholds for responses and the interactive effects of other stressors, such as temperature and hypercapnia. Although important progress has been made, our Review identifies knowledge gaps that need to be addressed if we are to fully understand the impact of climate change on the developmental plasticity of the oviparous vertebrate cardiovascular system.

Evolution and divergence of teleost adrenergic receptors: why sometimes 'the drugs don't work' in fish.

Adrenaline and noradrenaline, released as hormones and/or neurotransmitters, exert diverse physiological functions in vertebrates, and teleost fishes are widely used as model organisms to study adrenergic regulation; however, such investigations often rely on receptor subtype-specific pharmacological agents (agonists and antagonists; see Glossary) developed and validated in mammals. Meanwhile, evolutionary (phylogenetic and comparative genomic) studies have begun to unravel the diversification of adrenergic receptors (ARs) and reveal that whole-genome duplications and pseudogenization events in fishes results in notable distinctions from mammals in their genomic repertoire of ARs, while lineage-specific gene losses within teleosts have generated significant interspecific variability. In this Review, we visit the evolutionary history of ARs (including α1-, α2- and ß-ARs) to highlight the prominent interspecific differences in teleosts, as well as between teleosts and other vertebrates. We also show that structural modelling of teleost ARs predicts differences in ligand binding affinity compared with mammalian orthologs. To emphasize the difficulty of studying the roles of different AR subtypes in fish, we collate examples from the literature of fish ARs behaving atypically compared with standard mammalian pharmacology. Thereafter, we focus on specific case studies of the liver, heart and red blood cells, where our understanding of AR expression has benefited from combining pharmacological approaches with molecular genetics. Finally, we briefly discuss the ongoing advances in 'omics' technologies that, alongside classical pharmacology, will provide abundant opportunities to further explore adrenergic signalling in teleosts.

The air-breathing Alaska blackfish (Dallia pectoralis) suppresses brain mitochondrial reactive oxygen species to survive cold hypoxic winters.

The Alaska blackfish (Dallia pectoralis) is the only air-breathing fish in the Arctic. In the summer, a modified esophagus allows the fish to extract oxygen from the air, but this behavior is not possible in the winter because of ice and snow cover. The lack of oxygen (hypoxia) and near freezing temperatures in winter is expected to severely compromise metabolism, and yet remarkably, overwintering Alaska blackfish remain active. To maintain energy balance in the brain and limit the accumulation of reactive oxygen species (ROS), we hypothesized that cold hypoxic conditions would trigger brain mitochondrial remodeling in the Alaska blackfish. To address this hypothesis, fish were acclimated to warm (15 °C) normoxia, cold (5 °C) normoxia or cold hypoxia (5 °C, 2.1-4.2 kPa; no air access) for 5-8 weeks. Mitochondrial respiration, ADP affinity and H202 production were measured at 10 °C in isolated brain homogenates with an Oroboros respirometer. Cold acclimation and chronic hypoxia had no effects on mitochondrial aerobic capacity or ADP affinity. However, cold acclimation in normoxia led to a suppression of brain mitochondrial H202 production, which persisted and became more pronounced in the cold hypoxic fish. Overall, our study suggests cold acclimation supresses ROS production in Alaska blackfish, which may protect the fish from oxidative stress when oxygen becomes limited during winter.

Thermal preference does not align with optimal temperature for aerobic scope in zebrafish (Danio rerio).

Warming is predicted to have negative consequences for fishes by causing a mismatch between oxygen demand and supply, and a consequent reduction in aerobic scope (AS) and performance. This oxygen and capacity limited thermal tolerance (OCLTT) hypothesis features prominently in the literature but remains controversial. Within the OCLTT framework, we hypothesised that fish would select temperatures that maximise their AS, and thus their performance. We tested this hypothesis using intermittent flow respirometry to measure AS at, above (+2.5°C) and below (-2.5°C) the self-selected, preferred temperature (Tpref) of individual zebrafish (Danio rerio). AS was greatest 2.5°C above Tpref, which was driven by an increase in maximal metabolic rate. This mismatch between Tpref and the optimal temperature for AS suggests that factor(s) aside from AS maximisation influence the thermal preference of zebrafish.

Absence of atrial smooth muscle in the heart of the loggerhead sea turtle (Caretta caretta): a re-evaluation of its role in diving physiology.

Contraction of atrial smooth muscle in the hearts of semi-aquatic emydid turtles regulates ventricular filling, and it has been proposed that it could regulate stroke volume during characteristic rapid transitions in cardiac output associated with diving. For this hypothesis to be supported, atrial smooth muscle should be widely distributed in diving Testudines. To further understand the putative function and evolutionary significance of endocardial smooth muscle in Testudines, we studied the hearts of loggerhead sea turtles, Caretta caretta (n=7), using immunohistochemistry and histology. Surprisingly, we found no evidence of prominent atrial smooth muscle in C. caretta. However, smooth muscle was readily identified in the sinus venosus. Our results suggest that atrial smooth muscle does not contribute to the diving capabilities of C. caretta, indicating that the possible roles of smooth muscle in emydid turtle hearts require a re-evaluation. In sea turtles, the sinus venosus may instead contribute to regulate cardiac filling.

Inhibition of the hERG potassium channel by phenanthrene: a polycyclic aromatic hydrocarbon pollutant.

The lipophilic polycyclic aromatic hydrocarbon (PAH) phenanthrene is relatively abundant in polluted air and water and can access and accumulate in human tissue. Phenanthrene has been reported to interact with cardiac ion channels in several fish species. This study was undertaken to investigate the ability of phenanthrene to interact with hERG (human Ether-à-go-go-Related Gene) encoded Kv11.1 K+ channels, which play a central role in human ventricular repolarization. Pharmacological inhibition of hERG can be proarrhythmic. Whole-cell patch clamp recordings of hERG current (IhERG) were made from HEK293 cells expressing wild-type (WT) and mutant hERG channels. WT IhERG1a was inhibited by phenanthrene with an IC50 of 17.6 ± 1.7 µM, whilst IhERG1a/1b exhibited an IC50 of 1.8 ± 0.3 µM. WT IhERG block showed marked voltage and time dependence, indicative of dependence of inhibition on channel gating. The inhibitory effect of phenanthrene was markedly impaired by the attenuated inactivation N588K mutation. Remarkably, mutations of S6 domain aromatic amino acids (Y652, F656) in the canonical drug binding site did not impair the inhibitory action of phenanthrene; the Y652A mutation augmented IhERG block. In contrast, the F557L (S5) and M651A (S6) mutations impaired the ability of phenanthrene to inhibit IhERG, as did the S624A mutation below the selectivity filter region. Computational docking using a cryo-EM derived hERG structure supported the mutagenesis data. Thus, phenanthrene acts as an inhibitor of the hERG K+ channel by directly interacting with the channel, binding to a distinct site in the channel pore domain.

Adrenergic prolongation of action potential duration in rainbow trout myocardium via inhibition of the delayed rectifier potassium current, IKr.

Catecholamines mediate the 'fight or flight' response in a wide variety of vertebrates. The endogenous catecholamine adrenaline increases heart rate and contractile strength to raise cardiac output. The increase in contractile force is driven in large part by an increase in myocyte Ca2+ influx on the L-type Ca current (ICaL) during the cardiac action potential (AP). Here, we report a K+- based mechanism that prolongs AP duration (APD) in fish hearts following adrenergic stimulation. We show that adrenergic stimulation inhibits the delayed rectifier K+ current (IKr) in rainbow trout (Oncorhynchus mykiss) cardiomyocytes. This slows repolarization and prolongs APD which may contribute to positive inotropy following adrenergic stimulation in fish hearts. The endogenous ligand, adrenaline (1 µM), which activates both α- and ß-ARs reduced maximal IKr tail current to 61.4 ± 3.9% of control in atrial and ventricular myocytes resulting in an APD prolongation of ~20% at both 50 and 90% repolarization. This effect was reproduced by the α-specific adrenergic agonist, phenylephrine (1 µM), but not the ß-specific adrenergic agonist isoproterenol (1 µM). Adrenaline (1 µM) in the presence of ß1 and ß2-blockers (1 µM atenolol and 1 µM ICI-118551, respectively) also inhibited IKr. Thus, IKr suppression following α-adrenergic stimulation leads to APD prolongation in the rainbow trout heart. This is the first time this mechanism has been identified in fish and may act in unison with the well-known enhancement of ICaL following adrenergic stimulation to prolong APD and increase cardiac inotropy.

Warmer, faster, stronger: Ca2+ cycling in avian myocardium.

Birds occupy a unique position in the evolution of cardiac design. Their hearts are capable of cardiac performance on par with, or exceeding that of mammals, and yet the structure of their cardiomyocytes resembles those of reptiles. It has been suggested that birds use intracellular Ca2+ stored within the sarcoplasmic reticulum (SR) to power contractile function, but neither SR Ca2+ content nor the cross-talk between channels underlying Ca2+-induced Ca2+ release (CICR) have been studied in adult birds. Here we used voltage clamp to investigate the Ca2+ storage and refilling capacities of the SR and the degree of trans-sarcolemmal and intracellular Ca2+ channel interplay in freshly isolated atrial and ventricular myocytes from the heart of the Japanese quail (Coturnix japonica). A trans-sarcolemmal Ca2+ current (ICa) was detectable in both quail atrial and ventricular myocytes, and was mediated only by L-type Ca2+ channels. The peak density of ICa was larger in ventricular cells than in atrial cells, and exceeded that reported for mammalian myocardium recorded under similar conditions. Steady-state SR Ca2+ content of quail myocardium was also larger than that reported for mammals, and reached 750.6±128.2 µmol l-1 in atrial cells and 423.3±47.2 µmol l-1 in ventricular cells at 24°C. We observed SR Ca2+-dependent inactivation of ICa in ventricular myocytes, indicating cross-talk between sarcolemmal Ca2+ channels and ryanodine receptors in the SR. However, this phenomenon was not observed in atrial myocytes. Taken together, these findings help to explain the high-efficiency avian myocyte excitation-contraction coupling with regard to their reptilian-like cellular ultrastructure.

Ocean warming and hypoxia affect embryonic growth, fitness and survival of small-spotted catsharks, Scyliorhinus canicula.

Elasmobranchs are key to a healthy marine ecosystem but are under threat from human activities, such as destructive fisheries and shark finning. Embryos of oviparous elasmobranchs may be further challenged during development by rising temperatures and falling dissolved oxygen concentrations in their intertidal environment. However, the impact of climate change on survival and growth of oviparous elasmobranchs is still poorly understood. Here, we investigate the effects of temperature and hypoxia on the growth and survival of small-spotted catshark (Scyliorhinus canicula) embryos by incubating eggs in normoxia 15°C, normoxia 20°C, hypoxia 15°C, or hypoxia 20°C. Incubation under the elevated temperature increased the embryonic growth rate, yolk consumption rate and Fulton's condition factor at hatching, whilst decreasing the total length and body mass of newly hatched sharks. Under low oxygen conditions (50% air saturation) the survival rate of S. canicula embryos dropped significantly and the temperature-induced increase in Fulton's condition factor was reversed. Together, these data demonstrate both the individual and compound effects of elevated temperature and hypoxia on the survival and growth during early ontogeny of a ubiquitous, coastal elasmobranch, S. canicula.

Developmental plasticity of cardiac anoxia-tolerance in juvenile common snapping turtles ( Chelydra serpentina).

For some species of ectothermic vertebrates, early exposure to hypoxia during embryonic development improves hypoxia-tolerance later in life. However, the cellular mechanisms underlying this phenomenon are largely unknown. Given that hypoxic survival is critically dependent on the maintenance of cardiac function, we tested the hypothesis that developmental hypoxia alters cardiomyocyte physiology in a manner that protects the heart from hypoxic stress. To test this hypothesis, we studied the common snapping turtle, which routinely experiences chronic developmental hypoxia and exploits hypoxic environments in adulthood. We isolated cardiomyocytes from juvenile turtles that embryonically developed in either normoxia (21% O2) or hypoxia (10% O2), and subjected them to simulated anoxia and reoxygenation, while simultaneously measuring intracellular Ca2+, pH and reactive oxygen species (ROS) production. Our results suggest developmental hypoxia improves cardiomyocyte anoxia-tolerance of juvenile turtles, which is supported by enhanced myofilament Ca2+-sensitivity and a superior ability to suppress ROS production. Maintenance of low ROS levels during anoxia might limit oxidative damage and a greater sensitivity to Ca2+ could provide a mechanism to maintain contractile force. Our study suggests developmental hypoxia has long-lasting effects on turtle cardiomyocyte function, which might prime their physiology for exploiting hypoxic environments.

Thermal acclimation and seasonal acclimatization: a comparative study of cardiac response to prolonged temperature change in shorthorn sculpin.

Seasonal thermal remodelling (acclimatization) and laboratory thermal remodelling (acclimation) can induce different physiological changes in ectothermic animals. As global temperatures are changing at an increasing rate, there is urgency to understand the compensatory abilities of key organs such as the heart to adjust under natural conditions. Thus, the aim of the present study was to directly compare the acclimatization and acclimatory response within a single eurythermal fish species, the European shorthorn sculpin (Myoxocephalus scorpio). We used current- and voltage-clamp to measure ionic current densities in both isolated atrial and ventricular myocytes from three groups of fish: (1) summer-caught fish kept at 12°C ('summer-acclimated'); (2) summer-caught fish kept at 3°C ('cold acclimated'); and (3) fish caught in March ('winter-acclimatized'). At a common test temperature of 7.5°C, action potential (AP) was shortened by both winter acclimatization and cold acclimation compared with summer acclimation; however, winter acclimatization caused a greater shortening than did cold acclimation. Shortening of AP was achieved mostly by a significant increase in repolarizing current density (IKr and IK1) following winter acclimatization, with cold acclimation having only minor effects. Compared with summer acclimation, the depolarizing L-type calcium current (ICa) was larger following winter acclimatization, but again, there was no effect of cold acclimation on ICa Interestingly, the other depolarizing current, INa, was downregulated at low temperatures. Our further analysis shows that ionic current remodelling is primarily due to changes in ion channel density rather than current kinetics. In summary, acclimatization profoundly modified the electrical activity of the sculpin heart while acclimation to the same temperature for >1.5 months produced very limited remodelling effects.

3D ultrastructural organisation of calcium release units in the avian sarcoplasmic reticulum.

Excitation-contraction coupling in vertebrate hearts is underpinned by calcium (Ca2+) release from Ca2+ release units (CRUs). CRUs are formed by clusters of channels called ryanodine receptors on the sarcoplasmic reticulum (SR) within the cardiomyocyte. Distances between CRUs influence the diffusion of Ca2+, thus influencing the rate and strength of excitation-contraction coupling. Avian myocytes lack T-tubules, so Ca2+ from surface CRUs (peripheral couplings, PCs) must diffuse to internal CRU sites of the corbular SR (cSR) during centripetal propagation. Despite this, avian hearts achieve higher contractile rates and develop greater contractile strength than many mammalian hearts, which have T-tubules to provide simultaneous activation of the Ca2+ signal through the myocyte. We used 3D electron tomography to test the hypothesis that the intracellular distribution of CRUs in the avian heart permits faster and stronger contractions despite the absence of T-tubules. Nearest edge-edge distances between PCs and cSR, and geometric information including surface area and volume of individual cSR, were obtained for each cardiac chamber of the white leghorn chicken. Computational modelling was then used to establish a relationship between CRU distance and cell activation time in the avian heart. Our data suggest that cSR clustered close together along the Z-line is vital for rapid propagation of the Ca2+ signal from the cell periphery to the cell centre, which would aid in the strong and fast contractions of the avian heart.

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.

Recognition software successfully aids the identification of individual small-spotted catsharks Scyliorhinus canicula during their first year of life.

Eighteen captive small-spotted catsharks Scyliorhinus canicula were successfully identified from hatching to 1 year of age using the free computer recognition software, I3 S classic. The effect of increasing the time interval between recognition attempts on the accuracy of the software was investigated, revealing that recognition fiedelity decreases with increasing time intervals for younger (0 to 15 weeks), but not older (15 weeks onwards) sharks. Identification by I3 S was validated using genetic analyses of seven microsatellite markers, revealing a 100% success rate. Thus, this non-invasive recognition method can be used as an inexpensive and effective alternative to invasive tagging, improving animal welfare and complementing ex-situ conservation methods.

Stress-Activated Kinase Mitogen-Activated Kinase Kinase-7 Governs Epigenetics of Cardiac Repolarization for Arrhythmia Prevention.

BACKGROUND: Ventricular arrhythmia is a leading cause of cardiac mortality. Most antiarrhythmics present paradoxical proarrhythmic side effects, culminating in a greater risk of sudden death. METHODS: We describe a new regulatory mechanism linking mitogen-activated kinase kinase-7 deficiency with increased arrhythmia vulnerability in hypertrophied and failing hearts using mouse models harboring mitogen-activated kinase kinase-7 knockout or overexpression. The human relevance of this arrhythmogenic mechanism is evaluated in human-induced pluripotent stem cell-derived cardiomyocytes. Therapeutic potentials by targeting this mechanism are explored in the mouse models and human-induced pluripotent stem cell-derived cardiomyocytes. RESULTS: Mechanistically, hypertrophic stress dampens expression and phosphorylation of mitogen-activated kinase kinase-7. Such mitogen-activated kinase kinase-7 deficiency leaves histone deacetylase-2 unphosphorylated and filamin-A accumulated in the nucleus to form a complex with Krüppel-like factor-4. This complex leads to Krüppel-like factor-4 disassociation from the promoter regions of multiple key potassium channel genes (Kv4.2, KChIP2, Kv1.5, ERG1, and Kir6.2) and reduction of their transcript levels. Consequent repolarization delays result in ventricular arrhythmias. Therapeutically, targeting the repressive function of the Krüppel-like factor-4/histone deacetylase-2/filamin-A complex with the histone deacetylase-2 inhibitor valproic acid restores K+ channel expression and alleviates ventricular arrhythmias in pathologically remodeled hearts. CONCLUSIONS: Our findings unveil this new gene regulatory avenue as a new antiarrhythmic target where repurposing of the antiepileptic drug valproic acid as an antiarrhythmic is supported.

Macro- and micromechanical remodelling in the fish atrium is associated with regulation of collagen 1 alpha 3 chain expression.

Numerous pathologies lead to remodelling of the mammalian ventricle, often associated with fibrosis. Recent work in fish has shown that fibrotic remodelling of the ventricle is 'reversible', changing seasonally as temperature-induced changes in blood viscosity alter haemodynamic load on the heart. The atrial response to varying haemodynamic load is less understood in mammals and completely unexplored in non-mammalian vertebrates. To investigate atrial remodelling, rainbow trout were chronically cooled (from 10 ± 1 to 5 ± 1 °C) and chronically warmed (from 10 ± 1 to 18 ± 1 °C) for a minimum of 8 weeks. We assessed the functional effects on compliance using ex vivo heart preparations and atomic force microscopy nano-indentation and found chronic cold increased passive stiffness of the whole atrium and micromechanical stiffness of tissue sections. We then performed histological, biochemical and molecular assays to probe the mechanisms underlying functional remodelling of the atrial tissue. We found cooling resulted in collagen deposition which was associated with an upregulation of collagen-promoting genes, including the fish-specific collagen I alpha 3 chain, and a reduction in gelatinase activity of collagen-degrading matrix metalloproteinases (MMPs). Finally, we found that cooling reduced mRNA expression of cardiac growth factors and hypertrophic markers. Following long-term warming, there was an opposing response to that seen with cooling; however, these changes were more moderate. Our findings suggest that chronic cooling causes atrial dilation and increased myocardial stiffness in trout atria analogous to pathological states defined by changes in preload or afterload of the mammalian atria. The reversal of this phenotype following chronic warming is particularly interesting as it suggests that typically pathological features of mammalian atrial remodelling may oscillate seasonally in the fish, revealing a more dynamic and plastic atrial remodelling response.