RESUMEN
A regular heartbeat is essential for maintaining the homeostasis of the vertebrate body. However, environmental pollutants, oxygen deficiency and extreme temperatures can impair heart function in fish. In this Review, we provide an integrative view of the molecular origins of cardiac arrhythmias and their functional consequences, from the level of ion channels to cardiac electrical activity in living fish. First, we describe the current knowledge of the cardiac excitation-contraction coupling of fish, as the electrical activity of the heart and intracellular Ca2+ regulation act as a platform for cardiac arrhythmias. Then, we compile findings on cardiac arrhythmias in fish. Although fish can experience several types of cardiac arrhythmia under stressful conditions, the most typical arrhythmia in fish - both under heat stress and in the presence of toxic substances - is atrioventricular block, which is the inability of the action potential to progress from the atrium to the ventricle. Early and delayed afterdepolarizations are less common in fish hearts than in the hearts of endotherms, perhaps owing to the excitation-contraction coupling properties of the fish heart. In fish hearts, Ca2+-induced Ca2+ release from the sarcoplasmic reticulum plays a smaller role than Ca2+ influx through the sarcolemma. Environmental changes and ion channel toxins can induce arrhythmias in fish and weaken their tolerance to environmental stresses. Although different from endotherm hearts in many respects, fish hearts can serve as a translational model for studying human cardiac arrhythmias, especially for human neonates.
Asunto(s)
Arritmias Cardíacas , Peces , Animales , Arritmias Cardíacas/fisiopatología , Arritmias Cardíacas/etiología , Peces/fisiología , Ambiente , Calcio/metabolismoRESUMEN
Polycyclic aromatic hydrocarbons (PAHs) are embryo- and cardiotoxic to fish that might be associated with improper intracellular Ca2+ management. Since sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) is a major regulator of intracellular Ca2+, the SERCA activity and the contractile properties of rainbow trout (Oncorhynchus mykiss) ventricle were measured in the presence of 3- and 4-cyclic PAHs. In unfractionated ventricular homogenates, acute exposure of SERCA to 0.1-1.0 µM phenanthrene (Phe), retene (Ret), fluoranthene (Flu), or pyrene (Pyr) resulted in concentration-dependent increase in SERCA activity, except for the Flu exposure, with maximal effects of 49.7-83 % at 1 µM. However, PAH mixture did not affect the contractile parameters of trout ventricular strips. Similarly, all PAHs, except Ret, increased the myotomal SERCA activity, but with lower effect (27.8-40.8 % at 1 µM). To investigate the putative chronic effects of PAHs on SERCA, the atp2a2a gene encoding trout cardiac SERCA was expressed in human embryonic kidney (HEK) cells. Culture of HEK cells in the presence of 0.3-1.0 µM Phe, Ret, Flu, and Pyr for 4 days suppressed SERCA expression in a concentration-dependent manner, with maximal inhibition of 49 %, 65 %, 39 % (P < 0.05), and 18 % (P > 0.05), respectively at 1 µM. Current findings indicate divergent effects of submicromolar PAH concentrations on SERCA: stimulation of SERCA activity in acute exposure and inhibition of SERCA expression in chronic exposure. The depressed expression of SERCA is likely to contribute to the embryo- and cardiotoxicity of PAHs by depressing muscle function and altering gene expression.
Asunto(s)
Oncorhynchus mykiss , Fenantrenos , Hidrocarburos Policíclicos Aromáticos , Animales , Humanos , Oncorhynchus mykiss/metabolismo , Fenantrenos/toxicidad , Fenantrenos/metabolismo , Hidrocarburos Policíclicos Aromáticos/toxicidad , Hidrocarburos Policíclicos Aromáticos/metabolismo , ATPasas Transportadoras de Calcio/metabolismo , Estrés del Retículo Endoplásmico , Calcio/metabolismoRESUMEN
Heavy metals are harmful to aquatic animals by disrupting their ionic balance. Here, we compare the effects of three metals, zinc (Zn), nickel (Ni) and manganese (Mn) on Na,K-ATPase activity in gills and kidneys in fish species with different ecophysiological characteristics. Crucian carp (Carassius carassius), a cold-dormant species, and rainbow trout (Oncorhynchus mykiss), a cold-active species, were acclimated to 2 °C and 18 °C, and branchial and renal Na,K-ATPase activities were measure in the presence of Zn, Ni and Mn. Under basal conditions, species-, tissues- and temperature-dependent differences appeared in Na,K-ATPase activity. Renal Na,K-ATPase activity was higher in trout than carp, and cold-acclimation increased Na,K-ATPase activity in both species. Cold-acclimation reduced branchial Na,K-ATPase activity in carp, but no acclimation effect was found in trout. In both species and tissues, Zn stimulated Na,K-ATPase in concentration-dependent manner at 0.1 to 3 µM. At 30 µM, Zn strongly inhibited both branchial and renal Na,K-ATPase in both species. Inhibition by Zn was stronger in trout than carp, but no differences existed between acclimation groups in either species. Ni (0.1-3.0 µM) stimulated renal Na,K-ATPase in crucian carp but not in rainbow trout. At 30 µM, Ni depressed the renal Na,K-ATPase of carp back to the control level. Mn had no statistically significant effect on Na,K-ATPase in either species. At low concentrations, Zn and Ni impose an energetic cost to fish by increasing ATP consumption in Na,K-ATPase activity. At higher concentrations, Zn, but not Ni and Mn, strongly inhibit renal and branchial Na,K-ATPase. Due to differences in baseline activity level and acclimation-induced changes in renal and branchial Na,K-ATPase, metal pollution may impair ion regulation of fish in species-specific manner and depending on season.
Asunto(s)
Carpas , Oncorhynchus mykiss , Contaminantes Químicos del Agua , Animales , Carpas/metabolismo , Oncorhynchus mykiss/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Contaminantes Químicos del Agua/toxicidad , Metales/farmacología , Sodio/metabolismo , Riñón/metabolismo , BranquiasRESUMEN
ABSTRACT: In cardiac myocytes, the slow component of the delayed rectifier K+ current (IKs) ensures repolarization of action potential during beta-adrenergic activation or when other repolarizing K+ currents fail. As a key factor of cardiac repolarization, IKs should be present in model species used for cardiovascular drug screening, preferably with pharmacological characteristics similar to those of the human IKs. To this end, we investigated the effects of inhibitors and activators of the IKs on KCNQ1 and KCNQ1 + KCNE1 channels of the zebrafish, an important model species, in Chinese hamster ovary cells. Inhibitors of IKs, chromanol 293B and HMR-1556, inhibited zebrafish IKs channels with approximately similar potency as that of mammalian IKs. Chromanol 293B concentration for half-maximal inhibition (IC50) of zebrafish IKs was at 13.1 ± 5.8 and 13.4 ± 2.8 µM for KCNQ1 and KCNQ1+KCNE1 channels, respectively. HMR-1556 was a more potent inhibitor of zebrafish IKs channels with IC50 = 0.1 ± 0.1 µM and 1.5 ± 0.8 µM for KCNQ1 and KCNQ1 + KCNE1 channels, respectively. R-L3 and mefenamic acid, generally identified as IKs activators, both inhibited zebrafish IKs. R-L3 almost completely inhibited the current generated by KCNQ1 and KCNQ1 + KCNE1 channels with similar potency (IC50 1.1 ± 0.4 and 1.0 ± 0.4 µM, respectively). Mefenamic acid partially blocked zebrafish KCNQ1 (IC50 = 9.5 ± 4.8 µM) and completely blocked KCNQ1 + KCNE1 channels (IC50 = 3.3 ± 1.8 µM). Although zebrafish IKs channels respond to IKs inhibitors in the same way as mammalian IKs channels, their response to activators is atypical, probably because of the differences in the binding domain of KCNE1 to KCNQ1. Therefore, care must be taken when translating the results from zebrafish to humans.
Asunto(s)
Canal de Potasio KCNQ1 , Canales de Potasio con Entrada de Voltaje , Animales , Células CHO , Cricetinae , Cricetulus , Canal de Potasio KCNQ1/genética , Canal de Potasio KCNQ1/metabolismo , Ácido Mefenámico , Canales de Potasio con Entrada de Voltaje/metabolismo , Pez CebraRESUMEN
Cadmium (Cd2+ ) is cardiotoxic to fish, but its effect on the electrical excitability of cardiac myocytes is largely unknown. To this end, we used the whole-cell patch-clamp method to investigate the effects of Cd2+ on ventricular action potentials (APs) and major ion currents in rainbow trout (Oncorhynchus mykiss) ventricular myocytes. Trout were acclimated to +4 °C, and APs were measured at the acclimated temperature and elevated temperature (+18 °C). Cd2+ (10, 20, and 100 µM) altered the shape of the ventricular AP in a complex manner. The early plateau fell to less positive membrane voltages, and the total duration of AP prolonged. These effects were obvious at both +4 °C and +18 °C. The depression of the early plateau is due to the strong Cd2+ -induced inhibition of the L-type calcium (Ca2+ ) current (ICaL ), whereas the prolongation of the AP is an indirect consequence of the ICaL inhibition: at low voltages of the early plateau, the delayed rectifier potassium (K+ ) current (IKr ) remains small, delaying repolarization of AP. Cd2+ reduced the density and slowed the kinetics of the Na+ current (INa ) but left the inward rectifier K+ current (IK1 ) intact. These altered cellular and molecular functions can explain several Cd2+ -induced changes in impulse conduction of the fish heart, for example, slowed propagation of the AP in atrial and ventricular myocardia (inhibition of INa ), delayed relaxation of the ventricle (prolongation of ventricular AP duration), bradycardia, and atrioventricular block (inhibition of ICaL ). These findings indicate that the cardiotoxicity of Cd2+ in fish involves multiple ion currents that are directly and indirectly altered by Cd2+ . Through these mechanisms, Cd2+ may trigger cardiac arrhythmias and impair myocardial contraction. Elevated temperature (+18 °C) slightly increases Cd2+ toxicity in trout ventricular myocytes. Environ Toxicol Chem 2021;40:2874-2885. © 2021 SETAC.
Asunto(s)
Oncorhynchus mykiss , Potenciales de Acción , Animales , Cadmio/toxicidad , Cardiotoxicidad , Células Musculares , Miocitos Cardíacos/fisiologíaRESUMEN
Heat tolerance of heart rate in fish is suggested to be limited by impaired electrical excitation of the ventricle due to the antagonistic effects of high temperature on Na+ (INa) and K+ (IK1) ion currents (INa is depressed at high temperatures while IK1 is resistant to them). To examine the role of Na+ channel proteins in heat tolerance of INa, we compared temperature dependencies of zebrafish (Danio rerio, warm-dwelling subtropical species) and rainbow trout (Oncorhynchus mykiss, cold-active temperate species) ventricular INa, and INa generated by the cloned zebrafish and rainbow trout NaV1.4 and NaV1.5 Na+ channels in human embryonic kidney (HEK) cells. Whole-cell patch-clamp recordings showed that zebrafish ventricular INa has better heat tolerance and slower inactivation kinetics than rainbow trout ventricular INa. In contrast, heat tolerance and inactivation kinetics of zebrafish and rainbow trout NaV1.4 channels are similar when expressed in the identical cellular environment of HEK cells. The same applies to NaV1.5 channels. These findings indicate that thermal adaptation of ventricular INa is largely achieved by differential expression of Na+ channel alpha subunits: zebrafish that tolerate higher temperatures mainly express the slower NaV1.5 isoform, while rainbow trout that prefer cold waters mainly express the faster NaV1.4 isoform. Differences in elasticity (stiffness) of the lipid bilayer and/or accessory protein subunits of the channel assembly may also be involved in thermal adaptation of INa. The results are consistent with the hypothesis that slow Na+ channel kinetics are associated with increased heat tolerance of cardiac excitation.
Asunto(s)
Oncorhynchus mykiss , Animales , Corazón , Humanos , Isoformas de Proteínas/genética , Temperatura , Pez CebraRESUMEN
At critically high temperature, cardiac output in fish collapses as a result of depression of heart rate (bradycardia). However, the cause of bradycardia remains unresolved. To investigate this, rainbow trout (Oncorhynchus mykiss; acclimated at 12°C) were exposed to acute warming while electrocardiograms were recorded. From 12°C to 25.3°C, electrical excitation between different parts of the heart was coordinated, but above 25.3°C, atrial and ventricular beating rates became partly dissociated because of 2:1 atrioventricular (AV) block. With further warming, atrial rate increased to a peak value of 188±22 beats min-1 at 27°C, whereas the ventricle rate peaked at 124±10 beats min-1 at 25.3°C and thereafter dropped to 111±15 beats min-1 at 27°C. In single ventricular myocytes, warming from 12°C to 25°C attenuated electrical excitability as evidenced by increases in rheobase current and the size of critical depolarization required to trigger action potential. Depression of excitability was caused by temperature-induced decrease in input resistance (sarcolemmal K+ leak via the outward IK1 current) of resting myocytes and decrease in inward charge transfer by the Na+ current (INa) of active myocytes. Collectively, these findings show that at critically high temperatures AV block causes ventricular bradycardia owing to the increased excitation threshold of the ventricle, which is due to changes in the passive (resting ion leak) and active (inward charge movement) electrical properties of ventricular myocytes. The sequence of events from the level of ion channels to cardiac function in vivo provides a mechanistic explanation for the depression of cardiac output in fish at critically high temperature.
Asunto(s)
Bloqueo Atrioventricular , Oncorhynchus mykiss , Aclimatación , Animales , Bloqueo Atrioventricular/veterinaria , Corazón , Frecuencia Cardíaca , Ventrículos Cardíacos , Miocitos Cardíacos , TemperaturaRESUMEN
Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous contaminants in aqueous environments. They affect cardiovascular development and function in fishes. The 3-ring PAH phenanthrene has recently been shown to impair cardiac excitation-contraction coupling by inhibiting Ca2+ and K+ currents in marine warm-water scombrid fishes. To see if similar events take place in a boreal freshwater fish, we studied whether the PAHs phenanthrene and retene (an alkylated phenanthrene) modify the action potential (AP) via effects on Na+ (INa ), Ca2+ (ICaL ), or K+ (IKr , IK1 ) currents in the ventricular myocytes of the rainbow trout (Oncorhynchus mykiss) heart. Electrophysiological characteristics of myocytes were measured using whole-cell patch clamp. Micromolar concentrations of phenanthrene and retene modified the shape of the ventricular AP, and retene profoundly shortened the AP at low micromolar concentrations. Both PAHs increased INa and reduced ICaL and IKr , but retene was more potent. Neither of the PAHs had an effect on IK1 . Our results show that phenanthrene and retene affect cardiac function in rainbow trout by a mechanism that involves multiple cardiac ion channels, and the final outcome of these changes (shortening of AP) is opposite to that observed in scombrid fishes (prolongation of AP). The results also show that retene and aryl hydrocarbon receptor (AhR) agonist have an additional mechanism of toxicity besides the previously known AhR-mediated, transcription-dependent one. Environ Toxicol Chem 2019;38:2145-2153. © 2019 SETAC.
Asunto(s)
Potenciales de Acción/efectos de los fármacos , Miocitos Cardíacos/efectos de los fármacos , Oncorhynchus mykiss/metabolismo , Hidrocarburos Policíclicos Aromáticos/toxicidad , Contaminantes Químicos del Agua/toxicidad , Animales , Miocitos Cardíacos/fisiología , Fenantrenos/toxicidad , Receptores de Hidrocarburo de Aril/agonistas , Receptores de Hidrocarburo de Aril/metabolismo , Contaminantes Químicos del Agua/químicaRESUMEN
Electrical excitability (EE) is vital for cardiac function and strongly modulated by temperature and external K+ concentration ([K+]o), as formulated in the hypothesis of temperature-dependent deterioration of electrical excitability (TDEE). As little is known about EE of arctic stenothermic fishes, we tested the TDEE hypothesis on ventricular myocytes of polar cod (Boreogadus saida) and navaga (Eleginus nawaga) of the Arctic Ocean and those of temperate freshwater burbot (Lota lota). Ventricular action potentials (APs) were elicited in current-clamp experiments at 3, 9 and 15°C, and AP characteristics and the current needed to elicit APs were examined. At 3°C, ventricular APs of polar cod and navaga were similar but differed from those of burbot in having a lower rate of AP upstroke and a higher rate of repolarization. EE of ventricular myocytes - defined as the ease with which all-or-none APs are triggered - was little affected by acute temperature changes between 3 and 15°C in any species. However, AP duration (APD50) was drastically reduced at higher temperatures. Elevation of [K+]o from 3 to 5.4â mmolâ l-1 and further to 8â mmolâ l-1 at 3, 9 and 15°C strongly affected EE and AP characteristics in polar cod and navaga, but had a lesser effect in burbot. In all species, ventricular excitation was resistant to acute temperature elevations, while small increases in [K+]o severely compromised EE, in particular in the marine stenotherms. This suggests that EE of the heart in these Gadiformes species is resistant against acute warming, but less so against the simultaneous temperature and exercise stresses.
Asunto(s)
Potenciales de Acción/fisiología , Gadiformes/fisiología , Miocitos Cardíacos/fisiología , Potasio/fisiología , Función Ventricular/fisiología , Animales , Femenino , Masculino , TemperaturaRESUMEN
Calcium channels are necessary for cardiac excitation-contraction (E-C) coupling, but Ca2+ channel composition of fish hearts is still largely unknown. To this end, we determined transcript expression of Ca2+ channels in the heart of zebrafish (Danio rerio), a popular model species. Altogether, 18 Ca2+ channel α-subunit genes were expressed in both atrium and ventricle. Transcripts for 7 L-type (Cav1.1a, Cav1.1b, Cav1.2, Cav1.3a, Cav1.3b, Cav1.4a, Cav1.4b), 5 T-type (Cav3.1, Cav3.2a, Cav3.2b, Cav3.3a, Cav3.3b) and 6 P/Q-, N- and R-type (Cav2.1a, Cav2.1b, Cav2.2a, Cav2.2b, Cav2.3a, Cav2.3b) Ca2+ channels were expressed. In the ventricle, T-type channels formed 54.9%, L-type channels 41.1% and P/Q-, N- and R-type channels 4.0% of the Ca2+ channel transcripts. In the atrium, the relative expression of T-type and L-type Ca2+ channel transcripts was 64.1% and 33.8%, respectively (others accounted for 2.1%). Thus, at the transcript level, T-type Ca2+ channels are prevalent in zebrafish atrium and ventricle. At the functional level, peak densities of ventricular T-type (ICaT) and L-type (ICaL) Ca2+ current were 6.3±0.8 and 7.7±0.8â pAâ pF-1, respectively. ICaT mediated a sizeable sarcolemmal Ca2+ influx into ventricular myocytes: the increment in total cellular Ca2+ content via ICaT was 41.2±7.3â µmol l-1, which was 31.7% of the combined Ca2+ influx (129â µmol l-1) via ICaT and ICaL (88.5±20.5â µmol l-1). The diversity of expressed Ca2+ channel genes in zebrafish heart is high, but dominated by the members of the T-type subfamily. The large ventricular ICaT is likely to play a significant role in E-C coupling.
Asunto(s)
Canales de Calcio/metabolismo , Regulación de la Expresión Génica , Pez Cebra/fisiología , Animales , Calcio/metabolismo , Canales de Calcio/genética , Canales de Calcio Tipo T/genética , Canales de Calcio Tipo T/metabolismo , Atrios Cardíacos/metabolismo , Ventrículos Cardíacos/metabolismo , Células Musculares/metabolismo , Técnicas de Placa-Clamp , Pez Cebra/genéticaRESUMEN
Na+ channel α-subunit composition of the zebrafish heart and electrophysiological properties of Na+ current (INa) of zebrafish ventricular myocytes were examined. Eight Na+ channel α-subunits were expressed in both atrium and ventricle of the zebrafish heart. Nav1.5Lb, an orthologue to the human Nav1.5, was clearly the predominant isoform in both chambers representing 65.2⯱â¯4.1% and 83.1⯱â¯2.1% of all Na+ channel transcripts in atrium and ventricle, respectively. Nav1.4b, an orthologue to human Nav1.4, formed 34.1⯱â¯4.1 and 16.2⯱â¯2.0% of the Na+ channel transcripts in atrium and ventricle, respectively. The density of INa and the rate of action potential upstroke in zebrafish ventricular myocytes at 28⯰C were similar to those of human ventricles at the comparable temperature. Na+ channel isoforms and the main electrophysiological characteristics of the INa are largely similar in zebrafish and human hearts indicating evolutionary conservation of Na+ channel composition and function. The zebrafish INa differs from the human cardiac INa in terms of higher tetrodotoxin sensitivity (IC50-valueâ¯=â¯5.3⯱â¯0.1â¯nM) and slower inactivation kinetics. The zebrafish INa was inhibited with tricaine (MS-222) with an IC50-value of 1.2⯱â¯0.18â¯mM (336â¯mgâ¯l-1), suggesting some care in the use of MS-222 as an anesthetic.
Asunto(s)
Fenómenos Electrofisiológicos , Regulación de la Expresión Génica , Ventrículos Cardíacos/metabolismo , Sodio/metabolismo , Función Ventricular , Canales de Sodio Activados por Voltaje/metabolismo , Pez Cebra , Aminobenzoatos/farmacología , Animales , Fenómenos Electrofisiológicos/efectos de los fármacos , Ventrículos Cardíacos/efectos de los fármacos , Cinética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Tetrodotoxina/farmacología , Función Ventricular/efectos de los fármacosRESUMEN
Funny current (If), formed by hyperpolarization-activated cyclic nucleotide-gated channels (HCN channels), is supposed to be crucial for the membrane clock regulating the cardiac pacemaker mechanism. We examined the presence and activity of HCN channels in the brown trout (Salmo trutta fario) sinoatrial (SA) pacemaker cells and their putative role in heart rate (fH) regulation. Six HCN transcripts (HCN1, HCN2a, HCN2ba, HCN2bb, HCN3, and HCN4) were expressed in the brown trout heart. The total HCN transcript abundance was 4.0 and 4.9 times higher in SA pacemaker tissue than in atrium and ventricle, respectively. In the SA pacemaker, HCN3 and HCN4 were the main isoforms representing 35.8 ± 2.7 and 25.0 ± 1.5%, respectively, of the total HCN transcripts. Only a small If with a mean current density of -1.2 ± 0.37 pA/pF at -140 mV was found in 4 pacemaker cells out of 16 spontaneously beating cells examined, despite the optimization of recording conditions for If activity. If was not found in any of the 24 atrial myocytes and 21 ventricular myocytes examined. HCN4 coexpressed with the MinK-related peptide 1 (MiRP1) ß-subunit in CHO cells generated large If currents. In contrast, HCN3 (+MiRP1) failed to produce If in the same expression system. Cs+ (2 mM), which blocked 84 ± 12% of the native If, reversibly reduced fH 19.2 ± 3.6% of the excised multicellular pacemaker tissue from 53 ± 5 to 44 ± 5 beats/min (P < 0.05). However, this effect was probably due to the reduction of IKr, which was also inhibited (63.5 ± 4.6%) by Cs+ These results strongly suggest that fH regulation in the brown trout heart is largely independent on If.
Asunto(s)
Relojes Biológicos/fisiología , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/metabolismo , Miocardio/citología , Miocardio/metabolismo , Nodo Sinoatrial/fisiología , Trucha/metabolismo , Secuencia de Aminoácidos , Animales , Células CHO , Cesio/farmacología , Cricetinae , Cricetulus , Regulación de la Expresión Génica/fisiología , Atrios Cardíacos/citología , Atrios Cardíacos/metabolismo , Frecuencia Cardíaca/fisiología , Ventrículos Cardíacos/citología , Ventrículos Cardíacos/metabolismo , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/antagonistas & inhibidores , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/genética , Miocitos Cardíacos/metabolismo , Técnicas de Placa-ClampRESUMEN
The effects of sustained anoxia on cardiac electrical excitability were examined in the anoxia-tolerant crucian carp (Carassius carassius). The electrocardiogram (ECG) and expression of excitation-contraction coupling genes were studied in fish acclimatised to normoxia in summer (+18°C) or winter (+2°C), and in winter fish after 1, 3 and 6â weeks of anoxia. Anoxia induced a sustained bradycardia from a heart rate of 10.3±0.77 beats min-1 to 4.1±0.29 beats min-1 (P<0.05) after 5â weeks, and heart rate slowly recovered to control levels when oxygen was restored. Heart rate variability greatly increased under anoxia, and completely recovered under re-oxygenation. The RT interval increased from 2.8±0.34â s in normoxia to 5.8±0.44â s under anoxia (P<0.05), which reflects a doubling of the ventricular action potential (AP) duration. Acclimatisation to winter induced extensive changes in gene expression relative to summer-acclimatised fish, including depression in those genes coding for the sarcoplasmic reticulum calcium pump (Serca2a_q2) and ATP-sensitive K+ channels (Kir6.2) (P<0.05). Genes of delayed rectifier K+ (kcnh6) and Ca2+ channels (cacna1c) were up-regulated in winter fish (P<0.05). In contrast, the additional challenge of anoxia caused only minor changes in gene expression, e.g. depressed expression of Kir2.2b K+ channel gene (kcnj12b), whereas expression of Ca2+ (cacna1a, cacna1c and cacna1g) and Na+ channel genes (scn4a and scn5a) was not affected. These data suggest that low temperature pre-conditions the crucian carp heart for winter anoxia, whereas sustained anoxic bradycardia and prolongation of AP duration are directly induced by oxygen shortage without major changes in gene expression.
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Adaptación Fisiológica , Carpas/fisiología , Oxígeno/metabolismo , Aclimatación , Potenciales de Acción , Anaerobiosis , Animales , Carpas/genética , Electrocardiografía , Regulación de la Expresión Génica , Corazón/fisiología , Frecuencia Cardíaca , Estaciones del AñoRESUMEN
Temperature-induced changes in cardiac output (QÌ) in fish are largely dependent on thermal modulation of heart rate (fH), and at high temperatures QÌ collapses due to heat-dependent depression of fH This study tests the hypothesis that firing rate of sinoatrial pacemaker cells sets the upper thermal limit of fH in vivo. To this end, temperature dependence of action potential (AP) frequency of enzymatically isolated pacemaker cells (pacemaker rate, fPM), spontaneous beating rate of isolated sinoatrial preparations (fSA), and in vivo fH of the cold-acclimated (4°C) brown trout (Salmo trutta fario) were compared under acute thermal challenges. With rising temperature, fPM steadily increased because of the acceleration of diastolic depolarization and shortening of AP duration up to the break point temperature (TBP) of 24.0 ± 0.37°C, at which point the electrical activity abruptly ceased. The maximum fPM at TBP was much higher [193 ± 21.0 beats per minute (bpm)] than the peak fSA (94.3 ± 6.0 bpm at 24.1°C) or peak fH (76.7 ± 2.4 at 15.7 ± 0.82°C) (P < 0.05). These findings strongly suggest that the frequency generator of the sinoatrial pacemaker cells does not limit fH at high temperatures in the brown trout in vivo.
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Potenciales de Acción/fisiología , Relojes Biológicos/fisiología , Regulación de la Temperatura Corporal/fisiología , Frecuencia Cardíaca/fisiología , Miocitos Cardíacos/fisiología , Trucha/fisiología , Animales , Sistema de Conducción Cardíaco/citología , Sistema de Conducción Cardíaco/fisiologíaRESUMEN
Mobilization of glycogen stores was examined in the anoxic crucian carp (Carassius carassius Linnaeus). Winter-acclimatized fish were exposed to anoxia for 1, 3, or 6 weeks at 2 °C, and changes in the size of glycogen deposits were followed. After 1 week of anoxia, a major part of the glycogen stores was mobilized in liver (79.5 %) and heart (75.6 %), and large decreases occurred in gill (46.7 %) and muscle (45.1 %). Brain was an exception in that its glycogen content remained unchanged. The amount of glycogen degraded during the first anoxic week was sufficient for the anaerobic ethanol production for more than 6 weeks of anoxia. After 3 and 6 weeks of anoxia, there was little further degradation of glycogen in other tissues except the brain where the stores were reduced by 30.1 and 49.9 % after 3 and 6 weeks of anoxia, respectively. One week of normoxic recovery following the 6-week anoxia was associated with a complete replenishment of the brain glycogen and partial recovery of liver, heart, and gill glycogen stores. Notably, the resynthesis of glycogen occurred at the expense of the existing energy reserves of the body in fasting fish. These findings indicate that in crucian carp, glycogen stores are quickly mobilized after the onset of anoxia, with the exception of the brain whose glycogen stores may be saved for putative emergency situations.
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Carpas/metabolismo , Glucógeno/metabolismo , Hipoxia/metabolismo , Aclimatación , Acetatos/metabolismo , Amoníaco/metabolismo , Animales , Peso Corporal , Encéfalo/metabolismo , Etanol/metabolismo , Branquias/metabolismo , Corazón/fisiología , Ácido Láctico/metabolismo , Hígado/crecimiento & desarrollo , Hígado/metabolismo , Tamaño de los Órganos , Bazo/crecimiento & desarrolloRESUMEN
Pyrethroids are extensively used for the control of insect pests and disease vectors. Pyrethroids are regarded safe due to their selective toxicity: they are effective against insects but relatively harmless to mammals and birds. Unfortunately, pyrethroids are very toxic to fishes. The high toxicity of pyrethroids to fishes is only partly explained by slow metabolic elimination of pyrethroids, suggesting that some molecular targets in vital organs of the fish body are sensitive to pyrethroids. To this end we tested the effect of deltamethrin (DM) on fish (crucian carp, Carassius carassius) heart function in vitro. In sinoatrial preparations of the crucian carp heart DM (10 µM) caused irregularities in rate and rhythm of atrial beating and strong reductions in force of atrial contraction, thus indicating that DM is arrhythmogenic to the fish heart. Consistent with this, DM (10.0 µM) induced irregularities in electrical activity (surface electrocardiogram) of spontaneous beating hearts in vitro. In isolated ventricular myocytes, DM (0.1-30.0 µM) modified Na(+) current by slowing channel closing and shifting reversal potential and steady-state activation of the current to more negative voltages. Maximally about 48% of the cardiac Na(+) channels were affected by DM with a half-maximal effect occurring at the concentration of 1.3 µM. These findings indicate that DM can be cardiotoxic to the crucian carp and that these effects could be due to DM related changes in Na(+) channel function. These findings indicate that in addition to their neurotoxicity effects pyrethroid could also be cardiotoxic to fishes.
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Corazón/efectos de los fármacos , Nitrilos/toxicidad , Piretrinas/toxicidad , Animales , Carpas , Electrocardiografía , Corazón/fisiologíaRESUMEN
Seasonal changes in physiology of vertebrate animals are triggered by environmental cues including temperature, day-length and oxygen availability. Crucian carp (Carassius carassius) tolerate prolonged anoxia in winter by using several physiological adaptations that are seasonally activated. This study examines which environmental cues are required to trigger physiological adjustments for winter dormancy in crucian carp. To this end, crucian carp were exposed to changing environmental factors under laboratory conditions: effects of declining water temperature, shortening day-length and reduced oxygen availability, separately and in different combinations, were examined on glycogen content and enzyme activities involved in feeding (alkaline phosphatase, AP) and glycogen metabolism (glycogen synthase, GyS; glycogen phosphorylase, GP). Lowering temperature induced a fall in activity of AP and a rise in glycogen content and rate of glycogen synthesis. Relative mass of the liver, and glycogen concentration of liver, muscle and brain increased with lowering temperature. Similarly activity of GyS in muscle and expression of GyS transcripts in brain were up-regulated by lowering temperature. Shortened day-length and oxygen availability had practically no effects on measured variables. We conclude that lowering temperature is the main trigger in preparation for winter anoxia in crucian carp.
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Carpas/fisiología , Ayuno/fisiología , Glucógeno/metabolismo , Estaciones del Año , Temperatura , Fosfatasa Alcalina/genética , Fosfatasa Alcalina/metabolismo , Animales , Regulación Enzimológica de la Expresión Génica , Glucógeno Fosforilasa/genética , Glucógeno Fosforilasa/metabolismo , Glucógeno Sintasa/genética , Glucógeno Sintasa/metabolismo , Intestinos/enzimología , Hígado/anatomía & histologíaRESUMEN
Fishes are increasingly used as models for human cardiac diseases, creating a need for a better understanding of the molecular basis of fish cardiac ion currents. To this end we cloned KCNH6 channel of the crucian carp (Carassius carassius) that produces the rapid component of the delayed rectifier K(+) current (IKr), the main repolarising current of the fish heart. KCNH6 (ccErg2) was the main isoform of the Kv11 potassium channel family with relative transcript levels of 98.9% and 99.6% in crucian carp atrium and ventricle, respectively. KCNH2 (ccErg1), an orthologue to human cardiac Erg (Herg) channel, was only slightly expressed in the crucian carp heart. The native atrial IKr and the cloned ccErg2 were inhibited by similar concentrations of verapamil, terfenadine and KB-R7943 (P>0.05), while the atrial IKr was about an order of magnitude more sensitive to E-4031 than ccErg2 (P<0.05) suggesting that some accessory ß-subunits may be involved. Sensitivity of the crucian carp atrial IKr to E-4031, terfenadine and KB-R7943 was similar to what has been reported for the Herg channel. In contrast, the sensitivity of the crucian carp IKr to verapamil was approximately 30 times higher than the previously reported values for the Herg current. In conclusion, the cardiac IKr is produced by non-orthologous gene products in fish (Erg2) and mammalian hearts (Erg1) and some marked differences exist in drug sensitivity between fish and mammalian Erg1/2 which need to be taken into account when using fish heart as a model for human heart.
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Carpas/metabolismo , Canales de Potasio de Tipo Rectificador Tardío/antagonistas & inhibidores , Canales de Potasio Éter-A-Go-Go/antagonistas & inhibidores , Proteínas de Peces/antagonistas & inhibidores , Corazón/efectos de los fármacos , Miocardio/metabolismo , Bloqueadores de los Canales de Potasio/farmacología , Secuencia de Aminoácidos , Animales , Células CHO , Carpas/genética , Clonación Molecular , Cricetulus , Canales de Potasio de Tipo Rectificador Tardío/genética , Canales de Potasio de Tipo Rectificador Tardío/metabolismo , Relación Dosis-Respuesta a Droga , Canal de Potasio ERG1 , Canales de Potasio Éter-A-Go-Go/genética , Canales de Potasio Éter-A-Go-Go/metabolismo , Femenino , Proteínas de Peces/genética , Proteínas de Peces/metabolismo , Masculino , Potenciales de la Membrana , Modelos Animales , Datos de Secuencia Molecular , Potasio/metabolismo , ARN Mensajero/metabolismo , Especificidad de la Especie , TransfecciónRESUMEN
Electrophysiological properties and molecular background of the zebrafish (Danio rerio) cardiac inward rectifier current (IK1) were examined. Ventricular myocytes of zebrafish have a robust (-6.7 ± 1.2 pA pF(-1) at -120 mV) strongly rectifying and Ba(2+)-sensitive (IC50 = 3.8 µM) IK1. Transcripts of six Kir2 channels (drKir2.1a, drKir2.1b, drKir2.2a, drKir2.2b, drKir2.3, and drKir2.4) were expressed in the zebrafish heart. drKir2.4 and drKir2.2a were the dominant isoforms in both the ventricle (92.9 ± 1.5 and 6.3 ± 1.5%) and the atrium (28.9 ± 2.9 and 64.7 ± 3.0%). The remaining four channels comprised together less than 1 and 7 % of the total transcripts in ventricle and atrium, respectively. The four main gene products (drKir2.1a, drKir2.2a, drKir2.2b, drKir2.4) were cloned, sequenced, and expressed in HEK cells for electrophysiological characterization. drKir2.1a was the most weakly rectifying (passed more outward current) and drKir2.2b the most strongly rectifying (passed less outward current) channel, whilst drKir2.2a and drKir2.4 were intermediate between the two. In regard to sensitivity to Ba(2+) block, drKir2.4 was the most sensitive (IC50 = 1.8 µM) and drKir2.1a the least sensitive channel (IC50 = 132 µM). These findings indicate that the Kir2 isoform composition of the zebrafish heart markedly differs from that of mammalian hearts. Furthermore orthologous Kir2 channels (Kir2.1 and Kir2.4) of zebrafish and mammals show striking differences in Ba(2+)-sensitivity. Structural and functional differences needs to be taken into account when zebrafish is used as a model for human cardiac electrophysiology, cardiac diseases, and in screening cardioactive substances.
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Potenciales de Acción , Miocitos Cardíacos/metabolismo , Canales de Potasio de Rectificación Interna/metabolismo , Multimerización de Proteína , Proteínas de Pez Cebra/metabolismo , Secuencia de Aminoácidos , Animales , Bario/farmacología , Células Cultivadas , Células HEK293 , Atrios Cardíacos/citología , Ventrículos Cardíacos/citología , Humanos , Datos de Secuencia Molecular , Miocitos Cardíacos/fisiología , Bloqueadores de los Canales de Potasio/farmacología , Canales de Potasio de Rectificación Interna/química , Canales de Potasio de Rectificación Interna/genética , Subunidades de Proteína/química , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Pez Cebra , Proteínas de Pez Cebra/química , Proteínas de Pez Cebra/genéticaRESUMEN
Hagfishes and lampreys (order Cyclostomata) are living representatives of an ancient group of jawless vertebrates (class Agnatha). Studies on cyclostome hearts may provide insights into the evolution of the vertebrate heart and thereby increase our understanding of cardiac function in higher vertebrates, including mammals. To this end, electrical excitability of the heart in a basal vertebrate, the European river lamprey (Lampetra fluviatilis), was examined. Ion currents of cardiac myocytes, action potentials (APs) of atrial and ventricular muscle, and electrocardiogram (in vivo) were measured using the patch-clamp method, intracellular microelectrodes, and trailing wires, respectively. The characteristic features of fairly high heart rate (28.4 ± 3 beats min(-1)) and short AP duration (550 ± 44 and 122.1 ± 28.5 for ventricle and atrium, respectively) at low ambient temperature (5°C) are shared with cold-active teleost fishes. However, the ion current basis of the ventricular AP differs from that of other fishes. For inward currents, sodium current density (INa) is lower and calcium current density (ICa) higher than in teleost ventricles, while the kinetics of INa is slow and that of ICa is fast in comparison. Among the ventricular repolarizing currents, the delayed rectifier K(+) current is smaller than in myocytes of several teleost species. Unlike mammalian hearts, ATP-sensitive K(+) channels are constitutively open under normoxic conditions, thus contributing to negative resting membrane potential and repolarization of APs. Upstroke velocity of AP (5.4 ± 0.9 and 6.3 ± 0.6 V s(-1) for ventricular and atrial myocytes, respectively) is slower than in teleost hearts. Excitability of the lamprey heart seems to possess both primitive and advanced characteristics. Short APs are appropriate to support brief and vigorous contractions (in common with higher vertebrates), while relatively low AP upstroke velocities enable only relatively slow propagation of contraction over the heart.