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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 [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

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 atriumand 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±0.8 pA 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±7.3 μmol l-1, which was 31.7% of the combined Ca2+ influx (129 μmol l-1) viaICaT and ICaL (88.5±20.5 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. [ABSTRACT FROM AUTHOR]

Electrophysiological properties and molecular background of the zebrafish ( Danio rerio) cardiac inward rectifier current (I) were examined. Ventricular myocytes of zebrafish have a robust (−6.7 ± 1.2 pA pF at −120 mV) strongly rectifying and Ba-sensitive (IC = 3.8 μM) I. 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 block, drKir2.4 was the most sensitive (IC = 1.8 μM) and drKir2.1a the least sensitive channel (IC = 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-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. [ABSTRACT FROM AUTHOR]

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 micro-electrodes, 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. [ABSTRACT FROM AUTHOR]

Ectothermic vertebrates experience acute and chronic temperature changes which affect cardiac excitability and may threaten electrical stability of the heart. Nevertheless, ectothermic hearts function over wide range of temperatures without cardiac arrhythmias, probably due to special molecular adaptations. We examine function and molecular basis of the slow delayed rectifier K+ current (IKs) in cardiac myocytes of a eurythermic fish (Carassius carassius L.). IKs is an important repolarizing current that prevents excessive prolongation of cardiac action potential, but it is extremely slowly activating when expressed in typical molecular composition of the endothermic animals. Comparison of the IKs of the crucian carp atrial myocytes with the currents produced by homo- meric Kv7. 1 and heteromeric Kv7. 1/MinK channels in Chinese ham- ster ovary cells indicates that activation kinetics and pharmacological properties of the IKs are similar to those of the homomeric Kv7.1 channels. Consistently with electrophysiological properties and homomeric Kv7.l channel composition, atrial transcript expression of the MinK subunit is only 1.6-1.9% of the expression level of the Kv7. 1 subunit. Since activation kinetics of the homomeric Kv7. 1 channels is much faster than activation of the heteromeric Kv7. 1/ MinK channels, the homomeric Kv7. 1 composition of the crucian carp cardiac IKs is thermally adaptive: the slow delayed rectifier channels can open despite low body temperatures and curtail the duration of cardiac action potential in ectothermic crucian carp. We suggest that the homomeric Kv7.l channel assembly is an evolutionary thermal adaptation of ectothermic hearts and the heteromeric Kv7.1/MinK channels evolved later to adapt IKs to high body temperature of endotherms. [ABSTRACT FROM AUTHOR]

Electrical activity of the heart is assumed to be one of the key factors that set thermal tolerance limits for ectothermic vertebrates. Therefore, we hypothesized that in thermal acclimation--the duration of cardiac action potential and the repolarizing K+ currents that regulate action potential duration (APD)--the rapid component of the delayed rectifier K+ current (IKl) and the inward rectifier K+ current (IKr) are more plastic in eurythermal than in stenothermal fish species. The hypothesis was tested in six freshwater teleosts representing four different lish orders (Cadiformes, Cypriniformes, Perciformes, Salmoniformes) acclimated at +4°C (cold acclimation) or +18°C (warm acclimation). In cold acclimation, a compensatory shortening of APD occurred in all species regardless of thermal tolerances, life styles, or phylogenies of the fish, suggesting that this response is a common characteristic of the teleost heart. The strength of the response did not, however, obey simple eurythermy-stenothermy gradation but differed among the phylogenetic groups. Salmoniformes fish showed the greatest acclimation capacity of cardiac electrical activity, whereas the weakest response appeared in the perch (Perciformes) heart. The underlying ionic mechanisms were also partly phylogeny dependent. Modification of the IKr current was almost ubiquitously involved in acclimation response of fish cardiac myocytes to temperature, while the ability to change the IKl current under chronic thermal stress was absent or showed inverse compensation in Salmoniformes species. Thus, in Salmoniformes fish, the thermal plasticity of APD is strongly based on IKr, while other fish groups rely on both IKr and IKl. [ABSTRACT FROM AUTHOR]

The hypothesis of pacemaker level origin of thermal compensation in heart rate was tested by recording action potentials (AP) in intact sinoatrial tissue and enzymatically isolated pacemaker cells of rainbow trout acclimated at 4°C (cold) and 18°C (warm). With electrophysiological recordings, the primary pacemaker was located at the base of the sinoatrial valve, where a morphologically distinct ring of tissue comprising myocytes and neural elements was found by histological examination. Intrinsic beating rate of this pacemaker was higher in cold-acclimated (46 ± 6 APs/min) than warm-acclimated trout (38 ± 3 APs/min; P < 0.05), and a similar difference was seen in beating rate of isolated pacemaker cells (44 ± 6 vs. 38 ± 6 APs/min; P < 0.05), supporting the hypothesis that thermal acclimation modifies the intrinsic pacemaker mechanism of fish heart. Inhibition of sarcoplasmic reticulum (SR) with 10 μM ryanodine and 1 μM thapsigargin did not affect heart rate in either warm- or cold- acclimated trout at 11°C but reduced heart rate in warm-acclimated trout from 74 ± 2 to 42 ± 6 APs/min (P < 0.05) at 18°C. At 11°C, a half-maximal blockade of the delayed rectifier K+ current (IKr) with 0.1 μM E-4031 reduced heart rate more in warm-acclimated (from 45 ± 1 to 24 ± 5 APs/min) than cold-acclimated trout (56 ± 3 vs. 48 ± 2 APs/min), whereas IKr density was higher and AP duration less in cold-acclimated trout (P > 0.05). Collectively, these findings suggest that a cold-induced increase in AP discharge frequency is at least partly due to higher density of the IKr in the cold-acclimated trout, whereas contribution of SR Ca2+ release to thermal compensation of heart rate is negligible. [ABSTRACT FROM AUTHOR]

The present study examines the importance of the Na+ current (INa) in the excitability of atrial and ventricular myocardium of the rainbow trout heart. Whole-cell patch- clamp under reduced sarcolemmal Na+ gradient showed that the density of INa is similar in atrial and ventricular myocytes of the trout heart, and the same result was obtained when INa was elicited by chamber-specific action potentials (AP) in normal physiological saline solution. However, the maximum rate (Vmax) of AP upstroke, measured with microelectrodes in intact trout heart, was 21% larger in atrium than ventricle, and thus in variance with the similar INa density of the two myocyte types. Furthermore, Vmax calculated from the INa was 2.1 and 3.2 times larger for atrium and ventricle, respectively, than the values obtained from the APs. The discrepancy between INa of isolated myocytes and Vmax of intact muscle is only partly explained by the inward rectifier K+ current (IK1), which overlaps INa and decreases the net depolarising current. Clear differences exist in the voltage dependence of steady-state activation and inactivation as well as in the inactivation kinetics of INa between atrial and ventricular myocytes. As a result of a more negative voltage dependence of INa activation, smaller IK1 and higher input resistance of atrial myocytes, the voltage threshold for AP generation is more negative in atrium than ventricle of the trout heart. These findings suggest that atrial muscle is more readily excitable than ventricular muscle, and this difference is partly due to the properties of the atrial INa. [ABSTRACT FROM AUTHOR]