Ca2+ alternans (Ca-Alts) are alternating beat-to-beat changes in the amplitude of Ca2+ transients that frequently occur during tachycardia, ischemia, or hypothermia that can lead to sudden cardiac death. Ca-Alts appear to result from a variation in the amount of Ca2+ released from the sarcoplasmic reticulum (SR) between two consecutive heartbeats. This variable Ca2+ release has been attributed to the alternation of the action potential duration, delay in the recovery from inactivation of RYR Ca2+ release channel (RYR2), or an incomplete Ca2+ refilling of the SR. In all three cases, the RYR2 mobilizes less Ca2+ from the SR in an alternating manner, thereby generating an alternating profile of the Ca2+ transients. We used a new experimental approach, fluorescence local field optical mapping (FLOM), to record at the epicardial layer of an intact heart with subcellular resolution. In conjunction with a local cold finger, a series of images were recorded within an area where the local cooling induced a temperature gradient. Ca-Alts were larger in colder regions and occurred without changes in action potential duration. Analysis of the change in the enthalpy and Q10 of several kinetic processes defining intracellular Ca2+ dynamics indicated that the effects of temperature change on the relaxation of intracellular Ca2+ transients involved both passive and active mechanisms. The steep temperature dependency of Ca-Alts during tachycardia suggests Ca-Alts are generated by insufficient SERCA-mediated Ca2+ uptake into the SR. We found that Ca-Alts are heavily dependent on intra-SR Ca2+ and can be promoted through partial pharmacologic inhibition of SERCA2a. Finally, the FLOM experimental approach has the potential to help us understand how arrhythmogenesis correlates with the spatial distribution of metabolically impaired myocytes along the myocardium.
Cardiac physiology of fish models is an emerging field given the ease of genome editing and the development of transgenic models. Several studies have described the cardiac properties of zebrafish ( Denio rerio ). The goldfish ( Carassius auratus ) belongs to the same family as the zebrafish and has emerged as an alternative model with which to study cardiac function. Here, we propose to acutely study electrophysiological and systolic Ca 2+ signaling in intact goldfish hearts. We assessed the Ca 2+ dynamics and the electrophysiological cardiac function of goldfish, zebrafish, and mice models, using pulsed local field fluorescence microscopy, intracellular microelectrodes, and flash photolysis in perfused hearts. We observed goldfish ventricular action potentials (APs) and Ca 2+ transients to be significantly longer when compared to the zebrafish. The action potential half duration at 50% (APD 50 ) of goldfish was 370.38 ± 8.8 ms long, and in the zebrafish they were observed to be only 83.9 ± 9.4 ms. Additionally, the half duration of the Ca 2+ transients was also longer for goldfish (402.1 ± 4.4 ms) compared to the zebrafish (99.1 ± 2.7 ms). Also, blocking of the L-type Ca 2+ channels with nifedipine revealed this current has a major role in defining the amplitude and the duration of goldfish Ca 2+ transients. Interestingly, nifedipine flash photolysis experiments in the intact heart identified whether or not the decrease in the amplitude of Ca 2+ transients was due to shorter APs. Moreover, an increase in temperature and heart rate had a strong shortening effect on the AP and Ca 2+ transients of goldfish hearts. Furthermore, ryanodine (Ry) and thapsigargin (Tg) significantly reduced the amplitude of the Ca 2+ transients, induced a prolongation in the APs, and altogether exhibited the degree to which the Ca 2+ release from the sarcoplasmic reticulum contributed to the Ca 2+ transients. We conclude that the electrophysiological properties and Ca 2+ signaling in intact goldfish hearts strongly resembles the endocardial layer of larger mammals.
Bazmi and Escobar highlight a recent investigation of the mechanisms that regulate Ca2+ influx during sympathetic stimulation.
During production of this article, we inadvertently omitted an image from the Supplemental text. The beginning of the Supplemental text should have appeared as below. The error only exists in PDFs downloaded before March 9, 2021.
Autonomic regulation plays a central role in cardiac contractility and excitability in numerous vertebrate species. However, the role of autonomic regulation is less understood in fish physiology. Here, we used Goldfish as a model to explore the role of autonomic regulation. A transmural electrocardiogram recording showed perfusion of the Goldfish heart with isoproterenol increased the spontaneous heart rate, while perfusion with carbamylcholine decreased the spontaneous heart rate. Cardiac action potentials obtained via sharp microelectrodes exhibited the same modifications of the spontaneous heart rate in response to isoproterenol and carbamylcholine. Interestingly, the duration of the cardiac action potentials lengthened in the presence of both isoproterenol and carbamylcholine. To evaluate cardiac contractility, the Goldfish heart was perfused with the Ca2+ indicator Rhod-2 and ventricular epicardial Ca2+ transients were measured using Pulsed Local Field Fluorescence Microscopy. Following isoproterenol perfusion, the amplitude of the Ca2+ transient significantly increased, the half duration of the Ca2+ transient shortened, and there was an observable increase in the velocity of the rise time and fall time of the Ca2+ transient, all of which are compatible with the shortening of the action potential induced by isoproterenol perfusion. On the other hand, carbamylcholine perfusion significantly reduced the amplitude of the Ca2+ transient and increased the half duration of the Ca2+ transient. These results are interesting because the effect of carbamylcholine is opposite to what happens in classically used models, such as mouse hearts, and the autonomic regulation of the Goldfish heart is strikingly similar to what has been observed in larger mammalian models resembling humans.
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