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      Development of a rapid and economic in vivo electrocardiogram platform for cardiovascular drug assay and electrophysiology research in adult zebrafish

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          Abstract

          Zebrafish is a popular and favorable model organism for cardiovascular research, with an increasing number of studies implementing functional assays in the adult stage. For example, the application of electrocardiography (ECG) in adult zebrafish has emerged as an important tool for cardiac pathophysiology, toxicity, and chemical screen studies. However, few laboratories are able to perform such functional analyses due to the high cost and limited availability of a convenient in vivo ECG recording system. In this study, an inexpensive ECG recording platform and operation protocol that has been optimized for adult zebrafish ECG research was developed. The core hardware includes integration of a ready-to-use portable ECG kit with a set of custom-made needle electrode probes. A combined anesthetic formula of MS-222 and isoflurane was first tested to determine the optimal assay conditions to minimize the interference to zebrafish cardiac physiology under sedation. For demonstration, we treated wild-type zebrafish with different pharmacological agents known to affect cardiac rhythms in humans. Conserved electrophysiological responses to these drugs were induced in adult zebrafish and recorded in real time. This economic ECG platform has the potential to facilitate teaching and training in cardiac electrophysiology with adult zebrafish and to promote future translational applications in cardiovascular medicine.

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          International Conference on Harmonisation; guidance on E14 Clinical Evaluation of QT/QTc Interval Prolongation and Proarrhythmic Potential for Non-Antiarrhythmic Drugs; availability. Notice.

          (2005)
          The Food and Drug Administration (FDA) is announcing the availability of a guidance entitled "E14 Clinical Evaluation of QT/QTc Interval Prolongation and Proarrhythmic Potential for Non-Antiarrhythmic Drugs." The guidance was prepared under the auspices of the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH). The guidance provides recommendations to sponsors concerning clinical studies to assess the potential of a new drug to cause cardiac arrhythmias, focusing on the assessment of changes in the QT/QTc interval on the electrocardiogram as a predictor of risk. The guidance is intended to encourage the assessment of drug effects on the QT/QTc interval as a standard part of drug development and to encourage the early discussion of this assessment with FDA.
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            Real time electrocardiogram QRS detection using combined adaptive threshold

            Background QRS and ventricular beat detection is a basic procedure for electrocardiogram (ECG) processing and analysis. Large variety of methods have been proposed and used, featuring high percentages of correct detection. Nevertheless, the problem remains open especially with respect to higher detection accuracy in noisy ECGs Methods A real-time detection method is proposed, based on comparison between absolute values of summed differentiated electrocardiograms of one of more ECG leads and adaptive threshold. The threshold combines three parameters: an adaptive slew-rate value, a second value which rises when high-frequency noise occurs, and a third one intended to avoid missing of low amplitude beats. Two algorithms were developed: Algorithm 1 detects at the current beat and Algorithm 2 has an RR interval analysis component in addition. The algorithms are self-adjusting to the thresholds and weighting constants, regardless of resolution and sampling frequency used. They operate with any number L of ECG leads, self-synchronize to QRS or beat slopes and adapt to beat-to-beat intervals. Results The algorithms were tested by an independent expert, thus excluding possible author's influence, using all 48 full-length ECG records of the MIT-BIH arrhythmia database. The results were: sensitivity Se = 99.69 % and specificity Sp = 99.65 % for Algorithm 1 and Se = 99.74 % and Sp = 99.65 % for Algorithm 2. Conclusion The statistical indices are higher than, or comparable to those, cited in the scientific literature.
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              Combined use of MS-222 (tricaine) and isoflurane extends anesthesia time and minimizes cardiac rhythm side effects in adult zebrafish.

              As an important vertebrate model organism, zebrafish are typically studied at the embryonic stage to take advantage of their properties of transparency and rapid development. However, more and more studies require assays to be done on adults. Consequently, a good anesthetic is needed to sedate and immobilize the adult zebrafish during experimental manipulation. To date, MS-222 (tricaine methanesulfonate) is the only Food and Drug Administration approved anesthetic for aquaculture and is widely used by the zebrafish research community. Nevertheless, in adult zebrafish, MS-222 reduces heart rate and causes high mortality under long-term sedation. Consequently, adult zebrafish have limited research applications. In this study, we present a new anesthetic formula for the adult zebrafish that results in minimal side effects on its physiology under prolonged sedation. The combined use of MS-222 with isoflurane effectively extended the time of anesthesia, and the zebrafish recovered faster than when anesthetized with the traditional MS-222. Moreover, MS-222 + isoflurane did not cause reduction of heart rates, which enabled long-term electrocardiogram recording and microscopic observation on the adult zebrafish. Taken together, the new MS-222 + isoflurane formula will facilitate general applications of adult zebrafish in time-consuming experiments with minimal side effects on the model organism's overall physiology.
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                Author and article information

                Contributors
                drlawrenceliu@gmail.com
                yjchuang@life.nthu.edu.tw
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                30 October 2018
                30 October 2018
                2018
                : 8
                : 15986
                Affiliations
                [1 ]ISNI 0000 0004 0532 0580, GRID grid.38348.34, Department of Medical Science & Institute of Bioinformatics and Structural Biology, , National Tsing Hua University, ; Hsinchu, 30013 Taiwan
                [2 ]ISNI 0000 0004 0532 0580, GRID grid.38348.34, Department of Electrical Engineering, , National Tsing Hua University, ; Hsinchu, 30013 Taiwan
                [3 ]ISNI 0000 0004 0532 0580, GRID grid.38348.34, Department of Medical Science, , National Tsing Hua University, ; Hsinchu, 30013 Taiwan
                [4 ]ISNI 0000 0000 9476 5696, GRID grid.412019.f, Department of Biotechnology, College of Life Science, , Kaohsiung Medical University, ; Kaohsiung, 80708 Taiwan
                [5 ]ISNI 0000 0004 0573 007X, GRID grid.413593.9, Division of Cardiology, Department of Internal Medicine, , Hsinchu Mackay Memorial Hospital, ; Hsinchu, 30071 Taiwan
                Author information
                http://orcid.org/0000-0002-6022-0107
                Article
                33577
                10.1038/s41598-018-33577-7
                6207748
                30375400
                fc2523f8-5a07-4663-bf0e-4d756faca35e
                © The Author(s) 2018

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 14 May 2018
                : 27 September 2018
                Funding
                Funded by: FundRef https://doi.org/10.13039/501100004663, Ministry of Science and Technology, Taiwan (Ministry of Science and Technology of Taiwan);
                Award ID: 106-2311-B-007-009-MY3
                Award ID: 106-2311-B-007-009-MY3
                Award ID: 106-2311-B-007-009-MY3
                Award ID: 106-2311-B-007-009-MY3
                Award ID: 106-2311-B-007-009-MY3
                Award ID: 106-2311-B-007-009-MY3
                Award ID: 101-103-2325-B-007
                Award Recipient :
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