Cardioluminescence in transgenic zebrafish embryos: a calcium imaging tool to study drug effects and pathological modeling
bioRxiv (Cold Spring Harbor Laboratory)(2021)
摘要
Abstract The zebrafish embryo has emerged as an excellent model in cardiovascular research. The existing techniques to monitor Ca 2+ in the heart based on fluorescent Ca 2+ biosensors are limited due to phototoxicity and photobleaching. To overcome these issues, we have used bioluminescence. We generated a transgenic line expressing GFP-Aequorin in the heart, Tg ( cmlc2:GA ), and optimized an in vivo aequorin reconstitution protocol to improve the luminescence capacity. This allowed imaging Ca 2+ in long duration recordings in embryos of 3 to 5 days post-fertilization. The analogs diacetyl h -coelenterazine and f -coelenterazine enhanced the light output and signal-to-noise ratio from the embryos. With this cardioluminescence model, we monitored the time-averaged Ca 2+ levels and beat-to-beat Ca 2+ oscillations. Changes in Ca 2+ levels were observed by incubation with BayK8644, an L-type Ca 2+ channel agonist, the channel blocker nifedipine, and β-adrenergic blocker propranolol. Treatment of zebrafish embryos with terfenadine for 24 hours has been proposed as a model of heart failure. Tg ( cmlc2:GA ) embryos treated with terfenadine showed a 2:1 atrioventricular block and a decrease in the ventricular Ca 2+ levels.
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关键词
transgenic zebrafish embryos,calcium imaging tool
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