期刊
BIOMEDICINES
卷 9, 期 10, 页码 -出版社
MDPI
DOI: 10.3390/biomedicines9101294
关键词
zebrafish; calcium; heart; aequorin; coelenterazine; heart failure; imaging
资金
- Ministry of Science, Innovation and Universities, Spain [BFU2015-69874-R, PID2019-111456RB-100]
- EU FEDER-ERDF
- Consejeria de Educacion, Cultura y Deportes, Junta de Comunidades de Castilla-La Mancha [SBPLY/19/180501/000223]
- University of Castilla-La Mancha (UCLM) [2019-GRIN-27019, 2020-GRIN-29186, 2021-GRIN-31151]
Zebrafish embryos and larvae are an excellent model for cardiovascular research, allowing live imaging with genetically encoded biosensors to study cardiac cell behaviors, including calcium dynamics. The GFP-aequorin transgenic line provides a powerful platform for understanding calcium dynamics in heart failure and arrhythmia.
Zebrafish embryos and larvae have emerged as an excellent model in cardiovascular research and are amenable to live imaging with genetically encoded biosensors to study cardiac cell behaviours, including calcium dynamics. To monitor calcium ion levels in three to five days post-fertilization larvae, we have used bioluminescence. We generated a transgenic line expressing GFP-aequorin in the heart, Tg(myl7:GA), and optimized a reconstitution protocol to boost aequorin luminescence. The analogue diacetyl h-coelenterazine enhanced light output and signal-to-noise ratio. With this cardioluminescence model, we imaged the time-averaged calcium levels and beat-to-beat calcium oscillations continuously for hours. As a proof-of-concept of the transgenic line, changes in ventricular calcium levels were observed by Bay K8644, an L-type calcium channel activator and with the blocker nifedipine. The beta-adrenergic blocker propranolol decreased calcium levels, heart rate, stroke volume, and cardiac output, suggesting that larvae have a basal adrenergic tone. Zebrafish larvae treated with terfenadine for 24 h have been proposed as a model of heart failure. Tg(myl7:GA) larvae treated with terfenadine showed bradycardia, 2:1 atrioventricular block, decreased time-averaged ventricular calcium levels but increased calcium transient amplitude, and reduced cardiac output. As alterations of calcium signalling are involved in the pathogenesis of heart failure and arrhythmia, the GFP-aequorin transgenic line provides a powerful platform for understanding calcium dynamics.
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