A method to measure myocardial calcium handling in adult Drosophila.
A method to measure myocardial calcium handling in adult Drosophila.
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DOI：
10.1161/circresaha.110.238105
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发表时间：
2011-05-27
影响因子：
20.1
通讯作者：
Wolf MJ
中科院分区：
文献类型：
--
作者：
Lin N;Badie N;Yu L;Abraham D;Cheng H;Bursac N;Rockman HA;Wolf MJ
Normal cardiac physiology requires highly regulated cytosolic Ca2+ concentrations and abnormalities in Ca2+ handling are associated with heart failure. The majority of approaches to identify the components that regulate intracellular Ca2+ dynamics rely on cells in culture, mouse models, and human samples. However, a genetically robust system for unbiased screens of mutations that affect Ca2+ handling remains a challenge. We sought to develop a new method to measure myocardial Ca2+ cycling in adult Drosophila and determine whether cardiomyopathic fly hearts recapitulate aspects of diseased mammalian myocardium. Using engineered transgenic Drosophila that have cardiac-specific expression of Ca2+ sensing fluorescent protein, GCaMP2, we developed methods to measure parameters associated with myocardial Ca2+ handling. The following key observations were identified: (1) Control w1118 Drosophila hearts have readily measureable Ca2+-dependent fluorescent signals that are dependent on L-type Ca2+ channels and SR Ca2+ stores and originate from rostral and caudal pacemakers; (2) A fly mutant, held-up2 (hdp2) that has a point mutation in Troponin I and has a dilated cardiomyopathic phenotype demonstrates abnormalities in myocardial Ca2+ handling that include increases in the duration of the 50% rise in intensity to peak intensity, the half-time of fluorescence decline from peak, the full duration at half maximal intensity (FDHM) and decreases in the linear slope of decay from 80% to 20% intensity decay; and (3) Hearts from hdp2 mutants had reductions in caffeine-induced Ca2+ increases and reductions in ryanodine receptor (RyR) without changes in L-type Ca2+ channel transcripts compared to w1118. Our results show that the cardiac-specific expression of GCaMP2 provides a means to characterize propagating Ca2+ transients in adult fly hearts. Moreover, the adult fruit fly heart recapitulates several aspects of Ca2+ regulation observed in mammalian myocardium. A mutation in Drosophila that causes an enlarged cardiac chamber and impaired contractile function is associated with abnormalities in the cytosolic Ca2+ transient as well as changes in transcript levels of proteins associated with Ca2+ handling. This new methodology has the potential to permit an examination of evolutionarily conserved myocardial Ca2+ handing mechanisms by applying the vast resources available in the fly genomics community to conduct genetic screens to identify new genes involved in generated Ca2+ transients and arrhythmias.
