A Method to Measure Myocardial Calcium Handling in Adult Drosophila
Rationale: Normal cardiac physiology requires highly regulated cytosolic Ca2+ concentrations, and abnormalities in Ca2+ handling are associated with heart failure. The majority of approaches to identifying 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.
Objective: 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.
Methods and Results: 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, and decreases in the linear slope of decay from 80% to 20% intensity decay. (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 in comparison with w1118.
Conclusions: Our results show that the cardiac-specific expression of GCaMP2 provides a means of characterizing 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.
- Received December 1, 2010.
- Revision received March 30, 2011.
- Accepted April 1, 2011.
- © 2011 American Heart Association, Inc.