title: A method to measure myocardial calcium handling in adult Drosophila.
abstract: BACKGROUND
  Normal cardiac physiology requires highly regulated cytosolic Ca(2+) concentrations and abnormalities in Ca(2+) handling are associated with heart failure. The majority of approaches to identifying the components that regulate intracellular Ca(2+) dynamics rely on cells in culture, mouse models, and human samples. However, a genetically robust system for unbiased screens of mutations that affect Ca(2+) handling remains a challenge.
  
  
  OBJECTIVE
  We sought to develop a new method to measure myocardial Ca(2+) cycling in adult Drosophila and determine whether cardiomyopathic fly hearts recapitulate aspects of diseased mammalian myocardium.
  
  
  RESULTS
  Using engineered transgenic Drosophila that have cardiac-specific expression of Ca(2+)-sensing fluorescent protein, GCaMP2, we developed methods to measure parameters associated with myocardial Ca(2+) handling. The following key observations were identified: (1) Control w(1118) Drosophila hearts have readily measureable Ca(2+)-dependent fluorescent signals that are dependent on L-type Ca(2+) channels and SR Ca(2+) stores and originate from rostral and caudal pacemakers. (2) A fly mutant, held-up(2) (hdp(2)), that has a point mutation in troponin I and has a dilated cardiomyopathic phenotype demonstrates abnormalities in myocardial Ca(2+) 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 hdp(2) mutants had reductions in caffeine-induced Ca(2+) increases and reductions in ryanodine receptor (RyR) without changes in L-type Ca(2+) channel transcripts in comparison with w(1118).
  
  
  CONCLUSIONS
  Our results show that the cardiac-specific expression of GCaMP2 provides a means of characterizing propagating Ca(2+) transients in adult fly hearts. Moreover, the adult fruit fly heart recapitulates several aspects of Ca(2+) 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 Ca(2+) transient as well as changes in transcript levels of proteins associated with Ca(2+) handling. This new methodology has the potential to permit an examination of evolutionarily conserved myocardial Ca(2+)-handing mechanisms by applying the vast resources available in the fly genomics community to conduct genetic screens to identify new genes involved in generated Ca(2+) transients and arrhythmias.
authors.affiliation: Institute of Molecular Medicine, Peking University, Beijing, China.
authors.forename: Na
authors.initials: N
authors.lastname: Lin
authors.forename: Nima
authors.initials: N
authors.lastname: Badie
authors.forename: Lin
authors.initials: L
authors.lastname: Yu
authors.forename: Dennis
authors.initials: D
authors.lastname: Abraham
authors.forename: Heping
authors.initials: H
authors.lastname: Cheng
authors.forename: Nenad
authors.initials: N
authors.lastname: Bursac
authors.forename: Howard A
authors.initials: HA
authors.lastname: Rockman
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Wolf
mesh_terms: D000367:Age Factors; D000818:Animals; D030801:Animals, Genetically Modified; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D009202:Cardiomyopathies / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D003600:Cytosol / Q000378:metabolism; D004195:Disease Models, Animal; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster; D005260:Female; D015870:Gene Expression / Q000502:physiology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009200:Myocardial Contraction / Q000502:physiology*; D009206:Myocardium / Q000378:metabolism
chemical_list: D020746:Calcium Channels, L-Type; D029721:Drosophila Proteins; D008565:Membrane Proteins; C472859:tinc protein, Drosophila; D049452:Green Fluorescent Proteins