Figure 1

D. melanogaster exhibits negative staircase. (a) Semi intact preparation of a fly harboring the reporter system handC-GFP. (b) An amplified fluorescence image showing the fluorescent cardiomyocytes and pericardial cells. The arrow indicates the cell displacement trajectory. (c) Raw (above) and digitized (below) cell movements. (d) Semi intact preparation of a fly harboring the reporter system GCaMP3. (e) Images of intracellular Ca²⁺ transients tracked in the conical chamber. The recording represents an arrhythmic heart. (f) Amplified Ca²⁺ transient to indicate the different parameters analyzed. (g) Representative recording of mechanical activity showing lateral displacements of a single cell after three successive 0.5 Hz increments in stimulation frequency (ΔFr1, ΔFr2, ΔFr3) from basal (B) spontaneous heart rate. Right panel: twitches obtained after digitalization of images using an algorithm based in phyton language. (h) Left panel: representative line scans of calcium transient after three successive 0.5 Hz increments in stimulation frequency. Right panel: Digitized images of the respective line scans. (i) Quantification of mechanical activity (black symbols) and Ca²⁺ transients (white symbols), reveals the negative staircase in D. melanogaster. N = 20, 19, 13, 5 for mechanical activity data at basal, ΔFr1, ΔFr2, and Δ Fr3 respectively. N = 10, 10, 7, 4 for Ca²⁺ transients data at basal, ΔFr1, ΔFr2, and ΔFr3 respectively. (j) D. melanogaster exhibit significant FDAR only at the highest frequency. Quantification of half relaxation time of cell movement and Ca²⁺ transients. N = 11, 10, 5, 4 for mechanical activity data at basal, ΔFr1, ΔFr2, and ΔFr3 respectively. N = 9, 9, 8, 4 for Ca²⁺ transients data at basal, ΔFr1, ΔFr2, and ΔFr3 respectively. Two way ANOVA followed by Tukey’s test for multiple comparisons was used for statistical analysis.*, ^#P < 0.05.