Fig. 1: Periodic Ca²⁺ waves are triggered in the fly larval fat body by a brain-derived factor.

A A 3rd instar larva was placed in a narrow glass channel to restrict its mobility. Fat body-specific Ca²⁺ activity was visualized using Lpp-GAl4 > UAS-GCaMP5G. A kymograph of Ca²⁺ waves traveling from head to tail is shown. The result was repeated in more than 5 independent experiments. B Representative Ca²⁺ activity in the larval fat body within the midsection. Ca²⁺ intensity was plotted in arbitrary units (a.u.). C Fat body expressing GCaMP5G was dissected and cultured in Schneider’s Drosophila medium with or without different larval organs. D Fat body cultured alone showed little Ca²⁺ activity, while fat body cultured together with dissected brains showed prominent ICWs. The dynamic Ca²⁺ activities are highlighted in the lower panel by subtracting the constant background signal and showing the organ outline. The result was repeated in 6 independent experiments. E Quantification of Ca²⁺ activities in the cultured fat body under different conditions. N = 6, 6, 6, 6 fat bodies (left to right). F Brain-conditioned medium, prepared by incubating 20 dissected larval brains of the third instar with 400 μl of Schneider’s medium for one hour, was utilized to treat the isolated fat bodies. G Brain-conditioned medium before and after filtration triggered significant ICWs in cultured fat bodies. N = 3, 3 fat bodies. H Ca²⁺ activity triggered by proteinase K-digested brain-conditioned medium was significantly reduced compared to untreated brain-conditioned medium. Brain-conditioned medium was treated with 0.1 mg/mL proteinase K, DNAse I (1 U/mL), or RNAse A (1 μg/mL) for 1 h at 37 °C and then filtered with Pierce 10 K MWCO concentrator. The untreated brain-conditioned medium was processed similarly without the addition of proteinase K. N = 3, 3, 3, 3 fat bodies. Ca²⁺ intensity was presented using a linear colour scale (minimum = 0, maximum = 255) (A, D). Ordinary one-way ANOVA with Dunnett’s multiple comparisons was utilized in (E, H). Two-tail unpaired Student’s t-test was utilized in (G). Data were plotted as mean ± SD. Scale bars, 500 μm (A), 500 μm (D). Source data are provided as a Source Data file.