Fig. 6: NPF-releasing neurons convey internal state information to the feedback inhibitory neuron. | Nature Communications

Fig. 6: NPF-releasing neurons convey internal state information to the feedback inhibitory neuron.

From: Feeding state-dependent neuropeptidergic modulation of reciprocally interconnected inhibitory neurons biases sensorimotor decisions in Drosophila

Fig. 6

a, b Comparison of light microscopy images (GMR_SS01635 > UAS-GFP) with electron microscopy (EM) reconstruction images of DM-NPF neurons. c EM image shows neuropeptide-containing dense core vesicles in the DM-NPF neuron near one of its synapses with Handle-b. Red arrows: presynaptic neurons, DM-NPF annotated with yellow arrow, blue arrows: postsynaptic sites, magenta arrow: Handle-b (d−f) Baseline calcium fluorescence in DM-NPF ventral nerve cord projections upon 90 min sucrose feeding (d), starvation for 90 min (e) or 5 hours (f). g, h Locomotion analysis. Box plots display the median, interquartile range (IQR), and whiskers up to 1.5 × IQR. Outliers are plotted individually. g control larvae. h NPF knockdown in DM-NPF neurons. i−k Handle-b calcium responses with (UAS-GCaMP6s; 22E09-Gal4, NPF-LexA > LexAop-KIR) or without (UAS-GCaMP6s; 22E09-Gal4, NPF-LexA;+) DM-NPF silencing in fed larvae (i) Handle-B calcium responses, single trial of mechanosensory stimulation. j mean Handle-B calcium trace over time +/- SEM. k mean calcium response averaged during the stimulus. l−p Immunohistochemical labeling for NPFR in Handle-b (all images, scale bar = 5 µm). GMR_SS00888 split-Gal4 drives UAS-GCaMP6s in Handle-b, and LexAop-jRGECO1a is expressed under the control of the NPFR promoter using a T2A-LexA construct (magenta). GFP and dsRed antibodies increase detection sensitivity. l 4 neurons (N1-N4), 23 µm stack. The (m−p) whole thickness (5 to 7 µm) of each neuron (N1-N4) is shown. q−s Handle-B calcium responses upon NPFR knockdown (GMR_SS00888 > UAS-NPFR-RNAi; UAS-GCaMP6s) compared to control (GMR_SS00888 > UAS-GCaMP6s). q mean Handle-b calcium trace over time +/- SEM in control larvae fed, fed on sucrose or starved. r mean Handle-b calcium trace over time +/- SEM upon NPFR knockdown in larvae fed, fed on sucrose or starved. s mean calcium response averaged during the stimulus. f, k, s White line: mean, white dot: the median, colored dots: individual data points. j, q, r green dashed line: stimulus. (Statistics: d−f, k two-tailed T-test; g, h two-sided Mann-Whitney test with Bonferroni correction; s one-sided Mann-Whitney test; ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05). See also Supplementary Figs. 7 and 8. The source data and p-values are provided in Source Data 1, 5 and 6.

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