Figure 2. Dual-Color SCAPE Imaging of Proprioceptor Activity Dynamics in Crawling Larvae.

(A and A′) SCAPE imaging of a 410-Gal4, 20XUAS-IVS-GCaMP6f (×2), UAS-CD4tdTomato larva, ventral side, during forward crawling. Images are MIP of a full 168-μm-deep imaging volume (square root grayscale). (A) shows a ventral view (x-y) and (A′) shows a side view (y-z). GCaMP signal was extracted from the segmentally repeated vpda neurons indicated by circles. See Video S2.

(B) vpda soma response for the neurons tracked in (A). The distance between the measured neuron and the posterior neuron (posterior inter-cell distance) is plotted in dashed lines, and the GCaMP6f response is plotted in solid lines (quantified as the fractional change in the green/red fluorescence ratio ΔR/R₀; see STAR Methods and Figure S1). The dotted vertical line refers to the time point shown in (A).

(C and C′) Representative tdTomato (C) and GCaMP6f (C′) images showing increased activity in dendrites (arrowhead) and axons (arrow) during contraction. Images are cropped to show region of interest and are MIP of a 70-μm-depth range from a 160-μm-deep volume (square root grayscale). See Video S3.

(D) Mean calcium response (solid line) ± SD (ribbon) of vpda soma (3 animals, n = 22 cells [8, 10, and 4], 26 events [9, 13, and 4], respectively) during segment contraction, represented by mean posterior inter-cell distance (dashed line) ± SD (ribbon). Maximal segment contraction is set at “t = 0 s” for each event. ΔR/R₀ amplitude was normalized for each event.

Posterior is to the left for all images. Scale bars, 100 μm.