Figure 2. Propagation of touch signals in the fly ventral nerve cord (VNC).
(A) Left: Schematic of bristle recording configuration. Right: Responses of a bristle neuron to mechanical (red) and optogenetic (green) stimuli. Signals are band-pass filtered to facilitate spike identification (see Supplemental Experimental Procedures). All bristle neuron recordings are made from the same bristle on the prothoracic leg, near the femur-tibia joint (Figure S5A).
(B) Projection of a coronal stack through a region (~180 μm x 180 μm) of the prothoracic VNC showing resting GCaMP6f fluorescence. GCaMP6f is expressed pan-neuronally and imaged with a two-photon microscope. Outlined in white dashed lines are the neuropil regions (neuromeres); these regions do not contain neuronal cell bodies.
(C) Schematic of optogenetic bristle stimulation for calcium imaging experiments. The fly is positioned ventral side up. Light is directed at the femur/tibia joint of a prothoracic leg. The imaged region of the VNC is outlined in red.
(D) GCaMP signals recorded during periodic optogenetic stimulation of leg bristles. The left and right panels show color-coded ΔF/F responses of example neurons from a single imaging plane, illustrated in the center panel. Cross-correlation values, computed between each neuron’s ΔF/F signal and the stimulus waveform, are indicated alongside each trace.
(E) Map of all 699 neurons in the prothoracic region of a typical VNC, with individual neurons color-coded by their correlation value. In this projection, neurons with higher correlation are displayed on top. Neurons with correlation values below the threshold for statistical significance (0.19) are blue (n.s.).
(F) Top: distribution of correlation values between calcium signals (ΔF/F) and the stimulus waveform for all 699 neurons. Bottom: correlation values after shuffling the stimulus waveform; the 95th percentile of this distribution was taken as the threshold for significance.
(G) Correlation map of VNC neurons (same as panel E) but excluding motor neurons. The arrow points to the cluster of neurons that we identified for further scrutiny.