Fig. 3: In vivo optical recording of mild press (100 g)-induced neuronal activity in intact DRG neurons.

Optical voltage recordings of primary sensory neurons in response to a single mechanical force (100 g) applied to the hindpaw of naïve (a), CFA (b), or SN-CCI (c) animals. Insets, images of DRG cell bodies (white dotted lines) expressing ASAP4.4-Kv. Yellow lines indicate 1.1 kHz line scan regions where ASAP4.4-Kv optical recording signals were acquired. Each trace is the response of a single DRG neuron; (right) expanded view of the boxed region. Purple bars indicate the timing of the stimulus application. Representative line scan images in b are shown under the trace. Red arrow marks one of action potentials and red box indicates action potential burst. Scale bar: 5 μm. d Mean area under the curve (AUC) of ASAP4.4-Kv z-score signals (each dot represents an individual recorded neuron; number of cells = 4 from three biologically independent mice per group; Kruskal-Wallis test with Dunn’s post-hoc test) and GCaMP3 Ca²⁺ transients (violin plot with median (solid line) and quartiles (dashed lines); number of cells = 33 and 56 for naïve and CFA groups, respectively, from three biologically independent mice per group; two-tailed Mann-Whitney U-test) in L5 DRG neurons in response to 100 g press, and the number of total activated neurons (each dot represents one animal; number of biologically independent mice = 5 per group; two-tailed Mann-Whitney U-test). e In vivo intact DRG Ca²⁺ imaging from naïve or CFA-treated Pirt-GCaMP3 mice. (left) Averaged images before application of mild press (100 g). (right) Averaged images after application of mild press (100 g). White arrowheads indicate spontaneously activated neurons in the absence of applied stimulus. Yellow arrowheads indicate DRG neurons activated by mild press. Scale bar: 100 μm. Ca²⁺ transients from DRG neurons in response to mild press (100 g) applied to the hindpaw using Pirt-GCaMP3 mice (grey), from one (f) or three (g) animals for each treatment. Traces of averaged Ca²⁺ transients from each group are shown in black or orange. All data are presented as mean values ± SEM.