Fig. 7: LI-cTBUS induces mechanical analgesia through astrocytic TRPA1 activation. | Nature Communications

Fig. 7: LI-cTBUS induces mechanical analgesia through astrocytic TRPA1 activation.

From: Harnessing theta-gamma coupled brainwaves using ultrasound for spinal astrocyte revitalization and sustained neuropathic pain relief in mice

Fig. 7

a Top: Schematic illustrating conditional astrocytic TRPA1 knockdown using Gfap-CreERT2 mice and ex vivo calcium imaging in spinal cord slices, created in BioRender; Sung, Y. (2025) https://BioRender.com/1rwu7p9. Bottom: Representative confocal images showing Cre-dependent mCherry expression (red) and astrocytic GCaMP6s (green) expression in the dorsal horn, confirming astrocyte-specific viral targeting. Scale bar: 100 μm. b Top: Calcium transients (ΔF/F₀) from spinal astrocytes from PCI mice expressing scrambled shRNA show robust responses to single-pulse ultrasound (100, 200, 400 ms; pink-shaded bars) and to LI-cTBUS (40 s). Bottom: Astrocytes from Trpa1 knockdown mice show no detectable calcium response to either stimulation paradigm. Traces represent averaged ΔF/F₀ from defined regions of interest (ROIs). c Pharmacological inhibition of TRPA1 with HC030031 in WT mice abolished LI-cTBUS-induced astrocytic calcium elevations. Data are presented as mean ± SEM box-and-whisker plots, n = 5 (LI-cTBUS), n = 7 (LI-cTBUS + HC030031), *p = 0.031, Welch’s t-test. d Behavioral assessment of mechanical withdrawal thresholds in astrocytic Trpa1 knockdown mice treated with LI-cTBUS. LI-cTBUS failed to elicit analgesic responses in knockdown animals, confirming the functional requirement of astrocytic TRPA1. Data are presented as mean ± SEM and box-and-whisker plots, *p = 0.04, N = 3, one-way ANOVA with Tukey’s multiple comparison test. N number of mice, n number of recorded cells.

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