Extended Data Fig. 11: 0.5 mA ES at cutaneous or traditional non-acupoint regions failed to suppress inflammation, but ES at the forelimb LI10 acupoint can evoke PROKR2ADV neuron-dependent anti-inflammatory effects.
From: A neuroanatomical basis for electroacupuncture to drive the vagal–adrenal axis
a, b, low-intensity ES at the superficial ST36 region. Schematics (a) showing ES at the superficial, intradermal part of the ST36 region. Two electric needles were inserted through the epidermis (“Ep.”) and into the dermis (“De.”) at ST36 regions, with needles tilted to restrict them within the superficial dermis. This intradermal 0.5-mA ES failed to reduce LPS-induced TNF-α and IL-6 expression compared with sham 0 mA ES in C57BL/6J mice (b, n = 5 mice per group; two-side student’s unpaired t-test; for TNF-α: t8 = 0.218; NS, not significant, P = 0.833; for IL-6: t8 = 0.562; NS, P = 0.589). c, d, Low-intensity stimulation of the sural nerve. Schematics (c) showing ES at the middle region of the posterior hind leg, by inserting electric needles through the Chengjin (BL56) acupoint, with tips flanking the skin-innervating sural nerve (“Sn”). 0.5 mA ES at this acupoint failed to reduce TNF-α and IL-6 compared with sham 0 mA ES in C57BL/6J mice (d, n = 5 mice per group; two-side student’s unpaired t-test; for TNF-α: t8 = 0.375; NS, P = 0.718; for IL-6: t8 = 0.721; NS, P = 0.491). e-j, Low-intensity ES within the gastrocnemius (“G.”) muscle or the semitendinosus (“S.”) muscles. Schematics (e, h) showing ES at these two muscles. Two representative images (f, i) showing sparse innervation by PROKR2ADV-tdTomato+ fibers within these muscles, with the percentages of unit areas showing positive fibers (0.51 ± 0.19% for G. and 0.60 ± 0.11% for S. muscles) comparable to that seen in the outer TA muscle (0.51 ± 0.15%) and the abdominal wall muscles (0.59 ± 0.12%) shown in Extended Data Fig. 4h. n = 4 mice. In comparison with sham 0 mA ES, no impact on TNF-α and IL-6 production by 0.5 mA ES in either G. muscle (g, n = 5 mice per group. two-side student’s unpaired t-test; for TNF-α: t8 = 0.205; NS, P = 0.843; for IL-6: t8 = 0.861; NS, P = 0.415) or in S. mucle (j, n = 5 mice per group; two-side student’s unpaired t-test; for TNF-α: t8 = 0.468; NS, P = 0.652; for IL-6: t8 = 0.593; NS, P = 0.570) in C57BL/6J mice. k-m, Low-intensity ES at the forelimb acupoint LI10 (Shousanli). Schematics (k) and the image (l, top) showing ES at the forelimb acupoint LI10. Representative images (l) showing PROKR2ADV-tdTomato+ fibers within the deep branch of the radial nerve, and their innervations in radius periosteum, which were prominent at transverse sections at levels slightly distal (e.g., 1 mm) from the LI10 acupoint level. n = 3 mice. (m) Loss of 0.5 mA LI10 ES-evoked reduction of TNF-α and IL-6 in PROKR2ADV-Abl mice compared with control mice (two-way ANOVA, n = 5 mice per group; for TNF-α: F1, 16 = 20.384, P < 0.001; post-hoc Tukey test: *P = 0.011; NS, P = 0.562; for IL-6: F1, 16 = 14.296, P = 0.002; post-hoc Tukey test: **P = 0.004; NS, P = 0.728). (n) Loss of 0.5 mA LI10 ES-evoked reduction of TNF-α and IL-6 in mice with subdiaphragmatic vagotomy (“sVX”) compared with mice with sham surgery (two-way ANOVA, n = 5 mice per group; for TNF-α: F1, 16 = 22.875, P < 0.001; post-hoc Tukey test: **P = 0.004; NS, P = 0.697; for IL-6: F1, 16 = 18.065, P = 0.002; post-hoc Tukey test: **P = 0.004; NS, P = 0.57). TA: Anterior tibial muscle; T.: Tibia; F.: Fibula; Fe.: Femur; R.: Radius; U.: Ulna. Data are shown as mean ± SEM. Scale bars: 100 μm.
