Figure 7

The dural afferent neuron population lacks TRPM8-expressing neurons. (A) Representative images of TG sections from TRPM8^EGFPf/+ mice following dural DiI application or intradermal DiI injection at the whisker pad. The thick and thin arrowheads indicate DiI⁺ and EGFPf⁺ neurons, respectively. The arrows in the lower row indicate a DiI⁺ skin afferent neuron that is also EGFPf⁺. (B) The percentage of EGFPf⁺ neurons that are in the V₁/V₂ TG, the V₃ TG and the DiI⁺ dural afferent neuron population (n = 3 mice, on average, 480 V₁/V₂ neurons, 472 V₃, neurons and 206 DiI⁺ neurons were counted from each mouse; *** p < 0.001, two-tailed t-test, V₁/V₂ group versus DiI⁺ dura group). (C) The fraction of EGFPf⁺ neurons in the three TG divisions (n = 4 mice; on average, 1210 EGFPf⁺ neurons were counted from each mouse; *** p < 0.001, one-way ANOVA with post hoc Bonferroni test, all compared with the V₃ distribution). (D) The percentage of EGFPf⁺ V₁/V₂ TG neurons and DiI⁺ neurons innervating the skin over the whisker pad (n = 4 mice, on average, 448 V₁/V₂ neurons and 76 DiI⁺ neurons were counted from each mouse). (E) The sizes of EGFPf⁺ neurons (n = 363 neurons pooled from five mice) are significantly smaller than those of the neurons in the V₁/V₂ divisions of the TG (n = 2403 neurons pooled from 5 mice, p < 0.001, Mann–Whitney U test). (F) Cumulative distributions of the cross-sectional areas of the total TG neuron populations in the V₁/V₂ divisions (the same neurons as in E), the EGFPf⁻ V₁/V₂ TG neurons (n = 2040 neurons pooled from five mice), and the FG⁺ dural afferent neurons (the same neurons as in Figure 1F). The sizes of EGFPf⁻ V₁/V₂ TG neurons are comparable to those of the dural afferent neurons, and both populations are significantly larger than the total V₁/V₂ neurons (p < 0.001, Kruskal-Wallis ANOVA with Dunn’s post hoc test). (G) Nearest-neighbor measurement shows that both DiI⁺ dural afferent neurons and EGFPf⁺ neurons are randomly distributed in the TG (n = 182 EGFPf⁺ neurons and 619 DiI⁺ dural afferent neurons from three mice, the same neurons as in B). (H) A modified nearest-neighbor measurement shows that the EGFPf⁺ neurons are located farther away from the dural afferent population than would be predicted by random association, and vice versa (the same neurons as in G).