Fig. 4: Functional CLR/RAMP1 is expressed by HSCs and undergoes clathrin- and dynamin-mediated endocytosis, which underlies nociception.
From: Schwann cell endosome CGRP signals elicit periorbital mechanical allodynia in mice
a, b Effects of graded concentrations of CGRP on cAMP formation (n = 5 independent experiments). c, d Effects of graded concentrations of olcegepant on CGRP (100 nM)-evoked cAMP formation (n = 4 independent experiments). e Pharmacological targets. f Representative images of HSCs expressing Rab5a-GFP at 30 min after incubation with TAMRA-CGRP (100 nM). Arrows denote colocalization of TAMRA-CGRP and Rab5a-GFP. Arrowheads denote retention of a weak TAMRA-CGRP signal at the plasma membrane. Cells were preincubated with vehicle, Dyngo-4a (Dy4), Pitstop 2 (PS2), inactive analogs (PS2 and Dy4 inact) (all 30 µM) or sucrose (0.45 M) (n = 4 independent experiments). Scale, 10 µm. g Quantification of localization of TAMRA-CGRP in endosomes (data represent n = 949 veh 0 min, n = 1209 veh 30 min, n = 1111 Dy4 30 min, n = 1016 Dy4 inact 30 min, n = 1700 PS2 30 min, n = 714 PS2 inact 30 min, n = 1896 sucrose) and (h) quantification of the number of TAMRA-CGRP+ve endosomes (data represent n = 5 veh 0 min, n = 7 veh 30 min, n = 7 Dy4 30 min, n = 5 Dy4 inact 30 min, n = 7 PS2 30 min, n = 5 PS2 inact 30 min, n = 5 sucrose). i, j PMA induced by periorbital CGRP (1.5 nmol) or vehicle in C57BL/6 J male mice pretreated (0.5 h) with PS2, Dy4, PS2 or Dy4 inact (all 500 pmol) (n = 8 mice per group). k, l PMA induced by periorbital capsaicin (CPS, 50 pmol) or vehicle in C57BL/6 J male mice pretreated (0.5 h) with PS2, Dy4, PS2 or Dy4 inact (all 500 pmol) (n = 8 mice per group). Mean±SEM. ***P < 0.001 vs. Veh 0 min, and Veh/Veh; §§P < 0.01, §§§P < 0.001 vs. Veh 30 min, PS2 30 min, Dy4 30 min, CGRP/PS2 inact, CGRP/Dy4 inact, CPS/PS2 inact, CPS/Dy4 inact. 1-way (g, h) or 2-way (i–l) ANOVA, Bonferroni correction. Source data are provided as a Source Data file.
