Figure 3

Characterizing the localization of Grp in DRG, spinal cord, and dorsal root using anti-bombesin antiserum. (A-B) DRGs immunostained with antibodies against bombesin (red) and Cgrp (green). Arrow indicates a bombesin⁺;Cgrp⁺ neuron. (C-D) DRG neurons in adjacent sections are immunostained with anti-bombesin antiserum preabsorbed with purified Grp (red) and anti-Cgrp antibody (green). Preabsorption results in a complete loss of anti-bombesin immunoreactivity. (E-F) DRG neurons immunostained with antibodies against bombesin (red) and Pap (green). (G-H) DRG neurons immunostained with anti-bombesin anti-serum (red) and fluorescently-conjugated IB4 (green). (I-J) DRG neurons immunostained with antibodies against bombesin (red) and VGlut1 (green). Arrows in (E-J) indicate bombesin⁺ neurons that are not immunopositive for the costaining marker. (K-L) dSC section immunostained with antibodies against bombesin (red) and Cgrp (green). The dSC layers positive for bombesin are outlined with white dots and higher magnification of the staining is shown in the inset. (M-N) dSC section immunostained with anti-bombesin antiserum preabsorbed with purified Grp (red) and anti-Cgrp antibody (green), showing a complete loss of anti-bombesin immunoreactivity. (O-P) dSC section immunostained with antibodies against bombesin (red) and Pap (green). (Q-R) dSC section immunostained with anti-bombesin serum (red) and fluorescently-conjugated IB4 (green). (S-T) dSC section immunostained with antibodies against bombesin (red) and VGlut1 (green). (U) A high magnification confocal image of adult dSC immunostained with anti-bombesin (red) and anti-NeuN (green). (V-X) A higher magnification of wild type adult dorsal root immunostained with antibodies against bombesin (red) and Cgrp (green) reveals the presence of Cgrp, but not Grp, in axons projecting from the DRG to dSC. Dorsal root is outlined by dotted line. Arrow indicates the dSC anti-bombesin immunoreactivity. Scale bars = 50 μm (A-J), 100 μm (K-T), 5 μm (U), 30 μm (V-X).