In mice dorsal root ganglia (DRG), some neurons express calcitonin gene–related peptide (CGRP) without substance P (SP; CGRP+SP-). The projections and functions of these neurons are unknown. Therefore, we combined in vitro axonal tracing with multiple-labeling immunohistochemistry to neurochemically define these neurons and characterize their peripheral and central projections. Cervical spinal cord, DRG, and forepaw skin were removed from C57Bl/6 mice and multiple-labeled for CGRP, SP, and either marker for the sensory neuron subpopulations transient receptor potential vanilloid type 1 (TRPV1), neurofilament 200 (NF200), or vesicular glutamate transporter 2 (VGluT1). To determine central projections of CGRP+SP- neurons, Neurobiotin (NB) was applied to the C7 ventral ramus and visualized in DRG and spinal cord sections colabeled for CGRP and SP. Half (50%) of the CGRP-immunoreactive DRG neurons lacked detectable SP and had a mean soma size of 473 ± 14 μm2 (n = 5); 89% of the CGRP+SP- neurons expressed NF200 (n = 5), but only 32% expressed TRPV1 (n = 5). Cutaneous CGRP+SP- fibers were numerous within dermal papillae and around hair shafts (n = 4). CGRP+SP- boutons were prevalent in lateral lamina I and in lamina IV/V of the dorsal horn (n = 5). NB predominantly labeled fibers penetrating lamina IV/V, 6 ± 3% contained CGRP (n = 5), and 21 ± 2% contained VGluT1 (n = 3). CGRP+SP- afferent neurons are likely to be non-nociceptive. Their soma size, neurochemical profile, and peripheral and central targets suggest that CGRP+SP- neurons are polymodal mechanoceptors. J. Comp. Neurol. 523:2555–2569, 2015. © 2015 Wiley Periodicals, Inc.
The authors used multiple-labeling immunohistochemistry in combination with axonal tracing and high-resolution image analysis to characterize a subpopulation of large CGRP-expressing DRG neurons that lack SP and to determine their peripheral and central projections. The results suggest a possible function in mechanoception rather than nociception.