Ca2+-binding protein NECAB2 facilitates inflammatory pain hypersensitivity
Ming-Dong Zhang, … , Tibor Harkany, Tomas Hökfelt
Ming-Dong Zhang, … , Tibor Harkany, Tomas Hökfelt
Published June 12, 2018
Citation Information: J Clin Invest. 2018;128(9):3757-3768. https://doi.org/10.1172/JCI120913.
View: Text | PDF
Research Article Neuroscience

Ca2+-binding protein NECAB2 facilitates inflammatory pain hypersensitivity

Abstract

Pain signals are transmitted by multisynaptic glutamatergic pathways. Their first synapse between primary nociceptors and excitatory spinal interneurons gates the sensory load. In this pathway, glutamate release is orchestrated by Ca2+-sensor proteins, with N-terminal EF-hand Ca2+-binding protein 2 (NECAB2) being particular abundant. However, neither the importance of NECAB2+ neuronal contingents in dorsal root ganglia (DRGs) and spinal cord nor the function determination by NECAB2 has been defined. A combination of histochemical analyses and single-cell RNA-sequencing showed NECAB2 in small- and medium-sized C- and Aδ D-hair low-threshold mechanoreceptors in DRGs, as well as in protein kinase C γ excitatory spinal interneurons. NECAB2 was downregulated by peripheral nerve injury, leading to the hypothesis that NECAB2 loss of function could limit pain sensation. Indeed, Necab2–/– mice reached a pain-free state significantly faster after peripheral inflammation than did WT littermates. Genetic access to transiently activated neurons revealed that a mediodorsal cohort of NECAB2+ neurons mediates inflammatory pain in the mouse spinal dorsal horn. Here, besides dampening excitatory transmission in spinal interneurons, NECAB2 limited pronociceptive brain-derived neurotrophic factor (BDNF) release from sensory afferents. Hoxb8-dependent reinstatement of NECAB2 expression in Necab2–/– mice then demonstrated that spinal and DRG NECAB2 alone could control inflammation-induced sensory hypersensitivity. Overall, we identify NECAB2 as a critical component of pronociceptive pain signaling, whose inactivation offers substantial pain relief.

Authors

Ming-Dong Zhang, Jie Su, Csaba Adori, Valentina Cinquina, Katarzyna Malenczyk, Fatima Girach, Changgeng Peng, Patrik Ernfors, Peter Löw, Lotta Borgius, Ole Kiehn, Masahiko Watanabe, Mathias Uhlén, Nicholas Mitsios, Jan Mulder, Tibor Harkany, Tomas Hökfelt

×

Figure 1

NECAB2 expression in DRGs and spinal cord.

Options: View larger image (or click on image) Download as PowerPoint
NECAB2 expression in DRGs and spinal cord. (A and B) NECAB2 immunoreacti...
(A and B) NECAB2 immunoreactivity in DRGs from WT and Necab2–/– mice showing no residual immunosignal in the mice on a null background (B). (C and D) Coincident detection of NECAB2 and CGRP, a peptidergic marker (C), or IB4, a nonpeptidergic marker for nociceptors (D). (E and F) NECAB2 coexists with TH in C-LTMRs (E) or TrkB in Aδ D-hair LTMRs (F). (G and H) NECAB2 also colocalized with calbindin D28k (G) but not secretagogin (H) in DRGs. (I) Small-diameter VGLUT2::EGFP, but not VGLUT1+, neurons harbored NECAB2 in DRGs. (J and K) Neurochemical heterogeneity of NECAB2+ neurons in DRGs. (L) Molecular phenotyping of Necab2-expressing DRG neurons by reprocessing open-source, single-cell RNA-seq data (26). (M and N) NECAB2 immunoreactivity in spinal dorsal horn of WT mice (L) and its complete loss upon genetic ablation (Necab2–/–) (M). DAPI was used as a nuclear counterstain. (O) Colocalization of PKCγ and Necab2 mRNA in excitatory interneurons in spinal dorsal horn. The rectangle denotes the position of the inset. Projection image for enlarged inset in O is from 11-μm-thick tissue samples orthogonally scanned, with optical steps of 1 μm. Tissues from 2 or more mice were processed for histochemical analysis. Solid and open arrowheads point to colocalization and the lack thereof, respectively. Scale bars: 100 μm (A–I and M–O), 20 μm (J, K, and O, inset).