Ca2+-binding protein NECAB2 facilitates inflammatory pain hypersensitivity
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
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
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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

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Figure 4

Genetic deletion of NECAB2 gates inflammatory but not neuropathic pain.

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Genetic deletion of NECAB2 gates inflammatory but not neuropathic pain. ...
(A) Basal nociceptive sensation (upon noxious mechanical stimulus by pinprick) in Necab2–/– mice was intact (relative to WT mice) in a sex-independent manner (n = 5–9/group). (B–D) After SNI, Necab2–/– mice developed tactile allodynia (von Frey filaments, innocuous stimulus), mechanical hypersensitivity (noxious mechanical stimulus), and cold allodynia (acetone stimulus) to a degree equivalent to that of their WT counterparts (n = 12 for WT and n = 14 for Necab2–/– mice of both sexes). (E) Time course of edema in the hind paw, the typical symptom of inflammation, for WT and Necab2–/– mice (n >8/time point/group) after intraplantar injection of λ carrageenan (1%, 20 μl). (F) Time course of tactile allodynia after λ carrageenan application differed between Necab2–/– and WT mice. (G–I)Differential responses with the von Frey filament test after inflammation. Note the rapid behavioral recovery upon Necab2 deletion. (J–L) After SNI, secretagogin-null (Scgn–/–) mice (all male) developed tactile allodynia, mechanical hypersensitivity, and cold allodynia that were indistinguishable from what was observed in the WT littermates (n = 10/group). (M) Likewise, Scgn–/– and WT mice developed tactile allodynia to a similar degree upon λ carrageenan–induced inflammation (n = 6/group). Error bars in black and red correspond to WT and Scgn–/– mice, respectively. Behavioral data lacking normal distribution are presented as the median ± interquartile range and were statistically assessed by Mann-Whitney U test (e.g., von Frey filament test). Results of other behavioral assays (e.g., pinprick and acetone stimuli) are expressed as the mean ± SD and were statistically evaluated using a Student’s t test (**P < 0.01). BL, basal level.