Neurochemical and Ultrastructural Characterization of Unmyelinated Non-peptidergic C-Nociceptors and C-Low Threshold Mechanoreceptors Projecting to Lamina II of the Mouse Spinal Cord.

C-nociceptors (C-Ncs) and non-nociceptive C-low threshold mechanoreceptors (C-LTMRs) are two subpopulations of small unmyelinated non-peptidergic C-type neurons of the dorsal root ganglia (DRGs) with central projections displaying a specific pattern of termination in the spinal cord dorsal horn. Although these two subpopulations exist in several animals, remarkable neurochemical differences occur between mammals, particularly rat/humans from one side and mouse from the other. Mouse is widely investigated by transcriptomics. Therefore, we here studied the immunocytochemistry of murine C-type DRG neurons and their central terminals in spinal lamina II at light and electron microscopic levels. We used a panel of markers for peptidergic (CGRP), non-peptidergic (IB4), nociceptive (TRPV1), non-nociceptive (VGLUT3) C-type neurons and two strains of transgenic mice: the TAFA4Venus knock-in mouse to localize the TAFA4+ C-LTMRs, and a genetically engineered ginip mouse that allows an inducible and tissue-specific ablation of the DRG neurons expressing GINIP, a key modulator of GABABR-mediated analgesia. We confirmed that IB4 and TAFA4 did not coexist in small non-peptidergic C-type DRG neurons and separately tagged the C-Ncs and the C-LTMRs. We then showed that TRPV1 was expressed in only about 7% of the IB4+ non-peptidergic C-Ncs and their type Ia glomerular terminals within lamina II. Notably, the selective ablation of GINIP did not affect these neurons, whereas it reduced IB4 labeling in the medial part of lamina II and the density of C-LTMRs glomerular terminals to about one half throughout the entire lamina. We discuss the significance of these findings for interspecies differences and functional relevance.

Molecular Structure, Expression and Role of TAFA4 and its Receptor FPR1 in the Spinal Cord.

TAFA chemokine like family member 4 (TAFA4, also named FAM19A4) is a member of the TAFA chemokine like ligand or FAM19A family, which includes TAFA1, TAFA2, TAFA3, TAFA4, and TAFA5 (or FAM19A1, FAM19A2, FAM19A3, FAM19A4, and FAM19A5). They are also referred to as neurokines and are involved in the regulation of a diverse range of cellular processes, including chemotaxis of macrophages, phagocytosis, and release of reactive oxygen species (ROS). TAFA4 is a marker of C-low-threshold mechanoreceptors and is expressed predominantly in nociceptors, such as dorsal root ganglia (DRG). TAFA4 has been implicated in the sensory perception of pain in the spinal cord. Mice with deficiency of TAFA4 demonstrate altered excitability in lamina IIi neurons in DRG in addition to increased mechanical and chemical nociception following inflammation or injury. As a secreted protein, TAFA4 binds to cell surface receptor formyl peptide receptor 1 (FPR1), a G protein-coupled receptor to mediate the chemoattraction of macrophages, phagocytosis, and the inflammatory profile of macrophages. It also interacts with cell surface neurexin to mediate signalling across the synapse. Further understanding the mechanisms by which this conserved protein family regulates diverse biological processes such as in neuronal functions, inflammation, and tissue fibrosis will help to design therapeutic targets for the treatment of TAFA related diseases such as spinal cord injury and neuro-inflammatory disorders.

Diverse roles and modulations of IA in spinal cord pain circuits.

This review highlights recent findings of different amplitude ranges, roles, and modulations of A-type K+ currents (IA) in excitatory (GAD67-GFP-) and inhibitory (GAD67-GFP+) interneurons in mouse spinal cord pain pathways. Endogenous neuropeptides, such as TAFA4, oxytocin, and dynorphin in particular, have been reported to modulate IA in these pain pathways, but only TAFA4 has been shown to fully reverse the opposing modulations that occur selectively in LIIo GAD67-GFP- and LIIi GAD67-GFP+ interneurons following both neuropathic and inflammatory pain. If, as hypothesized here, Kv4 subunits underlie IA in both GAD67-GFP- and GAD67-GFP+ interneurons, then IA diversity in spinal cord pain pathways may depend on the interneuron-subtype-selective expression of Kv4 auxiliary subunits with functionally different N-terminal variants. Thus, IA emerges as a good candidate for explaining the mechanisms underlying injury-induced mechanical hypersensitivity.

TAFA4 relieves injury-induced mechanical hypersensitivity through LDL receptors and modulation of spinal A-type K+ current.

Pain, whether acute or persistent, is a serious medical problem worldwide. However, its management remains unsatisfactory, and new analgesic molecules are required. We show here that TAFA4 reverses inflammatory, postoperative, and spared nerve injury (SNI)-induced mechanical hypersensitivity in male and female mice. TAFA4 requires functional low-density lipoprotein receptor-related proteins (LRPs) because their inhibition by RAP (receptor-associated protein) dose-dependently abolishes its antihypersensitive actions. SNI selectively decreases A-type K+ current (IA) in spinal lamina II outer excitatory interneurons (L-IIo ExINs) and induces a concomitant increase in IA and decrease in hyperpolarization-activated current (Ih) in lamina II inner inhibitory interneurons (L-IIi InhINs). Remarkably, SNI-induced ion current alterations in both IN subtypes were rescued by TAFA4 in an LRP-dependent manner. We provide insights into the mechanism by which TAFA4 reverses injury-induced mechanical hypersensitivity by restoring normal spinal neuron activity and highlight the considerable potential of TAFA4 as a treatment for injury-induced mechanical pain.