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Research ArticleNeuroscience
Open Access | 
10.1172/JCI176227
1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
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1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
Find articles by Lee, S. in: JCI | PubMed | Google Scholar
1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
Find articles by Shah, S. in: JCI | PubMed | Google Scholar
1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
Find articles by Maffei, B. in: JCI | PubMed | Google Scholar
1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
Find articles by Berta, T. in: JCI | PubMed | Google Scholar
1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
Find articles by Du, X. in: JCI | PubMed | Google Scholar
1Department of Pharmacology; The Key Laboratory of Neural and Vascular Biology, Ministry of Education; The Key Laboratory of New Drug Pharmacology and Toxicology, Hebei Medical University, Shijiazhuang, Hebei, China.
2Pain Research Center, Department of Anesthesiology, University of Cincinnati Medical Center, Cincinnati, Ohio, USA.
3Faculty of Biological Sciences, School of Biomedical Sciences, University of Leeds, Leeds, United Kingdom.
4Department of Life Sciences, Manchester Metropolitan University, Manchester, United Kingdom.
Address correspondence to: Xiaona Du, Department of Pharmacology, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang, 050011 Hebei, China. Phone: 8631186266073; Email: du_xiaona@hotmail.com. Or to: Nikita Gamper, Faculty of Biological Sciences, University of Leeds, Woodhouse Lane, Leeds LS2 9JT, United Kingdom. Phone: 44.11.3343.7923; Email: n.gamper@leeds.ac.uk. Or to: Temugin Berta, Department of Anesthesiology, University of Cincinnati Medical Center, Medical Sciences Building, 231 Albert Sabin Way, Room 3408, Cincinnati, Ohio 45267, USA. Phone: 513.558.2434; Email: temugin.berta@uc.edu.
Authorship note: XL, ASP, and VP are co–first authors.
Find articles by Gamper, N. in: JCI | PubMed | Google Scholar
Authorship note: XL, ASP, and VP are co–first authors.
Published June 18, 2024 - More info
Published in Volume 134, Issue 16 on August 15, 2024
AbstractWe report that diazepam binding inhibitor (DBI) is a glial messenger mediating crosstalk between satellite glial cells (SGCs) and sensory neurons in the dorsal root ganglion (DRG). DBI is highly expressed in SGCs of mice, rats, and humans, but not in sensory neurons or most other DRG-resident cells. Knockdown of DBI results in a robust mechanical hypersensitivity without major effects on other sensory modalities. In vivo overexpression of DBI in SGCs reduces sensitivity to mechanical stimulation and alleviates mechanical allodynia in neuropathic and inflammatory pain models. We further show that DBI acts as an unconventional agonist and positive allosteric modulator at the neuronal GABAA receptors, particularly strongly affecting those with a high-affinity benzodiazepine binding site. Such receptors are selectively expressed by a subpopulation of mechanosensitive DRG neurons, and these are also more enwrapped with DBI-expressing glia, as compared with other DRG neurons, suggesting a mechanism for a specific effect of DBI on mechanosensation. These findings identified a communication mechanism between peripheral neurons and SGCs. This communication modulates pain signaling and can be targeted therapeutically.
Graphical Abstract
Introduction
Despite remarkable progress in our understanding of the fundamental biology of pain, current therapies for chronic pain are often inadequate and prone to serious side effects within the central nervous system (CNS). Hence, there is an increasing focus on the peripheral nociceptive pathways (1). Specifically, peripheral somatosensory ganglia, such as the dorsal root ganglion (DRG), emerge as an early gate within the somatosensory system (2–5). Because of the pseudounipolar morphology of the DRG neurons, the action potentials traveling from the peripheral nerve terminals to the spinal cord need to pass through the axonal bifurcations (T-junctions) at the DRG, where a propagating spike could fail (2, 3, 6–10). Accumulating evidence suggests that such failure does occur physiologically; moreover, it can be dynamically regulated, manifesting as filtering of the throughput firing frequency at the DRG (2–4, 6, 11). Although fundamentals of such filtering are only beginning to emerge, one revealed mechanism utilizes the ganglion’s intrinsic inhibitory GABAergic system (2, 3, 12). Indeed, peripheral sensory neurons abundantly express GABAA receptors (reviewed in ref. 13), and some of them are capable of producing and releasing GABA (2, 3, 14). Accordingly, direct ganglionic injections of GABAA agonists (2, 15) or GABA reuptake inhibitor (2, 16) provide strong relief of acute and chronic pain. Direct electrophysiological measurements of spike propagation through the DRG revealed that GABA applied to DRG induces spike filtering in the nociceptive fibers, while administration of GABAA antagonists reduces such filtering (3). These reports point to the existence of an inhibitory GABAergic tone at the DRG; this tone contributes to the ganglionic spike filtering, and it can be scaled up or down. Yet it is still unclear how transneuronal communication within the DRG is organized, especially given the fact that DRG neuron somata are individually wrapped by satellite glial cells (SGCs) (17). Being a physical barrier between the sensory neuron somata, SGCs are increasingly recognized as important actors in ganglionic communication (18–23). SGCs express receptors for some of the molecules released by the sensory neurons, particularly the purinergic P2Y receptors activated by ATP (20, 24). In turn, SGCs can also release neuroactive molecules, including ATP, chemokines, and cytokines (IL-1β, TNF-α) (23), which enable them to communicate with neurons and other SGCs. Additionally, SGCs are interconnected through gap junctions, especially connexin-43 (23), enabling complex SGC-to-SGC and SGC-to-neuron communications.
Here we report that SGCs abundantly produce and release a peptide called the diazepam binding inhibitor (DBI), also known as acyl-CoA–binding protein (ACBP). DBI is an 86–amino acid peptide belonging to the group of endogenous compounds exerting benzodiazepine-like effects, endozepines (25). DBI was discovered in the search for endogenous GABAA receptor modulators; it competed with [3H]diazepam for binding to crude synaptic membranes from rat cerebral cortex (26–29). DBI is primarily expressed in astrocytes in the brain, especially in the olfactory bulb, hypothalamus, and hippocampus (30); moreover, Dbi is one of the most transcribed genes in astrocytes (31). DRG-resident SGCs share many features of the CNS’s astrocytes (23, 32); expression of DBI in SGCs was also reported (33, 34). Accumulating evidence suggests that DBI acts as an endogenous GABAA receptor modulator, binding to the benzodiazepine binding site of the GABAA receptors to allosterically modulate its activity (25). Neuronal action of DBI is still under investigation, but both positive (PAM) and negative (NAM) allosteric modulation on GABAA receptors was reported (25, 35, 36). Given the emerging strong role of the GABA system in the control of peripheral nociceptive signaling, we investigated a potential role of DBI in nociception. We report that DBI, acting within DRGs, strongly antagonizes nociception induced by mechanical stimulation of the skin in vivo. Moreover, genetic overexpression of DBI antagonizes mechanical allodynia in the models of chronic inflammatory and neuropathic pain. Our data suggest that DBI acts as an unconventional agonist and a PAM at endogenous GABAA receptors in the mechanosensitive neurons within the ganglion. Such action enhances peripheral ganglionic “gating” of the mechanosensory input to the CNS. Our findings revealed a component in the somatosensory system’s peripheral gate machinery and may point toward new types of analgesia.
ResultsDBI is abundantly expressed in mouse, rat, and human SGCs. First, we tested the expression of DBI within the DRG (schematized in Figure 1A). Presence of the Dbi transcript was detected in both mouse and human DRG (Figure 1B). Analysis of human single-nucleus transcriptomic data (37) revealed coclustering of DBI with FABP7, a validated SGC marker (38) (Supplemental Figure 1A; supplemental material available online with this article; https://doi.org/10.1172/JCI176227DS1). Additionally, mouse RNA-Seq data (39) revealed the highest expression of Dbi in the SGCs, moderate expression in the non-myelinating Schwann cells, and low to non-detectable expression in other DRG-resident cells (Supplemental Figure 1B). Immunostaining identified strong abundance of DBI protein in structures enwrapping human (Figure 1C and Supplemental Figure 2, A and B), mouse (Figure 1E and Supplemental Figure 2, C and D), and rat (Figure 1D and Supplemental Figure 2, E and F) sensory neuron somata. DBI immunoreactivity did not overlap with neuronal markers β-tubulin III (mouse, Figure 1F; human, Supplemental Figure 2B), NF200, and peripherin (rat, Supplemental Figure 2F), nor with the macrophage marker IBA-1 and endothelial cell marker CD31 (mouse, Figure 1F). In contrast, strong colocalization with SGC markers FABP7 (mouse, Figure 1F and Supplemental Figure 2C), glutamine synthetase (mouse, Supplemental Figure 2D), and S100B (rat, Supplemental Figure 2E) was observed in DRG sections. Because S100B is also present in Schwann cells (40), we stained the sciatic nerve (Supplemental Figure 2G). A minority of S100B-positive cells in the nerve displayed DBI immunofluorescence, consistent with specific expression in non-myelinating Schwann cells (39) (Supplemental Figure 1B). DBI immunofluorescence colocalized with Dbi transcript signal (FISH) in an SGC-like pattern, supporting the specificity of our antibody (Figure 1F). Thus, DBI displays a highly specific expression pattern in mouse, rat, and human DRGs with the highest expression in the SGCs. We also used iDISCO (immunolabeling-enabled 3D imaging of solvent-cleared organs) clearance of the entire rat DRG in combination with light-sheet imaging (3) to visualize the SGC-like 3D pattern of DBI expression in the ganglion (Supplemental Video 1).
Figure 1DBI is a satellite glial cell marker. (A) Schematic of superposition of sensory neuron somata and satellite glial cells (SGCs) within the dorsal root ganglion (DRG). (B) Detection of Dbi mRNA expression in human and mouse DRG by reverse transcription (RT) PCR. (C–E) Sections of human (C), rat (D), and mouse (E) DRGs with sensory neuron somata identified with Nissl (green) staining; DBI immunofluorescence (red) forms a characteristic pattern consistent with SGC wrapping. (F) Colabeling of mouse DRG sections with DBI (red) and markers of DRG-resident cell types (all in green): TUBB3 (pan-neuronal marker), IBA-1 (macrophages), CD31 (endothelial cells), and FABP7 (SGCs). Bottom middle panel shows confocal orthogonal view of DBI (red) and FABP7 (green) colabeling of SGCs wrapping around a single sensory neuron somata; blue is DAPI. Bottom right panel shows colabeling of DBI immunofluorescence with the Dbi transcript (white) using FISH. All scale bars: 25 μm.
Glial DBI modulates sensitivity to mechanical stimuli. Next, we asked what physiological role the SGC-derived DBI might play. Acute knockdown of DBI in the DRG using an intrathecal siRNA in mice revealed a striking phenotype: strong increase in sensitivity to punctate and noxious mechanical stimuli (Figure 2, A and E, and Supplemental Figure 3, B–D and F–I), without a change in thermal sensitivity (either heat or cold; Figure 2, B and C). Sensitivity to innocuous mechanical stimulation (adhesive removal test; Figure 2D) was also increased. Thus, the punctate mechanical withdrawal threshold (von Frey) was more than halved (Figure 2A), responses to noxious pinprick were more than doubled (Supplemental Figure 3, D and H), and the latency to response in the alligator clip test dropped more than 4-fold (Figure 2E and Supplemental Figure 3I). Similar effects were recorded in male and female mice (Supplemental Figure 3, F–I). Sensorimotor coordination (rotarod) was not affected by DBI knockdown (Supplemental Figure 3E). Importantly, mechanical hypersensitivity induced by DBI knockdown was completely rescued by the intrathecal delivery of purified DBI (10 ng/site; Figure 2F).
Figure 2Knockdown of Dbi induces mechanical hypersensitivity in mice. (A–E) siRNA against Dbi (or a non-targeting control siRNA) was intrathecally injected (2 mg/site), and 48 hours later the following tests were performed: mechanical sensitivity (von Frey) test (A), cold allodynia (dry ice) test (B), Hargreaves test (C), adhesive removal test (tape assay) (D), and alligator clip test (E). Bars are mean ± SEM. *P < 0.05; **P < 0.01, significant difference for groups indicated by the connector line (unpaired t test). (F) Recovery of mechanical hypersensitivity (von Frey test) induced by intrathecal siRNA knockdown of DBI with intrathecal injection of recombinant DBI (10 ng/site). ###P < 0.001, significant difference from baseline; **P < 0.01, ***P < 0.001, significant difference from time-matched saline group (2-way repeated-measures ANOVA with Šidák’s post hoc test). (G) Schematic timeline for the viral DRG gene delivery and osmotic minipump experiment. Inset depicts DRG 8 weeks after injection with AAV9-U6-shDBI-CAG-EGFP virions; this image is included in the extended data set in Supplemental Figure 4A. Scale bars: 50 μm. (H) Mechanical sensitivity was monitored with the von Frey test during 42 days after the DRG injection of AAV9-U6-shDBI-CAG-EGFP virions or GFP control virions (1.1 × 1012 to 1.2 × 1012 viral genomes/mL; 2 μL). ***P < 0.001, significant difference from time-matched control group (2-way repeated-measures ANOVA with Tukey’s post hoc test). (I) Similar to H, but heat sensitivity was tested with the Hargreaves test. (J and K) Mechanical (J) and heat (K) sensitivity was monitored after the implantation of osmotic minipumps delivering recombinant DBI to the DRG (200 μM, 0.5 μL/h; see Methods) to the mice preinjected with the AAV9-U6-shDBI-CAG-EGFP virions. *P < 0.05; **P < 0.01, significant difference from time-matched control group (2-way repeated-measures ANOVA with Tukey’s post hoc test).
To produce sustained downregulation of DBI expression in the mouse DRG, we performed intra–L4 DRG injections of anti-Dbi shRNA construct incorporated into adeno-associated virions (AAV9-shDBI), which also contained EGFP. Two weeks after injection, the EGFP fluorescence was readily detectable in the DRG (Figure 2G and Supplemental Figure 4A) but not in the spinal cord (Supplemental Figure 5A), and Dbi transcript levels in the whole DRG were reduced by approximately 50% (in comparison with the animals receiving EGFP-only control virions; Supplemental Figure 6, A and B). Dbi transcript levels in the spinal cord were not affected (Supplemental Figure 5, C and D). Strikingly, AAV9-shDBI (but not AAV9-control) induced strong mechanical hypersensitivity, manifested in dramatic sensitization to mechanical, but not thermal, stimulation (Figure 2, H and I), which became significant at 14 days after viral infection and persisted for the duration of the experiment (42 days after injection). The hypersensitivity was partially alleviated by DRG delivery of recombinant DBI via the implanted minipump (Figure 2, G, J, and K). Notably, approximately 2-week delay is expected for AAV-mediated transgene expression in vivo (41). There was also no change to noxious mechanical sensitivity in the contralateral paw (Supplemental Figure 6, D and E). Sensitivity to innocuous mechanical stimulation (cotton swab test; not shown) was unchanged.
Viral constructs used in the above experiments carried general U6 promoter to drive shRNA expression; this would not target SGCs specifically. Hence, in the next experiments, we constructed AAV5 virions with DBI expression under control of the astroglial GFAP promoter (gfaABC1D), which has successfully been used for viral overexpression of genes in SGCs (42). In the first experiment we asked whether SGC-specific DBI overexpression would reduce sensitivity to noxious mechanical stimulation. To this end, we injected AAV5-gfaABC1D-DBI (or EGFP control) into L4 DRG of mice. This resulted in strong overexpression of DBI in the DRG with EGFP fluorescence often displaying a characteristic “ring” pattern (Figure 3, A and B, and Supplemental Figure 4B); expression of DBI in the spinal cord was not affected (Supplemental Figure 5, B–D). Glia-specific DBI overexpression reduced mechanical sensitivity on the ipsilateral side (significant from day 21 after viral injection; Figure 3C) with no contralateral effect (Figure 3E). Thermal sensitivity was minimally affected on the ipsilateral and contralateral sides (Figure 3, D and F).
Figure 3SGC-targeted DBI overexpression reduces mechanical sensitivity in naive mice and suppresses mechanical allodynia in neuropathic and inflammatory pain models. (A) Schematic timeline for the viral DRG gene delivery and chronic pain induction experiments. Inset depicts DRG 8 weeks after injection with AAV5-gfaABC1D-DBI-EGFP virions; this image is included in the extended data set in Supplemental Figure 4B. Scale bars: 50 μm. (B) Reverse transcription PCR confirmation of Dbi overexpression in the DRG. ***P < 0.001, significant difference from control group (unpaired t test). (C and D) Mechanical (von Frey; C) and heat (Hargreaves; D) sensitivity was monitored on the ipsilateral paws during 42 days after the DRG injec
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