Peripheral gating of mechanosensation by glial diazepam binding inhibitor
Xinmeng Li, Arthur Silveira Prudente, Vincenzo Prato, Xianchuan Guo, Han Hao, Frederick Jones, Sofia Figoli, Pierce Mullen, Yujin Wang, Raquel Tonello, Sang Hoon Lee, Shihab Shah, Benito Maffei, Temugin Berta, Xiaona Du, Nikita Gamper
Xinmeng Li, Arthur Silveira Prudente, Vincenzo Prato, Xianchuan Guo, Han Hao, Frederick Jones, Sofia Figoli, Pierce Mullen, Yujin Wang, Raquel Tonello, Sang Hoon Lee, Shihab Shah, Benito Maffei, Temugin Berta, Xiaona Du, Nikita Gamper
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Research Article Neuroscience

Peripheral gating of mechanosensation by glial diazepam binding inhibitor

Abstract

We 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.

Authors

Xinmeng Li, Arthur Silveira Prudente, Vincenzo Prato, Xianchuan Guo, Han Hao, Frederick Jones, Sofia Figoli, Pierce Mullen, Yujin Wang, Raquel Tonello, Sang Hoon Lee, Shihab Shah, Benito Maffei, Temugin Berta, Xiaona Du, Nikita Gamper

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Figure 4

DBI is released by SGCs.

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DBI is released by SGCs. (A) Detection of DBI by ELISA in extracellular ...
(A) Detection of DBI by ELISA in extracellular media from acutely extracted DRG (containing all the cell types within the ganglion, “Acute DRG”) and from purified mouse SGC culture (“Purified SGC culture”). Measurements were made after 30 minutes of incubation in control medium and in medium with high extracellular [K+] (10 or 100 mM). *P < 0.05; ***P < 0.001, significant difference from 0 mM KCl group (1-way ANOVA with Tukey’s post hoc test). (B–G) Detection of endozepine release by cultured purified rat SGCs using “reporter” HEK293 cells transfected with α1, β2, and γ2 subunits of GABAA receptors and a halide-sensitive EYFP mutant (H148Q/I152L; EYFP-QL). (B) Micrographs depicting coculture of reporter HEK293 cells (green) with purified primary rat SGC culture. Scale bars: 20 μm. (C) Schematic of experimental timeline. (D) Example of the experiment: Reporter HEK293 cells alone (black line) or in coculture with SGCs (red line) are imaged in the presence of 5 mM extracellular iodide. After introduction of the I–-containing solution, the perfusion is stopped for 5 minutes to allow releasable molecules to accumulate. When GABAA receptors are activated, I– enters the cells and produces EYFP-QL fluorescence quenching. GABA (5 μM) is added at the end of the experiment to authenticate fluorescence quenching. (E–G) Top panels display mean data for EYFP-QL quenching of reporter HEK293 cells only (black symbols) or in coculture with SGCs (blue symbols) in control conditions (E) or in the presence of the GABAA receptor blocker bicuculline (50 μM; F) or the benzodiazepine antagonist flumazenil (8 μM; G). Data from each biological replicate are color-coded. Bottom panels show frequency distribution histograms for the data sets summarized in the corresponding top panels. **P < 0.01, ***P < 0.001, significant difference from the reporter HEK293 cells only (Mann-Whitney test).