Research ArticleNeuroscience Open Access | 10.1172/JCI161507

Xueming Hu,1 Lixia Du,1 Shenbin Liu,1 Zhou Lan,1 Kaikai Zang,1 Jing Feng,1 Yonghui Zhao,1 Xingliang Yang,1 Zili Xie,1 Peter L. Wang,2 Aaron M. Ver Heul,1 Lvyi Chen,1 Vijay K. Samineni,1 Yan-Qing Wang,3 Kory J. Lavine,4 Robert W. Gereau IV,1 Gregory F. Wu,2,5 and Hongzhen Hu1

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Hu, X. in: JCI | PubMed | Google Scholar

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Yang, X. in: JCI | PubMed | Google Scholar

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Xie, Z. in: JCI | PubMed | Google Scholar |

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Wang, P. in: JCI | PubMed | Google Scholar

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

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1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Chen, L. in: JCI | PubMed | Google Scholar |

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Samineni, V. in: JCI | PubMed | Google Scholar |

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Wang, Y. in: JCI | PubMed | Google Scholar

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Lavine, K. in: JCI | PubMed | Google Scholar

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Gereau, R. in: JCI | PubMed | Google Scholar

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Wu, G. in: JCI | PubMed | Google Scholar |

1Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, and Washington University Pain Center and

2Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri, USA.

3Institute of Acupuncture and Moxibustion and Institute of Integrative Medicine; Department of Integrative Medicine and Neurobiology, School of Basic Medical Science; and State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Fudan University, Shanghai, China.

4Department of Internal Medicine, Cardiovascular Division and

5Department of Neurology, Washington University School of Medicine, St. Louis, Missouri, USA.

Address correspondence to: Hongzhen Hu, Department of Anesthesiology, Center for the Study of Itch and Sensory Disorders, Washington University School of Medicine in St. Louis, 660 S. Euclid Avenue, St. Louis, Missouri 63110, USA. Phone: 314.747.4317; Email: hongzhen.hu@wustl.edu. Or to: Gregory F. Wu, Department of Neurology, Washington University School of Medicine in St. Louis, 4523 Clayton Avenue, St. Louis, Missouri 63110, USA. Phone: 314.362.3293; Email: gfwu@wustl.edu.

Authorship note: XH, LD, and SL contributed equally to this work.

Find articles by Hu, H. in: JCI | PubMed | Google Scholar |

Authorship note: XH, LD, and SL contributed equally to this work.

Published January 26, 2023 - More info

Published in Volume 133, Issue 5 on March 1, 2023
J Clin Invest. 2023;133(5):e161507. https://doi.org/10.1172/JCI161507.
© 2023 Hu et al. This work is licensed under the Creative Commons Attribution 4.0 International License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Published January 26, 2023 - Version history
Received: May 2, 2022; Accepted: January 12, 2023 View PDF Abstract

Microglia, resident macrophages of the CNS, are essential to brain development, homeostasis, and disease. Microglial activation and proliferation are hallmarks of many CNS diseases, including neuropathic pain. However, molecular mechanisms that govern the spinal neuroimmune axis in the setting of neuropathic pain remain incompletely understood. Here, we show that genetic ablation or pharmacological blockade of transient receptor potential vanilloid type 4 (TRPV4) markedly attenuated neuropathic pain-like behaviors in a mouse model of spared nerve injury. Mechanistically, microglia-expressed TRPV4 mediated microglial activation and proliferation and promoted functional and structural plasticity of excitatory spinal neurons through release of lipocalin-2. Our results suggest that microglial TRPV4 channels reside at the center of the neuroimmune axis in the spinal cord, which transforms peripheral nerve injury into central sensitization and neuropathic pain, thereby identifying TRPV4 as a potential new target for the treatment of chronic pain.

Graphical Abstract Introduction

Whereas acute pain is a protective mechanism to avoid further damage, chronic pain is a maladaptive, debilitating condition affecting hundreds of millions of people worldwide (1, 2). About a fifth of people who have chronic pain predominantly have neuropathic pain as result of tissue or nerve injury (3). Although several drugs for relieving chronic pain are available, the use of current first-line pain medicines such as nonsteroidal antiinflammatory drugs, antidepressants, and opioids is limited by serious side effects, dependence, tolerance, and insufficient efficacy (4, 5). This has encouraged major efforts in both academia and industry to develop safer and more effective pain therapies. Notably, current treatments for chronic pain mainly focus on blocking neurotransmission in the pain pathway and have resulted only in limited success. Ironically, although nerve injury produces robust microgliosis, therapeutic strategies targeting microglia for chronic pain treatment remain largely overlooked.

Microglia, tissue-resident macrophages in the CNS, perform dynamic surveillance of their microenvironment to maintain tissue homeostasis. However, when exposed to pathological stimuli, microglia conversely change their functional phenotypes and secrete an excess of proinflammatory cytokines and reactive oxygen species, resulting in maladaptive neurological disorders (6). For instance, following peripheral nerve injury, microglia in the spinal cord convert injury signals from primary sensory neurons, leading to persistent central sensitization and chronic neuropathic pain (7, 8). However, although it has emerged as a vital event for the transition from acute to chronic pain state (9), the molecular mechanisms underlying the interactions between microglia and pain-transmitting spinal neurons are still poorly understood. Specifically, 2 key questions remain to be fully addressed: what is the molecular basis of microgliosis driven by peripheral nerve injury and how does microglial activation results in neuroplasticity leading to chronic neuropathic pain?

The transient receptor potential (TRP) channels are a group of Ca2+-permeable nonselective cation channels that are indispensable for transmitting a wide range of somatosensory stimuli, including itch, pain, temperature, and mechanosensation. Thus, they are typically considered molecular sensors in the primary sensory afferent nociceptors (10). Surprisingly, emerging evidence suggests that TRP channels are also widely expressed and have many physiological and pathological functions in both neurons and nonneuronal cells (11, 12). We and others have recently demonstrated that TRP vanilloid type 4 (TRPV4) expressed by macrophages and microglia is associated with diverse pathophysiological processes (13–15).

Here, we show that TRPV4 was functionally expressed in spinal resident microglia and TRPV4 expression was increased in a mouse model of spared nerve injury (SNI). Genetic ablation and pharmacological inhibition of microglia-expressed TRPV4 markedly suppressed SNI-induced neuropathic pain-related behaviors. Mechanistically, TRPV4 mediated microglial activation and proliferation and promoted functional and structural plasticity of excitatory spinal neurons by releasing lipocalin-2 (LCN2) from activated microglia after SNI. Collectively, our data identified the TRPV4-dependent neuroimmune axis as an essential component required for neuropathic pain generation and provided potential drug targets for the treatment of chronic neuropathic pain with decreased reliance on opioids.

Results

Functional expression of TRPV4 in spinal cord microglia. Although TRPV4 is traditionally believed to be functionally expressed by primary nociceptors (16–19), surprisingly, in the spinal cord, we found no TRPV4-eGFP expression in TRPV1+ primary–afferent terminals by using the Trpv4eGFP:Trpv1Cre/+:tdTomato reporter mice (Supplemental Figure 1A; supplemental material available online with this article; https://doi.org/10.1172/JCI161507DS1). Instead, we detected TRPV4-eGFP signals predominantly in IBA1+/Tmem119+ microglia and CD31+ endothelial cells (20), but not in NeuN+ neurons or GFAP+ astrocytes in the spinal cord (Figure 1, A–C, and Supplemental Figure 1B), suggesting that TRPV4 may participate in pain signaling through immunogenic instead of direct neurogenic mechanisms.

Figure 1

TRPV4 is expressed by spinal microglia. (A) Representative image of TRPV4-eGFP expression in the spinal cord of naive mice. Scale bar: 200 μm. (B and C) Representative images of TRPV4-expressing cells (TRPV4-eGFP+) with microglial marker IBA1, endothelial marker CD31, neuronal marker NeuN, and astrocytic marker GFAP in the spinal dorsal horn of naive mice (B), and quantification of the proportion of TRPV4-eGFP+/IBA1+ microglia and TRPV4-eGFP+/ CD31+ endothelial cells. n = 10 spinal slices from 3 mice (C). (B and C) Scale bar: 100 μm and 20 μm (zoom).

Given that TRPV4 is a Ca2+-permeable ion channel, we corroborated the observation of TRPV4 expression in spinal microglia using Ca2+ imaging with functional verification. We first performed in vitro Ca2+ imaging in cultured primary spinal microglia isolated from both newborn WT C57BL/6J and Trpv4–/– mice. Perfusion with a selective TRPV4 agonist GSK1016790A (GSK101) elicited robust intracellular Ca2+ ([Ca2+]i) responses, which were nearly abolished by coapplied TRPV4 antagonist GSK2193874 (GSK219, Figure 2, A–C). Consistent with pharmacological inhibition studies, GSK101 did not elicit any measurable [Ca2+]i responses in the Trpv4–/– spinal microglia, while ionomycin, used as positive control, elicited robust [Ca2+]i responses in all cells tested (Figure 2, D–F). To avoid potential issues related to the development and cell culture conditions using microglia from newborn mice, we further performed ex vivo Ca2+ imaging in acutely prepared spinal slices from adult Cx3cr1CreER/+:tdTomato:GCaMP6s mice (21). Perfusion with GSK101 elicited large [Ca2+]i transients in the spinal dorsal horn tdTomato+ cells, which was abolished by coapplied GSK219 (Figure 2, G–I). Moreover, we directly recorded GSK101-activated whole-cell currents in ex vivo spinal dorsal horn microglia using the whole-cell patch-clamp recording. Acute perfusion with GSK101 activated robust outwardly rectifying TRPV4-like currents in the spinal microglia from both adult Cx3cr1GFP/+ mice (Figure 2, J–L) and Cx3cr1CreER/+:tdTomato mice (Figure 2, O–Q), which were abolished by GSK219. Consistent with the pharmacological blockade experiments, the GSK101-activated TRPV4-like currents were completely absent in spinal microglia from adult Cx3cr1GFP/+:Trpv4–/– mice (Figure 2, M and N) or Cx3cr1CreER/+:tdTomato:Trpv4fl/fl mice (Figure 2, R and S). Collectively, these results suggest that microglia-expressed TRPV4 is the sole mediator of GSK101-induced responses.

Figure 2

TRPV4 mediates [Ca2+]i responses in spinal microglia. (A–C) Representative Ca2+ imaging (A), percentage (B), and time-lapse averaged Ca2+ traces (C) of in vitro responses to GSK101 (300 nM) and GSK219 (300 nM) exposure in primary spinal microglia from WT mice. (D–F) Representative Ca2+ imaging (D), percentage (E), and time-lapse averaged Ca2+ traces (F) of in vitro responses to GSK101 (300 nM) exposure in primary spinal microglia from WT and Trpv4–/– mice. Ionomycin (1 μM) was used as a positive control. n = 4 coverslips per group from 2 independent experiments. ***P < 0.001 by unpaired 2-tailed Student’s t test. Scale bar: 50 μm. (G–I) Representative Ca2+ imaging (G), time-lapse averaged Ca2+ traces (H), and ΔF/F peak amplitude (I) of ex vivo responses to GSK101 (300 nM) and GSK219 (300 nM) exposure in spinal microglia from Cx3cr1CreER/+:tdTomato:GCaMP6s mice. n = 4 mice per group. Scale bar: 50 μm. *P < 0.05 by unpaired 2-tailed Student’s t test. (J–N) GSK101-activated whole-cell membrane currents in spinal GFP+ microglia. Representative recorded GFP+ microglia images. Scale bar: 10 μm (J). Current-voltage curves and quantification of GSK101 (300 nM) and GSK219 (300 nM) on currents recorded at +100 mV from Cx3cr1GFP/+ mice (K and L) and Cx3cr1GFP/+:Trpv4–/– mice (M and N). (O–S) GSK101-activated whole-cell membrane currents in spinal tdTomato+ microglia. Representative recorded tdTomato+ microglia images. Scale bar: 10 μm (O). Current-voltage curves and quantification of GSK101 (300 nM) and GSK219 (300 nM) on currents recorded at +100 mV from Cx3cr1CreER/+:tdTomato mice (P and Q) and Cx3cr1CreER/+:tdTomato:Trpv4fl/fl mice (R and S). n = 8 cells from 3 mice per group. ***P < 0.001 by 1-way ANOVA with Bonferroni’s post hoc test. Data are represented as mean ± SEM.

TRPV4 was required for pain-related behaviors following spared nerve injury. Although sensory TRP channels are generally considered critical molecular sensors for mechanical, thermal, and chemical cues and contribute to both pain and itch sensations (10, 22–24), the role of centrally expressed TRPV4 in neuropathic pain remains poorly understood. To test this possibility, we generated a well-established mouse model of SNI (25) in both Trpv4–/– (global Trpv4 KO) and congenic WT C57BL/6J mice. Strikingly, compared with WT mice, Trpv4–/– mice had a significantly improved paw withdrawal threshold, a hallmark of mechanical allodynia, in both male and female mice following SNI (Figure 3, A and B). To complement genetic ablation studies, we administered TRPV4 antagonist GSK219 once daily through repeated i.p. injections for 7 consecutive days starting on day 1 after SNI in WT mice and observed a marked increase in paw withdrawal threshold in a dose- and time-dependent manner (Figure 3C). Moreover, acute application of GSK219 through a single intrathecal (i.t.) injection on day 7 after SNI also produced a time- and dose-dependent attenuation of the decreased paw withdrawal threshold caused by SNI (Figure 3D), suggesting that TRPV4 in the spinal cord is involved in both initiation and maintenance of neuropathic pain. Note that i.p. or i.t. injections of GSK219 alone had no significant effect on the paw withdrawal threshold (Supplemental Figure 2, A and B). Next, we directly tested whether the expression of TRPV4 in spinal resident microglia contributed to neuropathic pain using inducible Cx3cr1CreER/+:Trpv4fl/fl (microglia-specific Trpv4 cKO) mice 4 weeks after tamoxifen treatment. The SNI-induced mechanical allodynia was also attenuated in both male and female Cx3cr1CreER/+:Trpv4fl/fl mice, compared with Trpv4fl/fl control littermates (Figure 3, E–G). In marked contrast, mice ablating TRPV4 in either CDH5+ endothelial cells (Cdh5Cre/+:Trpv4fl/fl mice, Figure 3H), TRPV1+ primary sensory neurons (Trpv1Cre/+:Trpv4fl/fl mice, Supplemental Figure 3A), or CCR2+ blood-borne monocytes (Ccr2CreER/+:Trpv4fl/fl mice, Supplemental Figure 3B) exhibited no significant changes in SNI-induced mechanical allodynia.

Figure 3

Genetic or pharmacological inhibition of TRPV4 suppresses neuropathic pain. (A and B) Time course of paw withdrawal threshold following SNI in male (A) and female (B) WT control littermates and Trpv4–/– mice. n = 7–10 mice per group. Statistics were determined by 2-way repeated ANOVA with Bonferroni’s post hoc test. (C) Time course of paw withdrawal threshold following SNI in WT mice treated with repeated i.p. injection of vehicle or GSK219 (once per day from da