Contribution of dorsal horn CGRP-expressing interneurons to mechanical sensitivity

Löken, Line; Bráz, Joao; Etlin, Alexander; Sadeghi, Mahsa; Bernstein, Mollie; Jewell, Madison; Steyert, Marilyn; Kuhn, Julia; Hamel, Katherine; Llewellyn‐Smith, Ida J.; Basbaum, Allan I.

doi:10.7554/elife.59751

Cited by 27 publications

(15 citation statements)

References 51 publications

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“…For example, it was recently shown that CGRP is also expressed in a subset of dorsal horn interneurons, and their selective activation or ablation has behavioral consequences (Löken et al , DOI: 10.7554/eLife.59751). 1 The authors could cite these and other relevant literature to strengthen the point for developing this new tool. With the tool in hand, the authors explored the behavioral effects of ablating different subsets of primary sensory neurons and noted some differences.…”

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confidence: 99%

Tools for analysis and conditional deletion of subsets of sensory neurons

et al. 2021

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Background: Somatosensation depends on primary sensory neurons of the trigeminal and dorsal root ganglia (DRG). Transcriptional profiling of mouse DRG sensory neurons has defined at least 18 distinct neuronal cell types. Using an advillin promoter, we have generated a transgenic mouse line that only expresses diphtheria toxin A (DTA) in sensory neurons in the presence of Cre recombinase. This has allowed us to ablate specific neuronal subsets within the DRG using a range of established and novel Cre lines that encompass all sets of sensory neurons. Methods: A floxed-tdTomato-stop-DTA bacterial artificial chromosome (BAC) transgenic reporter line (AdvDTA) under the control of the mouse advillin DRG promoter was generated. The line was first validated using a Nav1.8Cre and then crossed to CGRPCreER (Calca), ThCreERT2, Tmem45bCre, Tmem233Cre, Ntng1Cre and TrkBCreER (Ntrk2) lines. Pain behavioural assays included Hargreaves’, hot plate, Randall-Selitto, cold plantar, partial sciatic nerve ligation and formalin tests. Results: Motor activity, as assessed by the rotarod test, was normal for all lines tested. Noxious mechanosensation was significantly reduced when either Nav1.8 positive neurons or Tmem45b positive neurons were ablated whilst acute heat pain was unaffected. In contrast, noxious mechanosensation was normal following ablation of CGRP-positive neurons but acute heat pain thresholds were significantly elevated and a reduction in nocifensive responses was observed in the second phase of the formalin test. Ablation of TrkB-positive neurons led to significant deficits in mechanical hypersensitivity in the partial sciatic nerve ligation neuropathic pain model. Conclusions: Ablation of specific DRG neuronal subsets using the AdvDTA line will be a useful resource for further functional characterization of somatosensory processing, neuro-immune interactions and chronic pain disorders.

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confidence: 99%

Tools for analysis and conditional deletion of subsets of sensory neurons

et al. 2021

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“…We found that compared with the vehicle-treated mice, the paw thermal withdrawal latency (PTWL) of T1DM mice dropped after STZ injection ( Figure 1(b) ; T1DM vs. control, F (1, 14) = 155.7, P < 0.0001, n = 8), indicating the occurrence of thermal hypersensitivity in T1DM mice. CGRP is a neuropeptide widely distributed in nociceptive circuits and is closely involved in pain transmission [ 41 ]. The immunostaining of the SDH suggested that the CGRP immunoreactivity of T1DM mice was more intense than that of control mice ( Figure 1(c) ).…”

Section: Resultsmentioning

confidence: 99%

Painful Diabetic Neuropathy Is Associated with Compromised Microglial IGF-1 Signaling Which Can Be Rescued by Green Tea Polyphenol EGCG in Mice

Chen

Yue

Tang

et al. 2022

Oxidative Medicine and Cellular Longevity

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Background. Painful diabetic neuropathy (PDN) is a frequent and troublesome complication of diabetes, with little effective treatment. PDN is characterized by specific spinal microglia-mediated neuroinflammation. Insulin-like growth factor 1 (IGF-1) primarily derives from microglia in the brain and serves a vital role in averting the microglial transition into the proinflammatory M1 phenotype. Given that epigallocatechin-3-gallate (EGCG) is a potent anti-inflammatory agent that can regulate IGF-1 signaling, we speculated that EGCG administration might reduce spinal microglia-related neuroinflammation and combat the development of PDN through IGF-1/IGF1R signaling. Methods. Type 1 diabetes mellitus (T1DM) was established by a single intraperitoneal (i.p.) injection of streptozotocin (STZ) in mice. The protein expression level of IGF-1, its receptor IGF1R, interleukin 1β (IL-1β), tumor necrosis factor-α (TNF-α), and inducible nitric oxide synthase (iNOS) was determined by Western blot or immunofluorescence. Results. The spinal IGF-1 expression markedly decreased along with the presence of pain-like behaviors, the spinal genesis of neuroinflammation (increased IL-1β, TNF-α, and Iba-1+ microglia), and the intensified M1 microglia polarization (increased iNOS+Iba-1+ microglia) in diabetic mice. IGF-1 could colocalize with neurons, astrocytes, and microglia, but only microglial IGF-1 was repressed in T1DM mice. Furthermore, we found that i.t. administration of mouse recombinant IGF-1 (rIGF-1) as well as i.t. or i.p. treatment with EGCG alleviated the diabetes-induced pain-like behaviors, reduced neuroinflammation (suppressed IL-1β, TNF-α, and Iba-1+ microglia), prevented the M1 microglia polarization (less iNOS+Iba-1+ microglia), and restored the microglial IGF-1 expression. Conclusions. Our data highlighted the importance of maintaining spinal IGF-1 signaling in treating microglia-related neuroinflammation in PDN. This study also provides novel insights into the neuroprotective mechanisms of EGCG against neuropathic pain and neuroinflammation through IGF-1 signaling, indicating that this agent may be a promising treatment for PDN in the clinical setting.

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“…Studies have identified that loss of Lbx1 leads to improper patterning of dorsal horn neurons and disrupted sensory afferent innervation 62,63 . Furthermore, Lbx1 has been utilized in intersectional approaches to target neurons implicated in mechanical hypersensitivity 64 , pain 65,66 and itch 28,67 . To further understand Lbx1 , we developed a method to target Lbx1 -lineage neurons in the adult.…”

Section: Resultsmentioning

confidence: 99%

Developmentally determined intersectional genetic strategies to dissect adult sensorimotor function

Bohic

Upadhyay

Keating

et al. 2022

Preprint

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SUMMARYImprovements in the speed and cost of expression profiling of neuronal tissues offer an unprecedented opportunity to define ever finer subgroups of neurons for functional studies. In the spinal cord, single cell RNA sequencing studies1,2support decades of work on spinal cord lineage studies3–5, offering a unique opportunity to probe adult function based on developmental lineage. While Cre/Flp recombinase intersectional strategies remain a powerful tool to manipulate spinal neurons6–8, the field lacks genetic tools and strategies to restrict manipulations to the adult mouse spinal cord at the speed at which new tools develop. This study establishes a new workflow for intersectional mouse-viral strategies to dissect adult spinal function based on developmental lineages in a modular fashion. To restrict manipulations to the spinal cord, we generate a brain-sparingHoxb8FlpOmouse line restricting Flp recombinase expression to caudal tissue. Recapitulating endogenousHoxb8gene expression9, Flp-dependent reporter expression is present in the caudal embryo starting day 9.5. This expression restricts Flp activity in the adult to the caudal brainstem and below.Hoxb8FlpOheterozygous and homozygous mice do not develop any of the sensory or locomotor phenotypes evident in Hoxb8 heterozygous or mutant animals10,11, suggesting normal developmental function of the Hoxb8 gene and protein inHoxb8FlpOmice. Compared to the variability of brain recombination in available caudal Cre and Flp lines12,13Hoxb8FlpOactivity is not present in the brain above the caudal brainstem, independent of mouse genetic background. Lastly, we combine theHoxb8FlpOmouse line with dorsal horn developmental lineage Cre mouse lines to express GFP in developmentally determined dorsal horn populations. Using GFP-dependent Cre recombinase viruses14and Cre recombinase-dependent inhibitory chemogenetics, we target developmentally defined lineages in the adult. We show how developmental knock-out versus transient adult silencing of the same RORβlineage neurons affects adult sensorimotor behavior. In summary, this new mouse line and viral approach provides a blueprint to dissect adult somatosensory circuit function using Cre/Flp genetic tools to target spinal cord interneurons based on genetic lineage.In briefWe describe the generation of aHoxb8FlpOmouse line that targets Flp-recombinase expression to the spinal cord, dorsal root ganglia, and caudal viscera. This line can be used in intersectional Cre/Flp strategies to restrict manipulations to the caudal nervous system. Additionally, we describe an intersectional genetics+viral strategy to convert developmental GFP expression into adult Cre expression, allowing for modular incorporation of viral tools into intersectional genetics. This approach allows for manipulation of a developmentally determined lineage in the adult. This strategy is also more accessible than traditional intersectional genetics, and can adapt to the constantly evolving available viral repertoire.Highlights-A newHoxb8FlpOmouse line allows Flp-dependent recombination in the spinal cord, dorsal root ganglia, and caudal viscera.-We observed no ectopic brain expression across mouse genetic backgrounds with theHoxb8FlpOmouse line.-Combining this new mouse line for intersectional genetics and a viral approach, we provide a novel pipeline to target and manipulate developmentally defined adult spinal circuits.

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Contribution of dorsal horn CGRP-expressing interneurons to mechanical sensitivity

Cited by 27 publications

References 51 publications

Tools for analysis and conditional deletion of subsets of sensory neurons

Tools for analysis and conditional deletion of subsets of sensory neurons

Painful Diabetic Neuropathy Is Associated with Compromised Microglial IGF-1 Signaling Which Can Be Rescued by Green Tea Polyphenol EGCG in Mice

Developmentally determined intersectional genetic strategies to dissect adult sensorimotor function

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