Tools for analysis and conditional deletion of subsets of sensory neurons.

Andrei L Okorokov,James J Cox,Federico Iseppon,Larissa De Clauser,Jing Zhao,Marta Alves-Simoes,Sonia Santana-Varela,Yury D Bogdanov,Samuel J Gossage,John N Wood,Ana P Luiz,Jane E Sexton,Shengnan Li

doi:10.12688/wellcomeopenres.17090.1

Andrei L Okorokov, James J Cox + Show 11 more

Open Access

https://doi.org/10.12688/wellcomeopenres.17090.1

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Abstract

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 Na v1.8 Cre and then crossed to CGRP CreER (Calca), Th CreERT2, Tmem45b Cre, Tmem233 Cre, Ntng1 Cre and TrkB CreER (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 Na v1.8 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.

Highlights

Somatosensory neurons have been classified into distinct sets first on the basis of action potential velocity, later with histochemical markers, and more recently by single cell transcriptional profiling[1,2,3,4]
Ablation of specific dorsal root ganglia (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
It was important to rule out effects on other tissues, and so we developed a floxed stop diphtheria toxin mouse driven by a sensory neuron specific promoter, advillin

Summary

Introduction

Somatosensory neurons have been classified into distinct sets first on the basis of action potential velocity, later with histochemical markers, and more recently by single cell transcriptional profiling[1,2,3,4]. 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. 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 TrkBpositive neurons led to significant deficits in mechanical hypersensitivity in the partial sciatic nerve ligation neuropathic pain model

Methods

Results

Conclusion