Altered potassium channel distribution and composition in myelinated axons suppresses hyperexcitability following injury
- Kings College London, United Kingdom
- Facultad de Ciencias Biologicas- Pontificia Universidad Catolica de Chile, Chile
- Facultad de Medicina, Pontificia Universidad Catolica de Chile, Chile
- The University of Queensland, Australia
- University of Oxford, United Kingdom
- Hospital Regional Universitario de Málaga. Servicio de Anestesiología, Spain
- De Montfort University, United Kingdom
- UT Health Science Center at San Antonio, United States
- Universidad Mayor, Chile
- Geroscience Center for Brain Health and Metabolism, Chile
- Pontificia Universidad Catolica de Chile, Chile
Figures
Figure 1
Kv1 channels expression in the naïve nerve and 21 days after sciatic nerve axotomy (note that the samples were collected from the neuroma site).
(a) Representative images of longitudinal nerve sections immunostained with Kv1 channels in green (Kv1.2, Kv1.4 and Kv1.6 respectively), a panNav antibody in red (to identify the node), and caspr in …
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Figure 1—source data 1
Source data for Figure 1.
- https://doi.org/10.7554/eLife.12661.004
Figure 2
Nav channel expression.
Representative sections of longitudinal nerves immunostained with Kv1.2 in green, a panNav antibody in red (to identify the node), and caspr in blue (to identify the paranode) from neuroma day 21. (a…
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Figure 2—source data 1
Source data for Figure 2.
- https://doi.org/10.7554/eLife.12661.006
Figure 3
Relocalization ok Kv1.2 and caspr2 at 7 days after neuroma.
(a) Representative longitudinal sections of nerves immuno-stained with Kv1.2 in green, a panNav antibody in red (to identify the node) and caspr in blue (to identify the paranode). Kv1.2 is …
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Figure 3—source data 1
Source data for Figure 3.
- https://doi.org/10.7554/eLife.12661.008
Figure 4
Ultrastructural anatomy of the node of Ranvier within the sciatic nerve following axotomy.
We used electron microscopy to look at the ultrastructure anatomy of the node. (a) Shows a diagram of the node, paranode and juxtaparanode and a low magnification section of this area in a …
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Figure 4—source data 1
Source data for Figure 4.
- https://doi.org/10.7554/eLife.12661.010
Figure 5
βII spectrin expression in naïve and neuroma nerves.
(a) Representative sections of longitudinal nerves immunostained with βII spectrin in green, a panNav antibody in red (to identify the node), and caspr in blue (to identify the paranode). βII …
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Figure 5—source data 1
Source data for Figure 5.
- https://doi.org/10.7554/eLife.12661.012
Figure 6
Expression of Kv1 channels in the DRG in the naïve state, 7 and 21 days after axotomy (neuroma).
(a) Representative sections of naive and neuroma day 21 DRG immunostained with Kv1.2, Kv1.4, and Kv1.6. Note that Kv1.2 expression in DRG cells and axonal juxtaparanodes (arrow heads) is reduced …
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Figure 6—source data 1
Source data for Figure 6.
- https://doi.org/10.7554/eLife.12661.014
Figure 7
Kv1 channels expression in the sural nerve of healthy volunteers (control) and from patients with painful Morton neuroma.
(a) Representative sections of longitudinal nerves immunostained with Kv1 channels in green (Kv1.2, Kv1.4 and Kv1.6 respectively), a panNav antibody in red (to identify the node), and caspr in blue …
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Figure 7—source data 1
Source data for Figure 7.
- https://doi.org/10.7554/eLife.12661.016
Figure 8
Kv1 channels expression in the dorsal roots of naïve animals and 21 days after spinal nerve transection (SNT).
(a) Representative sections of longitudinal dorsal roots immunostained with Kv1 channels in green (Kv1.2, Kv1.4 and Kv1.6, respectively), a panNav antibody in red (to identify the node), and caspr …
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Figure 8—source data 1
Source data for Figure 8.
- https://doi.org/10.7554/eLife.12661.018
Figure 9
Local application of α-DTX reinstates primary afferent hyperexcitability at later time points following nerve injury.
(a) Schematic illustration of 3-chamber recording system. 1) Recording chamber, 2) middle chamber, 3) stimulating chamber. The toxin was applied in chambers 1 or 2, respectively. (b) Following …
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Figure 9—source data 1
Source data for Figure 9.
- https://doi.org/10.7554/eLife.12661.020
Figure 10
Mechanical hypersensitivity is restored by blocking Kv1 channels.
Mechanical withdrawal thresholds were assessed by applying a range of Von Frey hairs to the skin over the neuroma site (labelled with a suture). Animals were randomised to receive either …
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Figure 10—source data 1
Source data for Figure 10.
- https://doi.org/10.7554/eLife.12661.022
Figure 11
Schematic illustration of the changes in Kv1 channels subunit composition and distribution within the nodal complex and the relationship to hyperexcitability.
In the naïve state, Kv1 channels (Kv1.1 and 1.2 shown in green) are localised to the juxtaparanode and separated from voltage-gated sodium channels at the node (red) by the paranode (blue). Acutely …
Author response image 1
Author response image 2
Tables
Table 1
Different antibodies used in the study.
| Antibody | Concentration used IHC WB | Company |
|---|---|---|
| Rabbit anti Pan voltage gated sodium channel (Cat No. S6936) | 1:1000 | Sigma-Aldrich |
| Mouse anti Kv1.2 (K14/16.2) | 1:100 1:500 | UC Davis/NIH NeuroMab Facility |
| Mouse anti Kv1.1 (K36/15.1) | 1:100 1:200 | UC Davis/NIH NeuroMab Facility |
| Mouse anti Kv1.4 (K13/31) | 1:100 1:200 | UC Davis/NIH NeuroMab Facility |
| Mouse anti Kv1.6 (K19/36) | 1:100 1:500 | UC Davis/NIH NeuroMab Facility |
| Guinea Pig anti Caspr | 1:1000 1:1000 | From Dr Manzoor Bhat - UT Health Science Center San Antonio (Bhat et al., 2001) |
| Rabbit anti Caspr2 (ab105581) | 1:500 1:400 | Abcam |
| Rabbit anti Pan Neurofascin | 1:500 | Gift from Prof Peter Brophy- University of Edinburgh (Pomicter et al., 2010) |
| Mouse anti βII spectrin (Clone 42) | 1:500 1:1000 | BD Bioscience |
| GAPDH | 1:10000 | Abcam |
| PGP 9.5 | 1:5000 | Ultraclone |
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IHC: Immunohistochemistry; WB: Western Blot analysis.
