Background & Aims

The factors that contribute to pain after nerve injury remain incompletely understood. Pain after laser-assisted in situ keratomileusis (LASIK) and photorefractive keratectomy (PRK), two surgical techniques commonly used to correct myopia, provides a very ‘neat’ model, in humans, to study the mechanism of nerve injury in trigeminal afferents and, therefore, the genetic basis of chronic post-surgical pain. These surgical techniques transect the distal axons of trigeminal ganglion neurons within the cornea accurately and reproducibly. However, after surgery, a subset of patients suffer intense and persistent pain, of unknown origin, described by patients as feeling like shards of glass in their eye.
Here we evaluated a TRPV1 variant, p.V527M, found in a 30-year-old woman who had developed corneal pain after LASIK/PRK surgery, reporting an Ocular Surface Disease Index (OSDI) score of 100 [1].

Methods

Human embryonic kidney (HEK) 293 cells were transiently transfected with plasmids encoding wild-type hTRPV1 (WT) and V527M and were analyzed by patch-clamp recordings. In a series of experiments WT and V527M were cotransfected (1:1) to reproduce the patient’s heterozygous state. To determine whether V527M mutation of TRPV1 leads to changes in neuronal excitability, current-clamp recordings were performed on cultured trigeminal neurons transfected with TRPV1 WT and V527M plasmids. Finally, by performing experiments with Fura-2, we tested the response to TRPV1 agonists (capsaicin, acidic pH, high temperature) to understand whether the mutation affects the Ca2+ permeability of the channel, leading to a gain- or loss-of-function phenotype.

Results

We found that V527M exhibits an enhanced response to acidic pH and high temperatures. Specifically, increased proton concentration induced a greater leftward shift in the activation curve of V527M compared to WT, resulting in activity of the mutant channel at acidic pH at more physiological membrane potentials. Furthermore, comparing responses to consecutive applications of different agonists, we found reduced capsaicin-induced desensitization in V527M. Finally, trigeminal neurons expressing V527M showed an enhanced response to acidic pH and increased sensitization by bradykinin, due to a higher affinity of the mutant for the arachidonic acid metabolite, 12-hydroxyethoxyetetraenoic acid (12-HETE), one of the inflammatory mediators released in the cornea after tissue damage.

Conclusions

These results suggest a new mechanism by which this TRPV1 mutation contribute to pain during inflammation after injury of trigeminal axons.

References

[1] Yuan JH, Schulman BR, Effraim PR, Sulayman DH, Jacobs DS, Waxman SG. Genomic analysis of 21 patients with corneal neuralgia after refractive surgery. Pain Rep. 2020 Jul 27;5(4):e826.

Presenting Author

Roberta Gualdani

Poster Authors

Roberta Gualdani

PhD

University of Louvain

Lead Author

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Philippe Gailly

Institute of Neuroscience, University of Louvain, Brussels (Belgium)

Lead Author

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Solène Barbeau

Institute of Neuroscience, University of Louvain, Brussels (Belgium)

Lead Author

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Deborah S. Jacobs

Massachusetts Eye and Ear, Boston, MA (USA)

Lead Author

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Sulayman Dib-Hajj

Neuroscience Research Center, 127A

Lead Author

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Stephen G. Waxman

Department of Neurology, Yale School of Medicine, New Haven, CT (USA)

Lead Author

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Topics

  • Mechanisms: Biological-Molecular and Cell Biology