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Home / Equine Research Bank / Equine Vet J / Optimisation and validation of immunohistochemical axonal ma...
Equine veterinary journal2021; 53(6); 1188-1198; doi: 10.1111/evj.13403

Optimisation and validation of immunohistochemical axonal markers for morphological and functional characterisation of equine peripheral nerves.

Abstract: Horses are affected by various peripheral nerve disorders but defining their aetiology and pathophysiology is hampered by limited understanding of associated morphological and pathological changes and involvement of specific axonal types. Objective: To investigate the hypothesis that selected antibody markers, used in conjunction with various tissue processing methods, would enable identification of axons with different functional modalities within a range of equine peripheral nerves. Methods: Optimisation and validation study. Methods: A range of antibodies were evaluated immunohistochemically via fluorescence confocal microscopy in cadaver equine nerve samples of primary motor, mixed or primary sensory functions (recurrent laryngeal, phrenic and plantar digital) within formalin-fixed paraffin-embedded (FFPE) and formalin-fixed frozen (FFF) tissues subjected to different antigen retrieval protocols. Results: Immunohistochemistry of FFPE-derived nerve samples with selected antibodies and specific antigen retrieval methods enabled identification of myelinated and unmyelinated axons, cholinergic, sympathetic and peptidergic axons. The recurrent laryngeal and phrenic nerves are composed of myelinated cholinergic (motor), myelinated sensory fibres, unmyelinated adrenergic (sympathetic) axons and unmyelinated peptidergic (sensory) axons. In contrast, as expected, the plantar digital nerve had no myelinated motor fibres being mainly composed of myelinated sensory fibres, unmyelinated sympathetic and unmyelinated peptidergic sensory axons. Conclusions: Attempts specifically to label parasympathetic fibres were unsuccessful in any nerve examined in both FFPE and FFF tissues. Conclusions: A panel of antibody markers can be used to reveal morphological and functional properties of equine nerves. Future work should enable better characterisation of morphological changes in equine neuropathies at various stages of disease development.
© 2021 The Authors. Equine Veterinary Journal published by John Wiley & Sons Ltd on behalf of EVJ Ltd.
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Publication Date: 2021-01-19 PubMed ID: 33338316DOI: 10.1111/evj.13403Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research study focuses on improving the understanding of peripheral nerve disorders in horses, by experimenting with different antibody markers and tissue processing methods to identify specific axonal types linked to horse nerve disorders. Various antibodies were tested on nerve samples and categorized based on different functions to enable more detailed characterisation of equine neuropathies and the potential identification of disease development stages.

Research Objective and Hypothesis

  • The study intended to better comprehend disorders affecting the peripheral nerves in horses, which is currently limited due to a lesser understanding of linked morphological and pathological changes, as well as involved axonal types. The researchers hypothesized that using certain antibody markers and tissue processing techniques might aid in the identification of axons of different functional modalities within a range of equine peripheral nerves.

Methods and Experiments

  • This was an optimisation and validation study where different antibodies were evaluated using immunohistochemical methods and fluorescence confocal microscopy. The experiments were performed on cadaver equine nerve samples, which could be functioning primarily as motor, mixed, or sensory nerves.
  • The nerve samples were treated with formalin and then either embedded in paraffin (FFPE) or frozen (FFF), followed by different antigen retrieval protocols.

Results and Observations

  • By using specific antibodies and antigen retrieval methods on FFPE-treated nerve samples, the researchers were able to distinguish myelinated and unmyelinated axons, as well as cholinergic, sympathetic, and peptidergic axons.
  • The recurrent laryngeal and phrenic nerves mainly consisted of myelinated cholinergic (motor), myelinated sensory fibres, unmyelinated adrenergic (sympathetic) axons, and unmyelinated peptidergic (sensory) axons.
  • On the contrary, the plantar digital nerve, as anticipated, did not have myelinated motor fibres, and was primarily composed of myelinated sensory fibres, unmyelinated sympathetic, and unmyelinated peptidergic sensory axons.

Conclusions and Future Work

  • Attempts to specifically label parasympathetic fibres did not succeed with any of the nerve samples examined with both FFPE and FFF tissue treatments.
  • The study concludes by recommending the use of a panel of antibody markers to reveal the morphological and functional characteristics of equine nerves. The researchers suggest that future work could enable a more detailed understanding of morphological changes in equine neuropathies during the various stages of disease development.

Cite This Article

APA
Almuhanna AH, Cahalan SD, Lane A, Goodwin D, Perkins J, Piercy RJ. (2021). Optimisation and validation of immunohistochemical axonal markers for morphological and functional characterisation of equine peripheral nerves. Equine Vet J, 53(6), 1188-1198. https://doi.org/10.1111/evj.13403

Publication

Equine veterinary journal
ISSN: 2042-3306
NlmUniqueID: 0173320
Country: United States
Language: English
Volume: 53
Issue: 6
Pages: 1188-1198

Researcher Affiliations

Almuhanna, Abdulaziz H
  • Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, Royal Veterinary College, Royal College Street, London, UK.
Cahalan, Stephen D
  • Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, Royal Veterinary College, Royal College Street, London, UK.
Lane, Annette
  • Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, Royal Veterinary College, Royal College Street, London, UK.
Goodwin, David
  • Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, Royal Veterinary College, Royal College Street, London, UK.
Perkins, Justin
  • Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, Royal Veterinary College, Royal College Street, London, UK.
Piercy, Richard J
  • Comparative Neuromuscular Diseases Laboratory, Department of Veterinary Clinical Sciences, Royal Veterinary College, Royal College Street, London, UK.

MeSH Terms

  • Animals
  • Axons
  • Horses
  • Immunohistochemistry
  • Nerve Fibers, Myelinated
  • Peripheral Nerves

Grant Funding

  • Horserace Betting Levy Board
  • Saudi Arabian Cultural Bureau (SACB)

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Citations

This article has been cited 4 times.
  1. Amari M, Brioschi FA, Auletta L, Ravasio G. Ultrasound-Guided Radiofrequency Ablation and Pulsed Radiofrequency Treatment for Chronic Lameness Due to Distal Forelimb Disease in Horses: A Pilot Study. Animals (Basel) 2025 Aug 10;15(16).
    doi: 10.3390/ani15162341pubmed: 40867669google scholar: lookup
  2. Iżycka-Świeszewska E, Gulczyński J, Sejda A, Kitlińska J, Galli S, Rogowski W, Sigorski D. Remarks on Selected Morphological Aspects of Cancer Neuroscience: A Microscopic Photo Review. Biomedicines 2024 Oct 14;12(10).
    doi: 10.3390/biomedicines12102335pubmed: 39457647google scholar: lookup
  3. Amari M, Rabbogliatti V, Ravasio G, Auletta L, Brioschi FA, Riccaboni P, Dell'Aere S, Roccabianca P. Development of an ultrasound-guided radiofrequency ablation technique in the equine cadaveric distal limb: histological findings and potential for treating chronic lameness. Front Vet Sci 2024;11:1437989.
    doi: 10.3389/fvets.2024.1437989pubmed: 39247124google scholar: lookup
  4. Bordoni B, Escher AR, Duczyński M. Proposal for Manual Osteopathic Treatment of the Phrenic Nerve. Cureus 2024 Apr;16(4):e58012.
    doi: 10.7759/cureus.58012pubmed: 38606024google scholar: lookup
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