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In This Article

  • Summary
  • Abstract
  • Introduction
  • Protocol
  • Representative Results
  • Discussion
  • Acknowledgements
  • Materials
  • References
  • Reprints and Permissions

Summary

Evaluation of motor recovery remains the benchmark outcome measure in experimental peripheral nerve studies. The isometric tetanic force measurement of the tibialis anterior muscle in the rat is an invaluable tool to assess functional outcomes after reconstruction of sciatic nerve defects. The methods and nuances are detailed in this article.

Abstract

Traumatic nerve injuries result in substantial functional loss and segmental nerve defects often necessitate the use of autologous interposition nerve grafts. Due to their limited availability and associated donor side morbidity, many studies in the field of nerve regeneration focus on alternative techniques to bridge a segmental nerve gap. In order to investigate the outcomes of surgical or pharmacological experimental treatment options, the rat sciatic nerve model is often used as a bioassay. There are a variety of outcome measurements used in rat models to determine the extent of nerve regeneration. The maximum output force of the target muscle remains the most relevant outcome for clinical translation of experimental therapies. Isometric force measurement of tetanic muscle contraction has previously been described as a reproducible and valid technique for evaluating motor recovery after nerve injury or repair in both rat and rabbit models. In this video, we will provide a step-by-step instruction of this invaluable procedure for assessment of functional recovery of the tibialis anterior muscle in a rat sciatic nerve defect model using optimized parameters. We will describe the necessary pre-surgical preparations in addition to the surgical approach and dissection of the common peroneal nerve and tibialis anterior muscle tendon. The isometric tetanic force measurement technique will be detailed. Determining the optimal muscle length and stimulus pulse frequency is explained and measuring the maximum tetanic muscle contraction is demonstrated.

Introduction

Loss of motor function following traumatic peripheral nerve injury has a significant impact on the quality of life and socioeconomic status of patients1,2,3. The prognosis of this patient population remains poor due to minimal improvements in surgical techniques over the years4. Direct end-to-end tension-free epineural repair forms the gold standard surgical reconstruction. However, in cases with extended nerve gaps interposition of an autologous nerve graft has proven to be superior5,6. The as....

Protocol

All animal procedures were performed with approval of the Institutional Animal Care and Use Committee (IACUC A334818).

1. Calibration of the force transducer

  1. Ensure that the computer is properly connected to the USB-6009 multifunctional I/O data acquisition (DAQ) device, which in turn should be connected to the force transducer.
    NOTE: Other rat strains and species may require a different load-cell force transducer as higher forces are to be expected 44

Representative Results

Five parameters are used to measure the ITF measurement. These include muscle tension (preload force), stimulus intensity (voltage), stimulus pulse frequency, stimulus duration of 0.4 ms and a delay of 2 ms. Prior to measuring the ITF, the optimal muscle tension has to be determined using two single twitch muscle contractions at an intensity of 2 V during the parameter test. These stimuli cause dorsiflexion of the paw and produce an output signal on the graph in the VI (Figure 5). These sing.......

Discussion

This protocol describes a previously validated method for acquiring accurate maximum ITF measurements of the TA muscle in the rat model32. The recovery of maximum strength after experimental nerve reconstruction treatments is of primary interest in the clinical setting as it proves that the nerve not only regenerated, but also made working connections with the target muscle. The ITF can be used in a small nerve gap model, such as the rat sciatic nerve model32, and with a fe.......

Acknowledgements

Research reported in this publication was supported by the National Institute of Neurological Disorders and Stroke of the National Institutes of Health under Award Number RO1 NS 102360. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

....

Materials

NameCompanyCatalog NumberComments
0.9% Sodium ChlorideBaxter Healthcare Corporation, Deerfield, IL, USAG130203
1 mm Kirshner wiresPfizer Howmedica, Rutherford, NJN/A
Adson Tissue ForcepsASSI, Westbury, NY, USAMTK-6801226
Bipolar electrode cablesGrass Instrument, Quincy, MAN/A
Bipolar stimulator deviceGrass SD9, Grass Instrument, Quincy, MAN/A
Cotton-tip ApplicatorsCardinal Health, Waukegan, IL, USAC15055-006
Curved Mosquito forcepsASSI, Westbury, NY, USAMTK-1201112
Force Transducer MDB-2.5Transducer Techniques, Temecula, CAN/A
Gauze Sponges 4x4Covidien, Mansfield, MA, USA2733
Ground cableGrass Instrument, Quincy, MAN/A
Isoflurane chamberN/AN/ACustom-made
KetamineKetalar, Par Pharmaceutical, Chestnut, NJ42023-115-10
LabView SoftwareNational Instruments, Austin, TX
LoopN/AN/ACustom-made
Microsurgical curved forcepsASSI, Westbury, NY, USAJFA-5B
Microsurgical scissorsASSI, Westbury, NY, USASAS-15R-8-18
Microsurgical straight forcepsASSI, Westbury, NY, USAJF-3
RetractorASSI, Westbury, NY, USAAG-124426
Scalpel Blade No. 15Bard-Parker, Aspen Surgical, Caledonia, MI, USA371115
Slim Body Skin StaplerCovidien, Mansfield, MA, USA8886803512
Subminiature electrodeHarvard Apparatus, Holliston, MAN/A
Surgical Nerve StimulatorCheckpoint Surgical LCC, Cleveland, OH, USA9094
Terrell IsofluranePiramal Critical Care Inc., Bethlehem, PA, USAH961J19A
Testing platformN/AN/ACustom-made
Tetontomy ScissorsASSI, Westbury, NY, USAASIM-187
Traceable Big-Digit Timer/StopwatchFisher Scientific, Waltham, MA, USAS407992
USB-6009 multifunctional I/O data acquisition (DAQ) deviceNational Instruments, Austin, TX779026-01
Vacuum Base HolderNoga Engineering & Technology Ltd., Shlomi, IsrealN/AAttached clamp is custom-made
Weight (10 g)Denver Instruments, Denver, CO, USA820010.4
Weight (20 g)Denver Instruments, Denver, CO, USA820020.4
Weight (50 g)Denver Instruments, Denver, CO, USA820050.4
XylazineXylamed, Bimeda MTC Animal Health, Cambridge, Canada1XYL002

References

  1. Taylor, C. A., Braza, D., Rice, J. B., Dillingham, T. The incidence of peripheral nerve injury in extremity trauma. American Journal of Physical Medicine & Rehabilitation. 87 (5), 381-385 (2008).
  2. Huckhagel, T., Nuchtern, J., Regelsberger, J., Lefering, R., TraumaRegister, D. G. U.

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