Behavioral Measures of Pain Thresholds

Michael S. Minett1, Kathryn Quick2, John N. Wood1

1 London Pain Consortium, Kings College London, London, United Kingdom, 2 Molecular Nociception Group, Wolfson Institute for Biomedical Research, University College London, London, United Kingdom
Publication Name:  Current Protocols in Mouse Biology
Unit Number:   
DOI:  10.1002/9780470942390.mo110116
Online Posting Date:  September, 2011
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Abstract

Pain afflicts a fifth of the population, and animal models have proven useful in target validation and analgesic drug development. Thresholds to pain are tested by applying a sensory stimulus, such as heat or pressure, and observing the resulting withdrawal behavior. Sensitized pain models involve provoking an inflammatory response or damaging the nerves themselves, and testing the changes in pain threshold. In this article, mouse models of acute mechanical and thermal pain and inflammatory, visceral, and neuropathic pain are discussed. These behavioral measures can be used to phenotype transgenic mice for target validation and mechanistic studies, as well as to screen potential analgesic compounds. Curr. Protoc. Mouse Biol. 1:383‐412 © 2011 by John Wiley & Sons, Inc.

Keywords: nociception; reflex; pain; neuropathic; inflammatory; visceral

     
 
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Table of Contents

  • Introduction
  • Strategic Planning
  • Testing Thresholds to Thermal Stimuli
  • Basic Protocol 1: Radiant Heat Tail‐Flick
  • Alternate Protocol 1: Hot Water Immersion Tail‐Flick
  • Alternate Protocol 2: Cold Water Immersion Tail‐Flick
  • Basic Protocol 2: Hot Plate: Nociceptive Response to Noxious Heat
  • Basic Protocol 3: Cold Plate: Noxious Cold
  • Basic Protocol 4: Hargreaves' Test: Withdrawal Threshold to Noxious Heat
  • Support Protocol 1: Alternative Measurement for Hargreaves' Apparatus
  • Basic Protocol 5: Acetone Evaporation Test: Innocuous Cooling and Cold Allodynia
  • Testing Thresholds to Mechanical Stimuli
  • Basic Protocol 6: Von Frey Test: Light Touch Perception Threshold
  • Support Protocol 2: Repeated Measures Von Frey Test
  • Alternate Protocol 3: Automatic Von Frey Test
  • Basic Protocol 7: Randall‐Selitto Test: Noxious Mechanical Pressure
  • Alternate Protocol 4: Digital Paw Pressure Test
  • Inflammatory Pain
  • Basic Protocol 8: Mechanical and/or Thermal Hyperalgesia in the Hindpaw
  • Alternate Protocol 5: Spontaneous Inflammatory Pain
  • Visceral Pain
  • Basic Protocol 9: Intracolonic Administration of Irritants
  • Support Protocol 3: Von Frey Hair
  • Neuropathic Pain: Hyperalgesic Allodynia Mediated by Peripheral Nerve Damage
  • Basic Protocol 10: Chronic Constriction Injury (CCI): Loose Ligation of the Sciatic Nerve
  • Alternate Protocol 6: Seltzer: Partial Ligation of the Sciatic Nerve
  • Alternate Protocol 7: Spinal Nerve Ligation (SNL)
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: Radiant Heat Tail‐Flick

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Mouse restrainer (consisting of base plate, body tube, and head cover)
  • Infrared heat source with built‐in timer and motion detector. (e.g., Tail Flick Analgesia Meter #33; IITC Life Science, http://www.iitcinc.com/)
  • Infrared heat‐flux radiometer (Ugo Basile, cat. no. 37300), for calibrating and maintaining apparatus

Alternate Protocol 1: Hot Water Immersion Tail‐Flick

  • Mice of optimal age (6 to 8 weeks)
  • Water bath
  • Thermometer (accurate to 0.5°C)
  • Timer

Alternate Protocol 2: Cold Water Immersion Tail‐Flick

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Hot plate apparatus with enclosure (e.g., Hot/Cold Plate; Ugo Basile, cat. no. 35100)
  • Surface thermometer, for calibrating and maintaining apparatus (optional)

Basic Protocol 2: Hot Plate: Nociceptive Response to Noxious Heat

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Cold plate apparatus with enclosure (e.g. Hot/Cold Plate; Ugo Basile, cat. no. 35100)
  • Surface thermometer, for calibrating and maintaining apparatus (optional)

Basic Protocol 3: Cold Plate: Noxious Cold

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Hargreaves' apparatus, infrared or light (IITC Life Science, cat. no. 390; http://www.iitcinc.com/; Ugo Basile, cat. no. 37370)
  • Raised glass pane with clear plastic enclosures
  • Optional: Infrared heat‐flux radiometer (Ugo Basile, cat. no. 37300), for calibrating and maintaining apparatus

Basic Protocol 4: Hargreaves' Test: Withdrawal Threshold to Noxious Heat

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Acetone
  • Test compartments with a mesh floor (e.g., Mesh Stand; IITC Life Science, cat. no. 410; http://www.iitcinc.com)
  • Acetone applicator (blunting two 33‐G needles and inserting them in either end of a 10‐cm length of thin tubing, and using a 5‐ml syringe, have approved successful previously; Fig. )
  • Two timers

Support Protocol 1: Alternative Measurement for Hargreaves' Apparatus

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Test compartments with a mesh floor (e.g., Mesh Stand; IITC Life Science, cat. no. 410; http://www.iitcinc.com)
  • Calibrated set of von Frey hairs (e.g., Stoelting Touch Test Sensory Evaluator)

Basic Protocol 5: Acetone Evaporation Test: Innocuous Cooling and Cold Allodynia

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Test compartments with a mesh floor (e.g., Mesh Stand; IITC Life Science, cat. no. 410; http://www.iitcinc.com)
  • Automatic von Frey apparatus (e.g., Electronic von Frey IITC Life Science, cat. no. 2390; http://www.iitcinc.com/)
  • Additional reagents and equipment for habituation of mice to von Frey apparatus ( protocol 9)

Basic Protocol 6: Von Frey Test: Light Touch Perception Threshold

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Mouse restrainer (consisting of base plate, body tube, and head cover)
  • Randall‐Selitto apparatus (e.g., Analgesy‐Meter; Ugo Basile, cat. no. 37215)

Support Protocol 2: Repeated Measures Von Frey Test

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Mouse restrainer
  • Digital Paw Pressure Analgesia Instrument (IITC Life Science, cat. no. 2500)

Alternate Protocol 3: Automatic Von Frey Test

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Hamilton syringe and 29‐G needle, or 0.5‐ml disposable insulin syringe
  • Inflammatory agent (see Table 11.1.1600 for concentrations and dosage)
  • Additional reagent and equipment for determining mechanical and thermal pain thresholds (see protocols above)

Basic Protocol 7: Randall‐Selitto Test: Noxious Mechanical Pressure

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Inflammatory agent (see Table 11.1.1600 for concentrations and dosage)
  • Clear, lidded Perspex box, ∼15 cm × 15 cm
  • Mirror
  • Stopwatches
  • Hamilton syringe and 29‐G needle or 0.5‐ml disposable insulin syringe

Alternate Protocol 4: Digital Paw Pressure Test

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Anesthetic (e.g., isoflurane)
  • Isoflurane
  • Oxygen
  • Irritant (mustard oil or capsaicin)
  • Catheter (0.6 mm diameter, ∼4 cm long; e.g., Harvard Apparatus, cat. no. 732851)
  • 1‐ml syringe
  • Test compartment with mesh floor (e.g., IITC Life Science, cat. no. 410; http://www.iitcinc.com/)

Basic Protocol 8: Mechanical and/or Thermal Hyperalgesia in the Hindpaw

  • Calibrated set of von Frey hairs (e.g., Stoelting Touch Test Sensory Evaluator)

Alternate Protocol 5: Spontaneous Inflammatory Pain

  Materials
  • Mice of optimal age (6 to 8 weeks)
  • Isoflurane
  • Oxygen
  • Iodine antiseptic solution
  • Apparatus for administering isoflurane anesthesia, including muzzle for mouse
  • Heat mat
  • Surgical swabs
  • Two size‐5 forceps
  • Size‐15 scalpel
  • 3‐0 non‐absorbable suture
  • Suture needles and needle holders
  • 3‐0 absorbable suture
  • Wound clips
  • Wound clip remover

Basic Protocol 9: Intracolonic Administration of Irritants

  • Dissection microscope

Support Protocol 3: Von Frey Hair

  • Surgical swabs
  • Retractor, glass hook: surgical hooks are commercially available; however these has proven awkward to use and can result in damage to the other sciatic nerve roots—using a Bunsen burner to melt and draw out glass Pasteur pipets into fine hooks has proven a more useful tool (Fig. )
  • Dissection microscope
  • Size‐7 forceps
  • Size‐11 scalpel
  • Micro‐scissors
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Figures

Videos

Literature Cited

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