Models of Visceral Pain: Colorectal Distension (CRD)

R. Carter W. Jones1, G.F. Gebhart1

1 The University of Iowa, Iowa City, Iowa
Publication Name:  Current Protocols in Pharmacology
Unit Number:  Unit 5.36
DOI:  10.1002/0471141755.ph0536s25
Online Posting Date:  September, 2004
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Abstract

The visceromotor response to balloon distension of the colon is a robust behavioral model of visceral nociception in rodents and is ideally suited for studying the visceral antinociceptive activity of drugs. This unit describes, in detail, quantification of this response with the use of electromyography in both rats and mice.

Keywords: Electromyography; Visceromotor; Colon; Balloon; Mouse; Rat

     
 
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Table of Contents

  • Basic Protocol 1: EMG Electrode Implantation in the Rat
  • Alternate Protocol 1: EMG Electrode Implantation in the Mouse
  • Basic Protocol 2: Preparing Rats for CRD
  • Alternate Protocol 2: Preparing MICE for CRD
  • Basic Protocol 3: CRD Testing Paradigms
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: EMG Electrode Implantation in the Rat

  Materials
  • Rats (400 to 425 g)
  • 50 mg/ml pentobarbital solution
  • Teflon‐coated stainless steel wire (Cooner Wire Sales)
  • 4‐0 silk suture with curved surgical needle attached (Ethicon)
  • Animal balance, accurate to 0.1 g
  • Small animal hair clippers
  • Scalpel or sharp pair of scissors
  • Sewing needle, ∼3.0‐cm length and 1‐mm diameter
  • Long, slender forceps

Alternate Protocol 1: EMG Electrode Implantation in the Mouse

  • Mice (25 to 30 g)
  • 17.5 mg/ml ketamine/2.5 mg/ml xylazine solution
  • Fine‐tipped forceps
  • 5‐0 silk suture with curved surgical needle attached (Ethicon)

Basic Protocol 2: Preparing Rats for CRD

  Materials
  • Rats with implanted abdominal EMG electrodes (see protocol 1)
  • Lubricant
  • Saline
  • 16‐G hypodermic needle
  • Tygon tubing (0.125‐in. i.d., 0.1875‐in. o.d., 0.03125‐in. thickness; Fisher Scientific)
  • Condom, unlubricated
  • Sewing thread
  • Fabric glove (e.g., gardening glove)
  • Alligator clips
  • Tape
  • CRD testing apparatus (see protocol 5 and Fig. )
  • Gauze pads, 2 × 2–in.

Alternate Protocol 2: Preparing MICE for CRD

  Materials
  • Mice implanted with abdominal EMG electrodes (see protocol 2)
  • Halothane
  • Lubricant
  • Saline
  • Polytetrafluoroethylene (PTFE)‐24 thin‐walled tubing (0.022‐in. i.d., 0.042‐in. o.d.; Cole‐Parmer)
  • 27‐G hypodermic needle
  • Polyethylene plastic (dimensions: 3‐cm length × 3‐cm width × 15‐µm thickness)
  • PVC rod, 5‐cm long and 1‐cm diameter
  • 5‐0 silk suture (Ethicon)
  • Razor blade
  • PE 240 polyethylene tubing (0.066‐in. i.d., 0.095‐in. o.d.; Intramedic)
  • 60‐ml plastic syringe (Becton‐Dickinson)
  • Small handsaw
  • Vaporized anesthetic delivery device (Draeger)
  • Restraint device
  • Baby sock, dark‐colored
  • CRD testing apparatus (see protocol 5 and Fig. )
  • Gauze

Basic Protocol 3: CRD Testing Paradigms

  Materials
  • Rat or mouse prepared for CRD (see protocol 3 or protocol 4)
  • Compressed nitrogen tank
  • Distension device
  • EMG recording equipment
  • Computer
  • Spike2 (Cambridge Electronic Design) or comparable recording software
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Figures

  •   FigureFigure 5.36.1 Rat (A) and mouse (B) EMG recording electrodes prior to implantation (top) and depicted after implantation (middle and bottom).
  •   FigureFigure 5.36.2 Overnight inflation of rat CRD balloons increases balloon compliance. Both balloons are inflated to ∼30 mmHg. The top balloon was inflated to ∼80 mmHg during the previous 12 hr, whereas, the bottom balloon was not.
  •   FigureFigure 5.36.3 CRD balloons for rat (top) and mouse (middle with sheath and bottom without sheath in place).
  •   FigureFigure 5.36.4 The CRD testing apparatus. Spike2 software running on the computer directs opening of the solenoid valves that restrict nitrogen gas flow into the colonic balloon and records data from the EMG electrodes. The amplifier/filter (A‐M Systems), oscilloscope (Tektronix), CED 1401 and Spike2 software (Cambridge Electronic Design), and nitrogen tank pressure regulator (Matheson Tri‐gas) are commercially available. The distension controller, valves, and CRD balloons are custom‐made.
  •   FigureFigure 5.36.5 Constructing a stimulus‐response function from the EMG recordings of the VMR to CRD in mice. (A) Representative EMG records during a short baseline period, a 20‐sec distension period to pressures from 15 to 60 mmHg, and a short post‐distension period. Analysis of the raw EMG records generates values for each distension that are expressed as a percent of the maximal response, i.e., the value obtained from analysis of the EMG record during the 60 mmHg distension stimulus. (B) A stimulus‐response function is generated by plotting the percent maximal response versus distension pressure.
  •   FigureFigure 5.36.6 Representative time course of antinociceptive drug effect on the visceromotor response (VMR) to constant‐pressure CRD in the mouse (reprinted with permission from Kamp et al., ). Subcutaneous morphine dose dependently inhibits responses to phasic distension at a constant balloon pressure of 45 mmHg.

Videos

Literature Cited

Literature Cited
   Burton, M.B. and Gebhart, G.F. 1998. Effects of kappa‐opioid receptor agonists on responses to colorectal distension in rats with and without acute colonic inflammation. JPET 285:707‐715.
   Coutinho, S.V., Meller, S.T., and Gebhart, G.F. 1996. Intracolonic zymosan produces visceral hyperalgesia in the rat that is mediated by spinal NMDA and non‐NMDA receptors. Brain Research 736:7‐15.
   Danzebrink, R.M. and Gebhart, G.F. 1990. Antinociceptive effects of intrathecal adrenoceptor agonists in a rat model of visceral nociception. JPET 253:698‐705.
   Danzebrink, R.M., Green, S.A., and Gebhart, G.F. 1995. Spinal mu and delta, but not kappa, opioid‐receptor agonists attenuate responses to noxious colorectal distension in the rat. Pain 63:39‐47.
   Friedrich, A.E. and Gebhart, G.F. 2000. Effects of spinal cholecystokinin receptor antagonists on morphine antinociception in a model of visceral pain in the rat. JPET 292:538‐544.
   Gebhart, G.F. and Sengupta, J.N. 1996. Evaluation of Visceral Pain. In Handbook of Methods in Gastrointestinal Pharmacology (T.S. Ganilla, ed.) pp.359‐373. CRC Press, New York.
   Julia, V., Su, X., Bueno, L., and Gebhart, G.F. 1999. Role of neurokinin‐3 receptors on responses to colorectal distension in the rat: Electrophysiological and behavioral studies. Gastroenterology 116:1124‐1131.
   Kamp, E.H., Beck, D.R., and Gebhart, G.F. 2001. Combinations of neurokinin receptor antagonists reduce visceral hyperalgesia. JPET 299:105‐113.
   Kamp, E.H., Jones III, R.C.W., and Gebhart, G.F. 2003. Quantitative assessment and characterization of visceral nociception and hyperalgesia in mice. Am. J. Physiol. Gastrointest. Liver Physiol. 284:G434‐G444.
   Lynn, P.A. and Blackshaw, L.A. 1999. In vitro recordings of afferent fibres with receptive fields in the serosa, muscle, and mucosa of rat colon. J. Physiology 518:271‐282.
   Maves, T.J. and Gebhart, G.F. 1992. Antinociceptive synergy between intrathecal morphine and lidocaine during visceral and somatic nociception in the rat. Anesthesiology 76:91‐99.
   Ness, T.J. and Gebhart, G.F. 1988. Colorectal distension as a noxious visceral stimulus: Physiologic and pharmacologic characterization of pseudoaffective reflexes in the rat. Brain Research 450:153‐169.
   Ness, T.J. and Gebhart, G.F. 1990. Visceral pain: A review of experimental studies. Pain 41:167‐234.
   Su, X., Joshi, S.K., Kardos, S., and Gebhart, G.F. 2002. Sodium channel blocking actions of the kappa‐opioid receptor agonist U50,488 contribute to its visceral antinociceptive effects. J. Neurophysiol. 87:1271‐1279.
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