Rat Model for Evaluation of Therapeutics on Peripheral Vascular Resistance

Roseanne S. Wexler1, James L. Ellis1, Gregory P. Dubé2

1 Surface Logix, Inc., Brighton, 2 Biopure Corporation, Cambridge
Publication Name:  Current Protocols in Pharmacology
Unit Number:  Unit 5.44
DOI:  10.1002/0471141755.ph0544s35
Online Posting Date:  December, 2006
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Described in this protocol is a rat preparation that serves as a sensitive, reliable in vivo assay of peripheral vascular resistance. Experiments are conducted on conscious animals 5 days following the surgical implantation of an arterial pressure catheter in the carotid artery and a transit time flow probe on the abdominal aorta. The transit time flow probe measures volume blood flow per unit time, making it possible to calculate a true vascular resistance, a reliable indicator of microvascular tone. After allowing appropriate recovery time between experiments, it is possible to use the animal preparation multiple times. This increases study efficiency by reducing the number of animals required and makes it possible to use paired statistical analysis since multiple treatment interventions, including control conditions, can be evaluated in each animal.

Keywords: blood flow; blood pressure; flow probe; hypertension; microvascular resistance; peripheral vascular resistance; Poiseuille's law; transit time flowmetry; vascular tone; vasodilation; rat

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Table of Contents

  • Strategic Planning
  • Basic Protocol 1: Preparation of Hypertensive Rats for Measurement of Peripheral Vascular Resistance
  • Basic Protocol 2: Measurement of Peripheral Vascular Resistance in Chronically Instrumented Rats
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
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Basic Protocol 1: Preparation of Hypertensive Rats for Measurement of Peripheral Vascular Resistance

  • Rats (e.g., SHR Stroke Prone, male, 16 to 19 weeks old; Charles River)
  • Saline (0.9% NaCl; Henry Schein), sterile
  • Prazosin oral suspension (see recipe)
  • 50 mg/ml sodium pentobarbital (Henry Schein)
  • Eye lubricant (e.g., Lacrilube)
  • 70% ethanol (VWR Scientific)
  • Surgical scrub (Henry Schein)
  • H 2O, sterile
  • KY jelly or appropriate lubricant
  • 2% lidocaine (Henry Schein)
  • Lock solution (see recipe)
  • Wound glue
  • Wound‐cleaning solution (see recipe)
  • Flow probe cuffs (Transonic Systems, Inc.)
  • Drill bit (approximate diameter 5/64 in.) in an electric drill or electric rotary tool
  • Flow probes, 20‐cm lead length (Transonic Systems, MC 1.5 PRB‐JS‐WC20‐CM4S‐GA)
  • Catheters: RenaPulse pressure tubing, i.d. 0.014 in., o.d. 0.033 in. (Braintree Scientific RPT 0‐33)
  • Ink marker, black Sharpie brand
  • Clipper blade, #40 (Henry Schein)
  • Hair clippers, electric (Henry Schein)
  • Cotton swabs, gauze
  • Heating pads (electric heat source)
  • Surgical board (e.g., small cutting board from kitchen supply store)
  • Temperature sensor probe (Cole‐Parmer, model EW‐92002‐51)
  • Temperature controller, Digi‐sense brand (Cole‐Parmer, model EW‐89000‐00)
  • Surgical instruments (Fine Science Tools): scissors; fine forceps (3‐mm tip); hemostats, large for tunneling; mosquito; 3‐mm microscissors
  • Suture: 2‐0 braided silk (Henry Schein), spool, cut to 4‐in. lengths for ligatures
  • Sterile drapes: for table, body, and heating pad cover (Henry Schein)
  • 1‐ml syringes
  • Microvascular clamps, 4 (Fine Science tools 0333‐01)
  • 23‐G needles, hypodermic
  • Catheter connector, 0.022 in. o.d., SS tubing cut to 2‐cm lengths (Small Parts Inc.)
  • Luer catheter connector: blunt needle 24‐G, 0.375 in. long, 0.0223 in. o.d. (Harvard Apparatus)
  • Catheter plug, 0.022 in. o.d. (SS wire SS B‐SWGX‐220 cut to 2‐cm lengths; Small Parts Inc.)
  • Trocar: 3.2 mm o.d, 2.5 mm i.d, 6 in. skin tunneling instrument (Harvard Apparatus)
  • Wound clips (Autoclip Applier; Harvard Apparatus), and 9 mm autoclips
  • Suture: 3‐0 braided silk for internal suture, C‐13 19‐mm cutting needle
  • Suture: Ethicon prolene 2‐0, 26 mm with 1/2 or 3/8 circle needle for cuff
  • Cages with clean bedding and food pellets (individual cage for each animal)
  • Heating pads, 2 per rat, Deltaphase (thermochemical heat source; Braintree Scientific, Inc.)
  • 45° to 47°C water bath
  • Gavage needle, 18‐ or 20‐G, 1.5‐ to 2‐in. in length (Harvard Apparatus)
NOTE: Perform all surgical procedures under aseptic conditions. Sterilize (with steam autoclave) all surgical ligatures, instruments and drapes. Sterilize flow probe and catheters in CIDEX (2.4% alkaline glutaraldehyde; Henry Schein) 1 to 2 hr then rinse in sterile water three times. Wear sterile surgical gloves.

Basic Protocol 2: Measurement of Peripheral Vascular Resistance in Chronically Instrumented Rats

  • Chronically instrumented spontaneously hypertensive rat (see protocol 1)
  • Heparinized saline (see recipe)
  • Compounds (vehicle, control, and test agents)
  • Swivel system consists of the following components:
    • Tether (extension cable; Transonic Systems), modified by Plastics One (see Fig. ), 12 in. long × 4.6 mm i.d.
    • Rodent fluid swivel, 23‐G; RS P4 (Lomir Biomedical)
    • Rodent swivel tether connector; RS T1 (Lomir Biomedical)
    • Electronic commutator, SL2+2C/SB 4 channel (contact representative for custom modifications; Plastics One) and molded plugs
  • Semi‐rigid tubing, e.g. Tygon (see Fig. )
  • Velcro (see Fig. )
  • Flowmeter (Transonic Systems, Inc.; model T402)
  • Blood pressure transducer (fluid‐filled; EMKA Technologies)
  • Three‐way stopcock (VWR)
  • Rodent jacket (Braintree Scientific)
  • Data acquisition equipment: e.g., EMKA Technologies
  • Data acquisition software: e.g., EMKA IOX, EMKA Technologies
  • Data reduction software: e.g., EMKA Datanalyst, EMKA Technologies
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Literature Cited

   Drost, C.J. 1978. Vessel diameter‐independent volume flow measurements using ultrasound. In Proceedings of San Diego Biomedical Symposium, Vol. 17, pp. 299‐302. San Diego Biomedical Society, San Diego.
   Drost, C.J. 1980. Volume Flow Measurement System. U.S. Patent #4,227,407, Cornell Research Foundation, Inc., Ithaca, N.Y.
   Moore, J. and Zouridakis, G. 2004. Biomedical Technology and Devices Handbook. CRC Press, Boca Raton, Fla.
   Reneman, R.S., Hoeks, A., and Spencer, M.P. 1979. Doppler ultrasound in the evaluation of the peripheral arterial circulation. Angiology 8:526‐538.
   Ritter, M. and Ringelstein, E.B. 2002. The venturi effect and cerebrovascular ultrasound. Cerebrovasc. Dis. 14:98‐104.
   Sutera, S.P. and Skalak, R. 1993. The History of Poiseuille's Law. Annu. Rev. Fluid Mech. 25:1‐20.
   Voci, P., Pizzutoa, F., and Romeo, F. 2004. Coronary flow: A new asset for the echo lab? Eur. Heart J. 25:1867‐1879.
Key reference
   Levy, M.N. and Berne, R.M. 2000. Cardiovascular Physiology, 8th Edition. Mosby, New York.
  This reference provides an excellent overview of cardiovascular physiology including the fundamental determinants of microvascular tone that, in turn, determine peripheral vascular resistance. Discussion includes the interplay between heart function, arterial contractile function, and neurohumoral regulation of these organ systems.
Internet Resources
  Braintree Scientific. Catheters, RenaPulse pressure tubing, Deltaphase® heating pads, Rodent jacket
  Cole Parmer. Digi‐Sense Temperature Controller, heating pads
  EMKA Technologies. Data acquisition equipment and software: EMKA IOX
  Data reduction software: EMKA Datanalyst
  Fine Science Tools. Surgical instruments, Microvascular clamps
  Harvard Apparatus. Luer catheter connector, Blunt needle 24‐G, Suture, rodent jacket
  Henry Schein. Suture, Nolvasan solution, surgical supplies, hair clippers, Prazosin, Lidocaine, and pharmaceuticals
  Lomir Biomedical. Swivel, fluid
  Plastics One Inc. Swivel, electronic commutator
  Sigma‐Aldrich. Glycerol, methylcellulose
  Small Parts Inc. Stainless steel wire and tubing for catheter plugs and connectors.
  Transonic Systems Inc. Transonic flow meter and perivascular flow probes, flow probe cuff
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