Information Resources for Institutional Animal Care and Use Committees 1985-1999 *************************

Pain Management and Humane Endpoints



Endpoints in Animal Study Proposals


Guideline revised and approved by the National Institutes of Health Animal Research Advisory Committee on March 8, 2000. It is available at http://oacu.od.nih.gov/ARAC/documents/ASP_Endpoints.pdf

Introduction

Experimental studies may involve procedures that cause clinical symptoms or morbidity in animals. The Animal Care and Use Committee must consider the selection of the most appropriate endpoint(s). This requires careful consideration of the scientific requirements of the study, the expected and possible adverse effects the research animals may experience (pain, distress, illness, etc.), the most likely time course and progression of those adverse effects, and the earliest most predictive indicators of present or impending adverse effects. The effective use of endpoints requires that properly qualified individuals perform both general and study specific observations of the research animals at appropriate time points. Optimally, studies are terminated when animals begin to exhibit clinical signs of disease if this endpoint is compatible with meeting the research objectives. Such endpoints are preferable to death or moribundity as endpoints since they minimize pain and distress. Efforts must be made to minimize pain and distress experienced by animals used in research.

Morbidity

Animal Study Proposals that include morbidity as an endpoint or that include animal procedures that have the potential to cause adverse sequella should address the following:

1.Criteria that establish when the endpoint has been reached.

A. There are several examples in the literature that might be considered, including:

1.Evaluation of five aspects of an animal's condition as described by Morton and Griffiths. These are body weight, physical appearance, measurable clinical signs, unprovoked behavior and response to external stimuli.

2.Clinical observations used in cancer research and toxicological studies as described by Montgomery. Parameters include changes in general appearance, skin and hair, eyes, nose, mouth and head, respiration, urine, feces and locomotion (see table 2 on page 339).

B. The clinical signs, depending on severity and duration, that may constitute an endpoint include, but are not limited to:

C. Additional signs in neoplasia studies that may constitute an endpoint include, but are not limited to:

1. A tumor burden greater than 10% bw, and in an adult mouse, a mean tumor diameter exceeding 20 mm or in an adult rat, a mean tumor diameter exceeding 40 mm. Formulas for calculating tumor size can be found in the literature (see tumor size ref.).

2.Tumors that ulcerate, become necrotic or infected.

D.Any animal found unexpectedly to be moribund, cachectic, or unable to obtain food or water.

2.A plan for monitoring the animals both before and after a change in any of the above aspects, providing care if appropriate, and increasing the level of monitoring. Monitoring or clinical care on weekends and holidays may require involvement of the investigative staff to supplement that provided by the animal care and veterinary staff.

3.Identification of personnel responsible for evaluation, record keeping, notification of the investigator and/or veterinarian and persons responsible for euthanasia. Checklists/ score sheets may be helpful in ensuring appropriate observations are made, consistently interpreted, and properly documented.

Death or Moribundity

While it is preferable to use the earliest endpoints compatible with the scientific requirements of each study, there are studies that require moribundity or mortality as an endpoint. The moribund condition is defined as a clinically irreversible condition leading inevitably to death. In these studies, animals are permitted to die or become moribund, as a result of experimental procedures. In some cases, pain relieving measures are not used because such measures may compromise the experimental integrity of the study. Examples of research proposals that may have death or moribundity as an endpoint include: infectious disease studies, drug and toxicity studies, and cancer research. The following guidelines are suggested to assist the IC Animal Care and Use Committees in reviewing proposals with death or moribundity as endpoints.

Animal Study Proposals utilizing death or moribundity as an endpoint should contain the following information:

1.The scientific rationale for death or moribundity as an endpoint, including:

A. What alternatives were considered, why morbidity as an endpoint cannot be used, and how alternatives will be used whenever possible.

B. Why pain relieving measures cannot be utilized.

C. Number of animals to be used and why this is the minimal number of animals required.

D. Whether animals will be euthanized when moribund and if not, what information is to be gained in the interval between moribundity and death.

2. A plan for the following animal care and monitoring procedures:

A. Animals involved in experiments that may lead to moribundity or death will be monitored daily by personnel experienced in recognizing signs of morbidity (illness, injury, or abnormal behavior) for at least the following: abnormal posture, rough hair coat, head tucked into abdomen, exudate around eyes and/ or nose, skin lesions, or abnormal breathing, difficulty with ambulation, decreased food or water intake, or self mutilation.

B. The frequency of observation will be increased when animals exhibit the above or other signs of moribundity. Monitoring on weekends and holidays may require involvement of the investigative staff to supplement that provided by the animal care and veterinary staff. Designated personnel, including a veterinarian, should be notified as soon as animals show signs of disease. An assessment of the animals' condition should be made as soon as possible and a plan of action established.

C. Consideration will be given to moving animals to individual cages when their condition deteriorates to the point that injury from other animals is likely. Dead animals must be promptly removed.

D. Written records will be kept of monitoring.

General endpoint references

Canadian Council on Animal Care (1998), Guidelines on: choosing an appropriate endpoint in experiments using animals for research, teaching and testing. Ottawa, Canada.

Hendriksen and Morton, ed. (1998), Humane Endpoints in Animal Experiments for Biomedical Research. Proceedings of the International Conference, 22-25 November 1998, Zeist, The Netherlands. Laboratory Animals Ltd, by Royal Society of Medicine Press Limited, London, England.

Institute for Laboratory Animal Research Journal (2000), Humane Endpoints for Animals Used in Biomedical Research and Testing. 41: No. 2

Morton and Griffiths (1985), Veterinary Record 116:431-43.

Montgomery (1990), Cancer Bulletin 42:230-237.

Toth (1997), Contemporary Topics 36:44-48.

Stokes (1999) Humane Endpoints in Animal Experiments for Laboratory Animals Used in Toxicity Testing. Proceedings of the 3rd World Congress on Alternatives and Animal use in the Life Sciences, 31 August - 2 September 1999, Bologana, Italy.

United Kingdom Co-ordinating Committee on Cancer Research (1997), UKCCCR Guidelines for the Welfare of Animals in Experimental Neoplasia, 2nd ed. London, England.

Tumor size references

Bullard et al. (1981), J. Neuropath. Exp. Neurol. 40:410-427.

Tomayko and Reynolds (1989), Cancer Chemother. Pharmacal. 24:148-154.

Sung et al. (1993), Cancer Research 53: 2092-2099.

Welch et al. (1994), Oncogene 9:255-262.

Hamm (1995), Contemporary Topics 34:69-71.


Guidelines for the Euthanasia of Mouse and Rat Fetuses and Neonates


These guidelines were revised and reapproved on November 10, 1998 by the National Institutes of Health Animal Research Advisory Committee and are available at http://oacu.od.nih.gov/ARAC/documents/Rodent_Euthanasia_Pup.pdf

The Report of the AVMA Panel on Euthanasia does not provide specific recommendations for the euthanasia of prenatal or neonatal animals. The following guidelines are suggested to assist individual Animal Care and Use Committees at the NIH in reviewing proposals which involve the use of rodent fetuses or neonates.

Fetuses:

  1. Fetuses up to 14 days in gestation: Neural development at this stage is minimal and pain perception is considered unlikely. Euthanasia of the mother or removal of the fetus should ensure rapid death of the fetus due to loss of blood supply and non-viability of fetuses at this stage of development.
  2. Fetuses 15 days in gestation to birth: The literature on the development of pain pathways suggests the possibility of pain perception at this time. Whereas fetuses at this age are not sensitive to inhalant anesthetics, euthanasia may be induced by the skillful injection of chemical anesthetics. Decapitation with surgical scissors, cervical dislocation, or rapid freezing (immersion in liquid nitrogen) are acceptable physical methods of euthanasia. When chemical fixation of the whole fetus is required, fetuses should be anesthetized prior to immersion in or perfusion with fixative solutions. Anesthesia may be induced by hypothermia (1) of the fetus, by injection of the fetus with a chemical anesthetic, or by deep anesthesia of the mother with a chemical agent that crosses the placenta, e.g., pentobarbital. The institute veterinarian should be consulted for considerations of fetal sensitivity to specific anesthetic agents. When fetuses are not required for study, the method chosen for euthanasia of a pregnant mother must ensure rapid death of the fetus.

Neonates:

a. Up to 14 days of age: Acceptable methods for the euthanasia of neonatal mice and rats include: injection of chemical anesthetics (e.g., pentobarbital), decapitation, or cervical dislocation. Additionally, these animals are sensitive to inhalant anesthetics; e.g., methoxyflurane (used with appropriate safety considerations). Immersion in liquid nitrogen may be used only for newborns; pups older than one day should be anesthetized prior to freezing with liquid nitrogen. Similarly, anesthesia should precede immersion or perfusion with chemical fixatives. Anesthesia may be induced by inhalant or injectable anesthetics; the institute veterinarian should be consulted for appropriate agents and dosages. Alternatively, when adequately justified, hypothermia1 may be used to induce anesthesia in pups six days of age or less.

b. Older than 14 days: Follow guidelines for adults.

In all cases, the person performing the euthanasia must be fully trained in the appropriate procedures.

(1) Phifer, C.B. and Terry, L.M. 1986. Use of hypothermia for general anesthesia in preweanling rodent. Physiol. & Behav. 38:887-890.


Bibliography



Amyx, H.L. (1987). Alternatives to the LD-50 in infectious disease studies. Scientists Center for Animal Welfare Newsletter 9(4): 1-2.
NAL call number: QL55 N48
Descriptors: reducing animal numbers, preventing pain, infectious dose 50, incubation time interval assay, alternatives, humane endpoints, infectious diseases, titration of infectious agents in live animals, diagnostic tools, clinical signs of disease.

Amyx, H.L. (1987). Control of animal pain and distress in antibody production and infectious disease studies. JAVMA 191(10): 1287-1289.
NAL call number: 41.8 Am3
Descriptors: Freund's complete adjuvant, alternative adjuvants, minimization of side-effects of Freund's by careful control of injection quantity and injection site selection, rabbits, inappropriate to use Freund's in feet of rabbits, rodent footpad injections, ascites, pristane, restraint methods, animal care personnel, principles for control of pain and distress, humane endpoints, monitoring health of animals, euthanasia, analgesics, experimental design, rodents, dogs, cats, primates.

Blogg, S.L., P.P. Townsend, P.J. Butler, and E.W. Taylor (1998). A method of anaesthesia and post-operative care for experimental procedures in avian species. Animal Technology 49(2): 101-109.
NAL call number: QL55 I5
Abstract: This paper describes a successful method of anaesthesia for prolonged and/or invasive avian surgery, along with an appropriate protocol for post-operative care. These methods were determined during a neuroanatomical study of two species of bird, the tufted duck (Aythya fuligula) and the domestic duck (Anas platyrhynchos), as part of an ongoing scientific study. Descriptions of current, relevant techniques of anaethesia suitable for this study, were not readily available, therefore we felt that details of our methods would be a valuable contribution to this field. A standard operating procedure for use in similar studies and an example of an avian post-operative score sheet are outlined.

Browder, E.J. (1995). Death as an endpoint. In Current Issues and New Frontiers in Animal Research, K.A.L. Bayne, M. Greene, and E.D. Prentice, (eds.), Greenbelt, Maryland: Scientists Center for Animal Welfare, pp. 25-29.
NAL call number: HV4913 C87 1995
Descriptors: LD50, scientific justification, humane endpoints, assessment of animals, temperature drop, rectal temperature, IACUC responsibilities, investigator responsibilities, animal care staff responsibilities, funding agency responsibilities, data collection, assessment of pain and distress, selected criteria for euthanasia of moribund animals.

Canadian Council on Animal Care (1999). CCAC guidelines on: Choosing an appropriate endpoint in experiments using animals for research, education, and testing. Ottawa, Ontario,Canada: Canadian Council on Animal Care. Available at http://www.ccac.ca/Documents/Standards/Guidelines/Appropriate_endpoint.pdf
Descriptors: animal observation, significant indicators of pain and distress, scoring significant behavioral and physiological observations to set endpoints, pilot studies to determine appropriate endpoint, determining frequency of observations, defining responsibility for observations, training personnel in clinical animal observations, role of the IACUC, monoclonal antibody production, cancer research, toxicology, pain research, infectious disease studies, vaccine trials, animal models with potential for significant levels of pain and distress, species specific signs of pain and distress, information sources, understanding normal animal behavior, recognition and assessment of pain and distress, laboratory animals, fish, farm animals, examples of observational checklists used to determine endpoints.

Coenen, A.M.L., W.H.I.M. Drinkenburg, R. Hoenderken, and E.L.J.M. van Luijtelaar (1995). Carbon dioxide euthanasia in rats: oxygen supplementation minimizes signs of agitation and asphyxia. Laboratory Animals 29(3): 262-268.
NAL call number: QL55.A1L3
Descriptors: experimental methods, behavior, electroencephalogram (EEG), electrocardiogram (ECG), effect of method-animals added to a box: completely filled with CO2, into which CO2 was streamed at a high flow rate, into which CO2 was streamed at a low flow rate, into which a mixture of CO2 and O2 was streamed at a fast rate, negative aspects of CO2 euthanasia can be prevented by an additional supply of oxygen.

Close, B., K. Banister, V. Baumans, E-M. Bernoth, N. Bromage, J. Bunyan, W. Erhardt, P. Flecknell, N. Gregory, H. Hackbarth, D. Morton, and C. Warwick (1997). Working party report: Recommendations for euthanasia of experimental animals: Part 2. Laboratory Animals 31(1): 1-32.
NAL call number: QL55.A1L3
Descriptors: This document was prepared for the European Commission to be used with Directive 86/609/EEC on animal welfare, excellent review of methods of euthanasia for fish, amphibians, reptiles, birds, rodents, rabbits, carnivores (dogs, cats, ferrets), large mammals (pigs, sheep, goats, cattle, horses), nonhuman primates, less commonly used species, each section includes information on overview of the species, recognition and confirmation of death, how to euthanize larvae or embryos, physical methods, chemical methods, unacceptable methods of euthanasia.

Cunningham, J.J. and D.C. Priddy (1999). A low-cost chamber for rodent CO2 euthanasia. Lab Animal 28(2):44-45.
NAL call number: QL55.A1L33
Descriptors: euthanasia, technique, low-cost method, humane.

Danneman, P.J., S. Stein, and S.O. Walshaw (1997). Humane and practical implications of using carbon dioxide mixed with oxygen for anesthesia or euthanasia of rats. Laboratory Animal Science 47(4): 376-385.
NAL call number: 410.9 P94
Abstract: A series of studies was undertaken to determine whether CO2 can be used as a humane as well as practical agent for euthanasia or anesthesia of rats. Human volunteers rated the degree of discomfort associated with breathing 50 to 100% CO2 mixed with oxygen. Increasing concentrations of CO2 were judged as progressively more noxious, from "highly unpleasant" for 50% CO2 to "painful" for 100% CO2. The practical aspects of anesthesia and euthanasia with 50 to 100% CO2 were studied, using male Sprague Dawley rats. Time to anesthesia and death were inversely related to CO2 concentration, as were the frequency and severity of adverse reactions, including seizures and hemorrhaging from the nose. The severity of edema and hemorrhage, which were observed on histologic examination of the lungs of all rats euthanized with CO2, were greatest in the animals exposed to the lowest concentrations. There were no significant effects of CO2 concentration on time to recumbency or recovery, and there were no significant effects of precharging versus not precharging the chamber on any of the parameters studied. It was concluded that, although CO2 can be used in a humane manner, the concentrations that are least likely to cause pain and distress are associated with the longest times to anesthesia and death, highest incidence of unwanted side effects, and most severe histologic changes in the lungs. Acceptably humane and reasonably practical euthanasia or anesthesia can be achieved using a nonprecharged chamber and a low gas flow rate so that conscious animals are never exposed to CO2 concentrations >70%.

Flecknell, P. (1997). Avoidance and alleviation of pain and distress. In Animal alternatives, welfare, and ethics, Proceedings of the 2nd World Congress on Alternatives and Animal Use in the Life Sciences, held in Utrecht, the Netherlands, 20-24 October 1996, L.F.M. Zutphen and M. Balls (eds.), Amsterdam, New York: Elsevier, pp. 241-245.
NAL call number: QL1.D48 v.27
Descriptors: pain assessment, animal behavior, terminal anesthesia, analgesics, anxiolytics, humane endpoints, housing, animal husbandry.

Flecknell, P. (1997). Medetomidine and atipamezole: Potential uses in laboratory animals. Lab Animal 26(2): 21-25.
NAL call number: QL55.A1L33
Descriptors: pharmacology, clinical uses, sedation, anesthesia, tables of suggested doses for cats, dogs, gerbils, guinea pigs, ferrets, hamsters, mice, pigs, rabbits, rats, sheep, marmosets, anesthetic combinations.

Foltz, C.J. and M. Ullman-Cullere (1999). Guidelines for assessing the health and condition of mice. Lab Animal 28(4): 28-32.
NAL call number: QL55.A1L33
Descriptors: transgenic mice, knockout mice, potentially debilitating phenotypes, humane endpoints, monitoring criteria, communications between veterinary staff and investigators, body condition scoring, health guidelines, barbering, fighting, malocclusion, rectal prolapse, tumors and masses, ulcerative dermatitis, vaginal or uterine prolapse, subtle health problems - activity/behavior, anemia, dehydration, diarrhea, hypothermia, preputial or vaginal discharge, additional health problems - abnormal breathing, abnormal locomotion, eye abnormality, head tilt, hyperactivity, lethargy, paresis, paralysis, ruffled fur, tremors.

Gentle M.J. (1992). Pain in birds. Animal Welfare 1(4): 235-247.
NAL call number: HV4701.A557
Descriptors: poultry, recognition and assessment of pain, nociceptors, behavioral and physiological responses to nociceptive stimulation, pain following trauma, animal welfare, debeaking, analgesics, pain, trauma, reviews.

Hamm, T.E. Jr. (1995). Proposed institutional animal care and use committee guidelines for death as an endpoint in rodent studies. Contemporary Topics in Laboratory Animal Science 34 (3): 69-71.
NAL call number: SF405.5.A23
Descriptors: laboratory animals, endpoints, animal welfare, committees, guidelines, rodents, death, regulations.

Hellyer, P. (1998). American College of Veterinary Anesthesiologists' position paper on the treatment of pain in animals. Journal of the American Veterinary Medical Association 213 (5): 628-630.
NAL call number: 41.8 Am3
Descriptors: animal welfare, pain, symptoms, treatment, anesthesia.

Jones, D.M. (1999). Carbon dioxide induced anesthesia has no effect on brain biogenic amine concentrations in mice. Laboratory Animal Science 49(3): 316-318.
NAL call number: 410.9 P94
Descriptors: humane death, norepinephrine, dopamine, serotonin, cortex, hippocampus, striatum, cerebellum, decapitation.

Kallman, R.F., J.M. Brown, J. Denekamp, R.P. Hill, J. Kummermehr, and K.-R. Trott (1985). The use of rodent tumors in experimental cancer therapy. Conclusions and recommendations from an international workshop. Cancer Research 45: 6541-6545.
NAL call number: 448.8 C16
Descriptors: tumor/host systems, model systems, tumor immunogenicity, host animals, assay procedures, tumor growth delay assay, cellular survival assays following excision, experimental variables that can affect assay procedures or the interpretation of results, animal health and husbandry, site of tumor implantation, tumor size, stress, anesthesia, hypothermia, experimental design, radiation exposure time, cell cycle effects, drug resistance, combined modality experiments, models for adjuvant therapy, ethical principles, tumors should not exceed 10 percent of body weight.

Kastello, D.A. (1990). Recognition and alleviation of pain and distress. Symposium: Animal Welfare Compliance for Study Directors Orlando, Florida: American College of Toxicology.
NAL call number: Videocassette no. 968
Descriptors: study director, veterinarian, ACUC, pain, humane endpoints.

Keefe, F.J., R.B. Fillingim, and D.A. Williams (1991). Behavioral assessment of pain: Nonverbal measures in animals and humans. ILAR News 33(1-2): 3-13.
NAL call number: QL55.A1I43
Descriptors: excellent review of pain induction methods for analgesia testing, tail flick, hot plate, flinch-jump, pinch, formalin, chronic pain paradigms, pain measurement methods, quantification of presence of analgesia, animals, humans, devices, observation methods.

Koolhaas J.M., V. Baumans, H.J.M. Blom, D. Holst von, P.J.A. Timmermans, P.R. Wiepkema, D. Von Holst, L.F.M. Zutphen van (ed.), V. Baumans (ed.), and A.C. Beynen (1993). Behaviour, stress and well-being. In Principles of laboratory animal science: A contribution to the humane use and care of animals and to the quality of experimental results. L.F.M. van Zutphen, V. Baumans, and A.C. Beynan (eds.), Amsterdam: Elsevier Science Publishers, pp. 75-99.
NAL call number: QL55.P762 1993
Descriptors: laboratory animals, animal experiments, adaptation, learning, environment, physiology, animal behaviour, animal welfare, stress, homeostasis, phylogeny, learning, ontogeny, interaction between animal and its environment, interaction between environment and physiology, autonomic nervous system, neuroendocrine system, functional significance of physiological stress responses, pathophysiology, well-being.

Kort,W.J., J.M. Hekking-Weijma, M.T. TenKate, V. Sorm, and R. van Strik (1998). A microchip implant system as a method to determine body temperature of terminally ill rats and mice. Laboratory Animals 32(3): 260-269.
NAL call number: QL55.A1L3
Abstract: Klebsiella pneumoniae was inoculated intratracheally into rats and mice, and the temperature of the animals was recorded twice daily using microchip transponders (ELAMS) implanted either s.c. or i.p. The microchip temperatures were compared with rectal temperatures taken at the same time. The results showed that ELAMS was easy to operate and there were no important drawbacks in the use of the system were observed. The temperatures measured by the transponders implanted s.c. and i.p. did not differ significantly from rectal temperatures. In 2 out of 3 experiments on rats, it was shown that when the temperatures reached values below 36C, the median survival time of the animals was 24 h. In an experiment on mice, the same median survival time was observed at 36C. In 1 experiment using rats, however, the disease was so acute that death occurred before any temperature drop. It is suggested that when a 36 C cutoff point is used instead of the time of death in this particular animal model, the statistical analysis is not altered, and it may spare animals further suffering for approximately 24 h. It is concluded that the ELAMS system of monitoring body temperature is simple and relatively stress free for laboratory animals.
Descriptors: body temperature, monitoring, microchips, transponders, animal welfare, experimental infectious diseases.

Liles, J.H. and P.A. Flecknell (1992). The use of non-steroidal anti-inflammatory drugs for relief of pain in laboratory rodents and rabbits. Laboratory Animals 26(4): 241-255.
NAL call number: QL55.A1L3
Descriptors: classification of non-opioid non-steroidal analgesics, pharmacology of NSAIDs, assessment of analgesic efficacy, thermal stimulus, electrical stimulus, chemical methods, mechanical forces, edema tests, analgesiometry and clinical pain, effective analgesic and lethal doses of NSAIDS in mice, adverse effects of NSAIDS and selection of dose rates, analgesic, anti-inflammatory, and lethal doses in rats, ulcerogenic doses in rats, use of NSAIDS in humans and animals, suggestions for uses in laboratory animals.

Louie, A., W. Liu, Q.F. Liu, A.C. Sucke, D.A. Miller, A. Battles, and M.H. Miller (1997). Predictive value of several signs of infection as surrogate markers for mortality in a neutropenic guinea pig model of Pseudomonas aeruginosa sepsis. Laboratory Animal Science 47(6): 617-623.
NAL call number: 410.9 P94
Abstract: Infected, neutropenic animals are used as experimental models to evaluate the relative efficacies of antimicrobial agents and host-pathogen-antibiotic interactions. In the past, these models used death as the study end point. Because of the concern about use of death as an end point, we evaluated the accuracy with which various signs of infection predicted mortality in a neutropenic guinea pig model of treated and untreated Pseudomonas aeruginosa sepsis. The potential surrogate markers studied included ruffled fur, respiratory distress, diarrhea, hunched posture, lethargy, abnormal neurologic movements (twitching, paralysis of a limb), inappetence for > 48 h, the inability to ambulate, and the inability of a supine animal to stand. In addition, we evaluated whether percentage of weight loss or change in daily food and water consumption were predictive of mortality. Animals were inspected for these signs at least every 4 h during the day and every 8 h in the evening. In treated and untreated animals, 100% of subjects that were unable to ambulate or to rise from the supine position died (positive predictive value for death was 100% for either sign). Guinea pigs that could not rise from a supine position expired between 1 and 8 h after this sign was observed. Those that could not ambulate died between 4 and 40 h after that sign was observed. In treated, and untreated animals, none of the survivors manifested either sign of disease (100% specificity for each sign). However, 59% of untreated and 69% of treated animals that were ambulatory were found dead at the next observation period, underscoring the rapidity with which this infection progresses to death when it enters its final stage. No other signs of infection distinguished animals that survived or died. Thus, the inability of neutropenic, infected guinea pigs to rise from a supine position and the inability to ambulate were the only signs that accurately predicted death and, therefore, are the only signs that can be used as surrogates for death in this experimental model of P. aeruginosa sepsis.
Descriptors: animal models, Pseudomonas aeruginosa, sepsis, surrogate markers, mortality prediction, weight losses, food intake, water intake, imipenem, tobramycin, ceftazidime, antibiotics, humane endpoints.

Montgomery, C.A. (1987). Control of animal pain and distress in cancer and toxicological research. JAVMA 191(10): 1277-1281.
NAL call number: 41.8 Am3
Descriptors: experimental design, choice of rodent strain, statistical assessment of studies, animal husbandry and disease control, housing and bedding, conduct and interpretation of a study, selected clinical observations used in toxicologic studies, route of chemical administration, selected criteria for euthanasia of moribund animals, veterinary care, training.

Morton, D.B. (1998). The recognition of adverse effects on animals during experiments and its use in the implementation of refinements. In Proceedings of the Joint ANZCCART/ NAEAC conference on Ethical Approaches to Animal-based Science, D. Mellor, M. Fisher, and G. Sutherland (eds.), Adelaide, Australia and Wellington, New Zealand: ANZCCART, pp. 61-67.

Morton, D.B. (1998). The use of score sheets in the implementation of humane end points. In Proceedings of the Joint ANZCCART/ NAEAC conference on Ethical Approaches to Animal-based Science, D. Mellor, M. Fisher, and G. Sutherland (eds.), Adelaide, Australia and Wellington, New Zealand: ANZCCART, pp. 75-82.

Morton, D.B. (1997). A scheme for the recognition and assessment of adverse effects in animals. In Animal alternatives, welfare, and ethics, Proceedings of the 2nd World Congress on Alternatives and Animal Use in the Life Sciences, held in Utrecht, the Netherlands, 20-24 October 1996, L.F.M. Zutphen and M. Balls (eds.), Amsterdam, New York: Elsevier, pp. 235-240.
NAL call number: QL1.D48 v.27
Descriptors: husbandry and experimental techniques, score sheets, body weight, activity, temperature, vocalization, abdominal distension, palpate, sunken eyes, dehydration, breathing, observation, humane endpoints.

Morton, D.B. and Griffiths, P.H.M. (1985). Guidelines on the recognition of pain, distress, and discomfort in experimental animals and an hypothesis for assessment. Veterinary Record 116(16): 431-436.
NAL call number: 41.8 V641
Descriptors: signs of pain, distress, and discomfort, aids in their interpretation, appearance, food and water intake, behavior, clinical signs, cardiovascular, respiratory, digestive, nervous and musculoskeletal, temperature, edema, swelling, discharges, urinary changes, quantitative assessment of pain, distress, and discomfort, bodyweight, relationships between signs and degree of pain, distress, and discomfort, interpretation of scoring from an overall assessment, species specific clinical signs indicating pain, rats, mice, rabbits, guinea pigs, dogs, cats, primates.

National Research Council (1992). Recognition and Alleviation of Pain and Distress in Laboratory Animals. Washington, D.C.: National Academy Press, 137 p.
NAL call number: SF996.5.R43 1992
Descriptors: biologic importance of pain and distress, stressors, distress models, basis of pain, basis of stress and distress not induced by pain, recognition and assessment of pain, stress and distress, control of pain, control of stress and distress, euthanasia.

Phifer, C.B. and L.M. Terry (1986). Use of hypothermia for general anesthesia in preweanling rodents. Physiology and Behavior 38(6): 887-890.
NAL call number: QP1.P4
Descriptors: rationale and background, techniques used to induce hypothermia, deep hypothermia for surgical manipulations, moderate hypothermia as adjunct, applications.

Raufer B. and M. Miller (1997). Euthanasia: an animal-care protocol. Pork 17(2): 32-37.
Descriptors: techniques, pain, euthanasia, animal welfare.

Redgate, E.S., M. Deutsch, and S.S. Boggs. Time of death of CNS tumor-bearing rats can be reliably predicted by body weight-loss patterns. Laboratory Animal Science 41(3): 269-273.
NAL call number: 410.9 P94
Descriptors: ACUC, central nervous system, antineoplastic drug, moribund state, alternative to death as endpoint, tumor growth, neoplasms.

Rockwell, S. (1987). Maintenance of tumor systems and appropriate treatment techniques for experimental tumors. In Rodent Tumor Models in Experimental Cancer Therapy, R.F. Kallman, ed., Elmsford, New York: Pergamon Press, pp.29-36.
NAL call number: RC261.A2R62
Descriptors: choice and maintenance of animals, effect of pathogenic microorganisms, contamination of personnel, appropriate treatment techniques, choice of appropriate tumor, effects of anesthesia and restraint, tumor size-a 200mm2 (about 3/8 inch X 3/4 inch) tumor in a 20-25 (about 1 ounce) gram mouse is roughly equivalent to a 600 gram (1 pound 5 ounces) tumor in a 60-75 kg (132-165 pounds) patient, effects of stress, induction of tumor hypoxia, host-mediated effects.

Rollin, B.E. (1997). Pain and ideology in human and veterinary medicine. Seminars in Veterinary Medicine and Surgery (Small Animals) 12(2): 56-60.
Descriptors: animal welfare, philosophy, analgesics, anesthetics, pain.

Siems, J.J. and Allen, S.D. (1989). Early euthanasia as an alternative to death in chronic infectious disease studies using a systemic Candida albicans model. Abstracts of the 89th Annual Meeting of the American Society for Microbiology 1989: 81 (B-304).
NAL call number: QR1.A5
Descriptors: humane endpoints, predicting time of animal's death, multiple regression analysis, use of variables, weight loss, temperature drop, tenting-an indicator of dehydration, ataxia, rough hair coat, assessment of animal conditions throughout experiment, prevention of post-mortem change of tissues and body fluids.

Shipp, K. and B.D. Woodward (1998). A simple exsanguination method that minimizes acute pre-anesthesia stress in the mouse: evidence based on serum corticosterone concentrations. Contemporary Topics in Laboratory Animal Science 37(5): 73-77.
NAL call number: SF405.5.A23
Descriptors: stress, anesthesia, corticosterone, blood collection, inhalation anesthetics, animal welfare, carbon dioxide.

Soma, L.R. (1987). Assessment of animal pain in experimental animals. In Effective Animal Care and Use Committees, F.B. Orlans, R.C. Simmonds, and W.J. Dodds (eds.), 71-74. Special Issue of Laboratory Animal Science, Bethesda, Maryland: Scientists Center for Animal Welfare, pp.71-74.
NAL call number: QL55.E4
Descriptors: pain responses of animals, surgical procedures, postoperative pain, procedures likely to require analgesics or sedatives, clinical assessment of pain, chronic pain, chronic illness.

Soothill, J.S., Morton, D.B., and Ahmad, A. (1992). The HID-50 (hypothermia inducing dose 50): An alternative to the LD-50 for measurement of bacterial virulence. International Journal of Experimental Pathology 73: 95-98.
Descriptors: rectal temperature, temperature drop, prediction of time of death, humane endpoint, euthanasia, alternatives.

Svendsen, P. and C. Rasmussen (1998). Anaesthesia of minipigs and basic surgical techniques. Scandinavian Journal of Laboratory Animal Science 25(SUPP 1): 31-43.
NAL call number: QL55.S322
Descriptors: miniature pigs, surgery, anesthesia, methods.

Tomasovic, S.P., Coghlan, L.G, Gray, K.N., Mastromarino, A.J., and Travis, E.L. (1988). IACUC evaluation of experiments requiring death as an endpoint: A cancer center's recommendations. Lab Animal 17(1): 31-34.
NAL call number: QL55.A1L33
Abstract: IACUCs in some institutions face special problems with protocols in which death is the required end point. A possible alternative in some cases may be euthanasia when the animals become moribund.

Townsend, P. (1993). Control of pain and distress in small laboratory animals. Animal Technology 44(3): 215-223.
NAL call number: QL55.I5
Abstract: The prevention and alleviation of pain and distress in laboratory animals is presented as a strategy incorporating management, animal husbandry and scientific techniques, involving all those concerned with animal care and use, including animal technicians, veterinary surgeons and scientists. The paper identifies causes of pain and distress throughout the experimental animals life and attempts to highlight practical ways of preventing or, if this is not possible, alleviating any pain or distress which the animal may endure. All of the strategies involve making the laboratory animal the focus of concern when designing experimental schedules, as the prevention of pain and distress is the first step to producing good science.
Descriptors: distressful situations-fear, loneliness, boredom, preemptive control-planning for avoidance, refinement of techniques to minimize discomfort, alleviation of pain, anesthesia, analgesia, review of drugs, humane endpoints, euthanasia, examples of score sheets.

Toth, L.A. (1997). The moribund state as an experimental endpoint. Contemporary Topics in Laboratory Animal Science 36(3): 44-48.
NAL call number: SF405.5.A23
Descriptors: pain scoring, distress, suffering in animals, experimental paradigms, use of objective information to predict death, humane endpoints, use of experimentally relevant clinical signs, body weight, body temperature, euthanasia.

Ullman-Culleré, M.H. and C.J. Foltz (1999). Body condition scoring: A rapid and accurate method for assessing health status in mice. Laboratory Animal Science 49(3): 319-323.
NAL call number: 410.9 P94
Descriptors: noninvasive methods, criteria for euthanasia, emaciation, underconditioned, well-conditioned, overconditioned, obese, tumors, aging studies.

United Kingdom Co-ordinating Committee on Cancer Research (UKCCCR) Guidelines for the Welfare of Animals in Experimental Neoplasia (Second Edition) (1998). British Journal of Cancer 77(1): 1-10.
Descriptors: animal welfare; experimental neoplasms, research design, analgesia, animal growth and development, disease models, gene therapy, neoplasm transplantation, neoplasms physiopathology, neoplasms, therapy, humane endpoints.

Van Loo, P.L.P., L.A. Everse, M.R. Bernsen, V. Baumans, L.J. Hellebrekers, C.L.J.J. Kruitwagen, and W.den Otter (1997). Analgesics in mice used in cancer research: reduction of discomfort? Laboratory Animals 31(4): 318-325.
NAL call number: QL55.A1L3
Descriptors: animal behavior, exploration, grooming, posture, food and water consumption, buprenorphine administered in Jello, chronic pain, advanced tumors, study could not document a positive effect of buprenorphine on discomfort, possible explanations are examined.

Wallace, J., J. Sanford, M.W. Smith, and K.V. Spencer (1990). The assessment and control of the severity of scientific procedures on laboratory animals. Laboratory Animals 24: 97-130.
NAL call number: QL55.A1L3
Descriptors: definitions of severity, assessment of severity, components, index of severity, identifying and scoring pain, control of the severity of scientific procedures, development of a program for control, management practices, training, animal husbandry, humane and experimental endpoints, behavioral changes, psychosocial influences, diseases, euthanasia, sensitivity of tissues and organs to challenges, recognition and assessment of pain and distress, potential responses and physiological signs are provided for the following species: mouse, guinea pig, rabbit, golden hamster, and gerbil, comprehensive indices of severity of scientific procedures are provided for administration of substances, collection of tissue and body fluids, surgical techniques, and restraint.

Wright, A.J. and R.J. Phillpotts (1998). Humane endpoints are an objective measure of morbidity in Venezuelan encephalomyelitis virus infection of mice. Archives of Virology 143(6): 1155-1162.
NAL call number: 448.3 Ar23
Descriptors: acute infection, symptoms, clinical aspects, animal welfare, equine encephalomyelitis virus, humane endpoints.


Useful World Wide Web Sites


American Veterinary Medical Association Panel on Euthanasia Report, 2000
http://www.aphis.usda.gov/ac/euthanasia.pdf
Euthanasia methods approved by the AVMA and referenced by USDA and PHS.

Assessment of Pain and Distress in Laboratory Animals
http://www.ahc.umn.edu/rar/pain&distress.html
Provided by Research Animal Resources, University of Minnesota

That Have the Potential To Cause Severe Or Chronic Pain or Distress
http://www.iacuc.emory.edu/documents/endpoints.pdf
Provided by Emory University

Guidelines For Endpoint Monitoring And Humane Termination
http://labanimals.stanford.edu/guidelines/endpoint.html
Provided by the Dean of Research, Stanford University

Guidelines on Death as an Endpoint
http://www.upstate.edu/iacuc/pdf/euthanasia_1_deathasendpoint.pdf
Provided by Department of Laboratory Animal Resources at State University of New York-Health Sciences Center in Syracuse, NY.

IACUC and Pain-Related Internet Resources
http://netvet.wustl.edu/pain.htm
Approximately 400 web-based resources relevant to pain or IACUC's are referenced. This collection of links attempts to provide those interested with an overview of some of the best information currently available on the Internet. The categories of resources listed include: major neuroscience directories, resource references, journals, meetings, government agencies and regulations, anesthesia and analgesia, academic departments, institutional animal care and use committees, animal behavior, ethics and alternatives to animal testing, commercial resources, as well as pain societies and animal research organizations.




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Updated June 18, 2014