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Establishing Humane Endpoints

Euthanasia Criteria

Humane Endpoints for Studies with Expected Morbidity/Mortality

Assessing Signs of Pain and Distress

References


Overview

The importance of minimizing discomfort, pain or distress that animals may experience during the conduct of biomedical research is well recognized and is the primary force behind the animal welfare regulations governing the humane and ethical use of animals in research.

Principal investigators are responsible for considering alternatives to the use of potentially painful procedures in animals, and for determining whether these alternatives are feasible for the study. Consultation with ULAR veterinarians or other experts in the specific animal models is recommended. A discussion of alternatives to painful or distressful procedures must be included in the animal use protocol application for review by the Institutional Animal Care and Use Committee (IACUC).

Euthanasia Criteria

Most experimental animal use protocols involve the euthanasia of study animals at a predetermined endpoint, even when the animals may be clinically healthy. However, in the event that animals become ill, debilitated, or experience unrelieved pain or distress, either as a result of spontaneous disease or as a result of experimental procedures, the criteria below must be utilized in deciding whether euthanasia is the most humane option. Animals experiencing one or more of the criteria listed below must be euthanized. Exceptions are permitted only if specifically approved by the IACUC (i.e. if clinical signs listed below are expected as part of the experiment and appropriate measures are taken to minimize pain or distress). Diagnostic tests may also be used to define endpoints (e.g. vital signs, hematology, serum chemistry, imaging).

If ULAR personnel identify animals exhibiting one or more of the conditions listed below, ULAR will attempt to contact the principal investigator or another responsible person. If a responsible person cannot be located, a ULAR veterinarian will authorize euthanasia. Researchers must comply with the veterinarian’s recommendations for treatment or euthanasia.


Weight Loss

  • In adult animals, loss of > 20 percent of body weight compared to the pre-study weight or to age-matched controls
  • In growing animals, or in animals whose body weight has not been recorded, or in tumor studies, weight loss must be assessed by body condition 1,2

Decrease in appetite

  • Complete anorexia for 24 hours in small rodents, up to 5 days in large animals
  • Partial anorexia (less than 50% of normal caloric requirement) for 3 days in rodents, 7 days in large animals

Weakness/inability to obtain feed or water

Inability or extreme reluctance to stand which persists for 24 hours (assuming that the animal has recovered from anesthesia).


Moribund state

An animal found to be in a state of dying with little likelihood of recovery, showing some or all of the following symptoms:  severe depression, lack of movement; complete anorexia, hypothermia.


Infection

Infection (either overt or indicated by abnormal body temperature or WBC parameters) which fails to respond to therapy within an appropriate time.


Tumor growth

  • Solid tumors estimated to exceed 10% of normal body weight.  Assuming 1 cm3  = 1 gram, the formula for the weight of the tumor is 4/3πr3
  • Example: 25 gram mouse, tumor not to exceed 1.7 cm in diameter; 300 gram rat, tumor not to exceed 3.5 cm in diameter.
  • Loss of body condition indicating that tumor growth is being supported by body and/or metabolic resources.
  • Tumor growth that impedes an animal's ability to ingest food or water or perform other normal bodily functions or to move about the cage or remain clean and dry.
  • Tumors that appear to be causing the animal pain or distress that cannot be relieved with analgesics or other palliative measures.
  • Evidence of tumor necrosis or ulceration indicating that the tumor has outgrown its blood supply.
  • Evidence of organ dysfunction and/or failure from either the primary or metastasized tumors.

Unrelieved pain/distress

Signs of significant pain and/or distress which are unresponse to analgesics/anesthesia, or as determined by a ULAR veterinarian.


Organ system dysfunction/failure, including but not limited to:

  • Respiratory: Labored breathing, cyanosis
  • Cardiovascular: Acute blood loss resulting in shock or anemia; cardiac failure
  • Gastrointestinal:  Severe vomiting or diarrhea; rectal prolapse; intestinal obstruction
  • Urogenital:  Renal failure characterized by elevated BUN or creatinine; urinary tract obstruction; ruptured bladder; uroperitoneum; vaginal, uterine, or penile prolapse
  • Nervous:  CNS depression; frequent or continuous seizures; severe paralysis; neurological conditions which impede movement or the animals' ability to eat or drink
  • Musculoskeletal: Muscle or bone damage resulting in severe pain or inablility to use a limb.
  • Integumental: Non-healing wounds or severe burns covering more than 10% of the body; self-mutilation; severe autophagia

Humane Endpoints for Studies with Expected Morbidity or Mortality

In most studies, animals must be humanely euthanized if they experience unrelieved pain or distress, based on the Euthanasia Criteria described in the section above. Studies with death as an endpoint 6,7 (also known as survival duration studies, LD50 etc.) are not approved by the UCI Institutional Animal Care and Use Committee unless this endpoint is scientifically justified in the animal use protocol.  The justification must include, at minimum:
  1. A clear and detailed description of the clinical signs that are expected as animals enter a moribund state and approach death
  2. An explanation for why animals cannot be euthanized as soon as such signs present themselves
  3. Discussion of what additional information may be gained in the interval between the first sign of moribund symptoms and actual death
  4. The method used to determine the minimum number of animals required to achieve statistically valid results.

At all times during this process, the well-being of the research animals must be balanced against the requirements of the study. If the study is approved with expected morbidity or death as an endpoint, investigators must monitor animals at least daily, including weekends and holidays, and keep detailed records of any observations and treatments.


Assessing Signs of Pain and Distress

To assess the possibility of pain and distress objectively, it is helpful to assign values to various observations of animal condition and behavior 8,9. The attached document presents an example of a scoring system for observations of possible animal distress - 0 (normal or mild) to 3 (severe changes from normal). The cumulative score gives an indication of the likelihood that the animal is experiencing pain or distress. Humane endpoints can be established and documented in the protocol based on criteria such as:
  • The total score (e.g., a total score of >5)
  • A high score in any one variable, regardless of the total score

Depending on the particular disease model, other parameters may be added to this assessment, such as the results of diagnostic tests. 


References

1 Foltz C.J. and Ullman-Cullere, M. 1999 Guidelines for Assessing the Health and Condition of Mice. Lab Animal 28(4): 28-31.
2 Ullman-Cullere, M. and Foltz, C.J. 1999 Body Condition Scoring: A Rapid and Accurate Method for Assessing Health Status in Mice. Lab. Anim. Sci. 49(3): 319-323.
3 Workman P, A Balmain, JA Hickman et al. 1988. UKCCCR guidelines for the welfare of animals in experimental neoplasia. Laboratory Animals 22:195-201.
4 Montgomery CA. 1990. Oncologic and Toxicologic Research: Alleviation and Control of Pain and Distress in Laboratory Animals. The Cancer Bulletin 42(4):230-237.
5 Rygaard J. 1994 Animal Models in Cancer Research. In Handbook of Laboratory Animal Science, Vol. II. Boca Raton, FL: CRC Press, Inc. pp 199-205.
6 Browder EJ. 1995. Death as an Endpoint. In Current Issues and New Frontiers in Animal research. Greenbelt, MD: Scientists Center for Animal Welfare. pp 25-29.
7 Olfert ED. 1995. Defining an Acceptable Endpoint in Invasive Experiments. AWIC Newsletter 6 (1): 3-7.
8 Morton, D.B. and P.H.M. Griffiths (1985). Guidelines on the recognition of pain and discomfort in experimental animals and an hypothesis for assessment. Veterinary Record 116: 431-436.
9 ILAR website: Recognizing Pain in Animals.

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