Guidelines for establishing, implementing and refining humane endpoints.
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A key component of refining studies involving laboratory animals is the implementation of humane endpoints. The use of humane endpoints in animal experiments describes the identification of clear, predictable and irreversible criteria which substitute for more severe experimental outcomes such as advanced pathology or death. By implementing humane endpoints pain and/or distress can be prevented or alleviated whilst still meeting experimental objectives. While humane endpoints should be considered for all experiments involving animals, they are essential in studies where potential severe suffering or death may be involved (e.g. acute toxicology, animal models of infection, cancer and neurodegenerative/inflammatory diseases). It is now widely accepted that ‘death as an endpoint to a procedure should be avoided as far as possible and replaced by earlier, humane endpoints .
Body temperature , body weight , behavioural changes (e.g. reduced exploration ), pathological changes observed using imaging technology  and blood oxygen saturation  are examples of criteria that have been used successfully to implement humane endpoints. When relevant criteria are identified, actions such as humane killing, modification of the experimental design, analgesic administration and/or treatment of the animals can then be carried out to prevent or alleviate pain and/or distress.
An example of a field where a number of criteria to implement humane endpoints have been successfully identified are studies using genetically altered mice to understand the pathophysiology of Huntington’s disease and develop potential therapies . Research groups have now identified a number of predictable, objective and irreversible criteria in several strains of mice that reliably occur during neurodegenerative processes. These include decreased exploratory activity [3, 7], increased thirst and drinking , as well as changes in specific behaviours such as hanging  and jumping . These specific criteria act as substitutes for the more severe endpoints that were used in previous animal studies (typically advanced pathology or death). The implementation of humane endpoints therefore allows scientists to carry out studies to further understand and develop treatments for disease whilst alleviating pain and/or distress that may have occurred had disease progressed.
Humane endpoints are best utilised with prospective planning for their use (i.e. not used ad hoc in order to address welfare concerns as they arise). There are several stages to consider when planning and carrying out studies involving animals to establish, implement and refine humane endpoints.
Establishing humane endpoints
- In designing an experiment, the researcher should clearly specify the experimental outcome he/she wishes to achieve. Specific criteria that will allow recognition of when the experimental outcomes have been met should be identified at this stage. Non-invasive technologies, including imaging, behavioural [3, 6, 8] and, or physiological monitoring (e.g. via biotelemetry), can be useful in recognizing when experimental outcomes have been met
- It is important to recognise that it is not always necessary for an animal model to share all properties of a disease or condition in humans; it may be sufficient that the animal model is similar in some relevant aspects (e.g. using animals to study arthritis may not require chronic painful joint disease to be manifested; just an increase or decrease in urinary cartilage breakdown products to be measured)
- The researcher should also identify any potential adverse effects and the times when pain or distress may occur. He/she should then work with the veterinary surgeon (e.g. NVS), animal welfare officer (e.g. NACWO) and animal technologists, in consultation with the existing literature, to place the potential adverse effects and associated clinical signs in the context of severity classification  and to establish criteria for humane endpoints (e.g. weight loss, behavioural changes)
- In some circumstances, pilot studies using small numbers of animals may be deemed necessary to determine the onset and progress of adverse effects and to identify criteria for humane endpoint
- Validation and monitoring are required to ensure robust predictivity of the humane endpoint without interference with scientific objectives
Implementing and refining humane endpoints
- In many studies pain and distress can be avoided by identification of non-clinical humane endpoint criteria  that occur prior to any observable suffering or clinical manifestation of a condition
- When indicators of pain are identified, analgesics should be administered at a dose and by a route appropriate for the study and species
- If animals are killed at humane endpoints, the methods used should be those most appropriate for the species and age of the animal
- The use of humane endpoints should be monitored and recorded throughout the experiment, and reviewed and changed as required. This information should be included when publishing the results of the study.
- There should be appropriate training and assessment of competency for all those engaged in monitoring animals for signs of adverse effects.
- 3Rs-Centre Utrecht Life Sciences. Humane endpoints in laboratory animal experimentation.
- Newcastle University. Assessing the health and welfare of laboratory animals.
- CCAC. Guidelines on choosing an appropriate endpoint in experiments using animals for research, teaching and testing.
- OECD. Guidance document on the recognition, assessment and use of clinical signs as humane endpoints for experimental animals used in safety evaluation.
- National Research Council. Humane endpoints for animals in pain
- Directive', C., 2010-63-EU, art. 13.3.
- Ray, M.A., et al., Identification of markers for imminent death in mice used in longevity and aging research. Journal of the American Association for Laboratory Animal Science, 2010. 49(3): p. 282-288.
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- Wood, N.I., et al., Increased thirst and drinking in Huntington's disease and the R6/2 mouse. Brains Research Bulletin, 2008. 76: p. 70-79.
- Steele, A.D., et al., The power of automated high-resolution behavior analysis revealed by its application to mouse models of Huntington's and prion diseases. Proc Natl Acad Sci U S A, 2007. 104: p. 1983-1988.
- Mochel, F., et al., Altered dopamine and serotonin metabolism in motorically asymptomatic R6/2 mice. PLoS One, 2011. 6(3): p. e18336.
- RSPCA , Categorising the severity of scientific procedures on animals, J.A. Smith and M. Jennings, Editors. 2004, RSPCA Research Animals Department. p. 45.
- Hendriksen, C., K. Cussler, and D. Morton, Use of humane endpoints to minimise suffering, in The COST manual of laboratory animal care and use, B. Howard, T. Nevalainen, and G. Peratta, Editors. 2010, CRC Press: Boca Raton, Florida, USA. p. 439.