Regulatory Basis
The Animal Welfare Act (AWA) regulations (Code of Federal Regulations, Title 9 Subchapter A, Parts 1, 2, and 3 and Animal Care Factsheet) requires that procedures involving animals avoid or minimize discomfort, distress and pain to the animals. Furthermore, the principal investigator is required to consider alternatives to procedures that may cause more than momentary or slight pain or distress to animals, and to provide a written narrative description of methods and sources e.g. the Animal Welfare Information Center (AWIC), used to determine that alternative were not available. These requirements are explained in United States Department of Agriculture (USDA) policies 11 and 12.

The Public Health Service policy (PHS policy) requires compliance with the AWA and the Guide for the Care and Use of Laboratory Animals. Procedures with animals will avoid or minimize discomfort, distress, and pain to the animals, consistent with sound research design.

The United States Government Principles for the Utilization and Care of Vertebrate Animals in Testing, Research, and Teaching was developed by the Interagency Research Animal Committee and adopted in 1985 by the White House Office of Science and Technology Policy. These principles stipulating investigator responsibilities mandate the transport, care and use of animals in accordance with the AWA and other applicable Federal laws, regulations and policies. The animals selected for a procedure should be of an appropriate species and quality and the minimum number required to obtain valid results. Methods such as mathematical models, computer simulations, and in vitro biological systems should be considered. Proper use of animals including, avoidance or minimization of discomfort, distress and pain consistent with sound scientific practices is imperative. Essentials for Animal Research, a primer for research personnel is a useful resource for investigators

The Institutional Animal Care and Use Committee
Federal regulations mandate that the Chief Executive Officer appoint an animal care and use committee (IACUC). The ACUC is qualified through the experience and expertise of its membership to provide guidance and oversee the university's animal care and use program and to maintain compliance with applicable laws, regulations and policies. One of the most common tasks the IACUC performs is review of animal use protocols. To secure approval and to comply with Federal regulations the principal investigator should completely address ways to avoid or minimize pain and distress and demonstrate that alternative methods were not available.

Pain and distress
Painful procedure in animals means any procedure resulting pain in excess of that caused by injections or other minor procedures. Currently there is no definition for distress, however, USDA has proposed the following working definition: 'A state in which an animal can not escape from, or adapt to, the external or internal stressors or conditions it experiences, resulting in negative effects upon its well-being.'

Examples of procedures that can be expected to cause more than momentary or slight pain include, but are not limited to, the following:

• Terminal Surgery is considered a painful procedure alleviated by anesthesia.
• Freund's Complete Adjuvant used for antibody production may produce from momentary or slight pain to severe pain depending on the product, procedure and species.
• Ocular and Skin Irritancy Testing. Reaction to the product being tested may cause pain.

Examples of procedures that may cause more than momentary or slight distress include, but are not limited to, the following:

• Food or water deprivation beyond that necessary for normal pre-surgical preparation.
• Noxious electrical shock that is not immediately escapable.
• Paralysis or immobility in a conscious animal.
• The AWA and the United States Government Principles for the Utilization and Care of Vertebrate Animals in Testing, Research, and Teaching prohibit the use of paralytic agents during surgery or other painful procedures in un-anesthetized animals.

Many procedures, including any of those in the lists above, may cause both pain and distress. An example of a procedure that can be expected to cause more than momentary or slight pain as well as distress would be a study involving extensive irradiation.

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How to meet regulatory requirements for alternatives
The AWA requires the principal investigator to consider alternatives to procedures that may cause pain or distress to animals and to provide a written narrative of the methods used and sources consulted to determine the availability of alternatives. Alternatives should be considered during planning of the animal use proposal and should form part of the routine scientific literature search and not be separate from it. If a bona fide alternative method is identified, the written narrative should justify why this alternative was not used.

Alternatives should aim at avoiding or minimizing discomfort, distress or pain without compromising research goals using the 3 Rs. The 3 Rs include: replacement with no-animal systems or less sentient animal species, reduction in the number of animals required to obtain scientifically valid data through better experimental design and refinement techniques that decrease or eliminate pain or distress. All through approaches should be considered.

A narrative enables the IACUC to determine that a reasonable and good faith effort was made to determine the availability of alternatives or alternative methods. The federal requirements can be met by a combination of any of the following:

• Database search (consult more than one major database): include names of databases searched, date the search was performed, period covered by the search, keywords and/or the search strategy used.
• Attendance of colloquia or conferences.
• Consultation with subject experts, include consultant's name, qualifications and date and content of the consult.

Animal Welfare Information Center (AWIC), an information service of the National Agricultural Library, was created specifically by congress to provide information on ways to minimize pain and distress. AWIC can formulate search strategy, select key words and databases, access unique databases, train in conducting effective alternatives searches, and perform no-cost or low-cost electronic database searches. AWIC has developed an anesthesia and analgesia database that is now available on ALTWEB.

Selected Alternatives Keywords suggested by AWIC

Note: Refinement alternatives are found using terms relevant to the area of study.

ALTWEB is the premier site for obtaining information on alternatives to pain and distress. This site is being developed in collaboration with USDA and OLAW, NIH among others. ALTWEB is managed by The Johns Hopkins Center for Alternatives to Animal Testing (CAAT). Example of a sample search from ALTWEB are included.

The University of California Center for Animal Alternatives has compiled an excellent resource of alternatives information and is a must see site. Among its unique attributes are search templates that are continually updated.

Other great sources for alternatives searches are NORINA, FRAME, and NetVet. Statistical Tools is provided for your convenience. Also check the Antibody Resource Page.

For federally mandated animal testing (e.g. testing product safety, efficacy or potency) include a citation of the appropriate government agency's regulation and guidance documents. Mandating agency guidelines may provide alternatives (e.g. refinements such as humane endpoints or replacements such as the Murine Local Lymph Node Assay) that are not included in the Code of Federal Regulations. If a mandating agency-accepted alternative is not used, the principal investigator should explain the reason in the written narrative.

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