OPRR, NIH
ARENA (Applied Research Ethics National Association)
Institutional Animal Care and Use Committee Guidebook
Note from Animal Welfare Information Center editor: Every effort has been made to minimize discrepancies between the electronic and paper version of this document. Due to wordprocessing limitations, spacing, fonts, and page numbering differ between versions. The paper copy, however, is the official document.
Foreword
Acknowledgements
A. The Institutional Animal Care and Use Committee (IACUC)
A-1 Background and History
A-2 Authority, Composition and Functions
B. IACUC Review of Proposals
B-1 Fundamental Issues, Criteria
B-1-1 Approval and Disapproval of Proposals
B-2 Special Issues
B-2-1 Minimization of Pain and Distress
B-2-2 Surgery
B-2-3 Euthanasia
B-2-4 Methodology
B-2-5 Role of the Attending Veterinarian
B-2-6 Personnel Qualifications
B-2-7 Hazardous Materials
B-2-8 Field Studies
B-2-9 Death as an Endpoint
C. IACUC Oversight of Animal Care and Use Program
C-1 Policies, Procedures and Resposibilities
C-1-1 Facility and Program Review
C-1-2 Program Evaluation
C-2 Animal Health and Husbandry
C-2-1 General
C-2-2 Animal Care
C-2-3 Facility Maintenance
C-3 Occupational Health
D. IACUC Evaluation of Animal Welfare Concerns
D-1 Policies, Procedures and Reporting
E. IACUC Recordkeeping and Reporting
E-1 Policy and Procedures
E-2 IACUC Staffing
E-3 Automated Information Systems
F. Special Considerations
F-1 Numbers and Species of Animals
F-2 Alternatives to the Use of Live Animals
F-3 Instructional Use of Animals
F-4 Farm Animals
F-5 Legal Concerns
G. Resources
H. Selected References
I. Acronyms
J. Sample Forms............(Not available on electronic version)
K. Local IACUC Documents...(Not available on electronic version)
Section
Table 1. Federally Mandated IACUC Functions
Table 2. Federal Criteria for Granting IACUC Approval
Table 3. Required Contents for an Institutional Training Program
Table 4. Example Alternative Endpoints for Studies with Potential Lethality
Table 5. Occupational Health Program for Animal Handlers
Table 6. Procedures Used for Consideration of Allegations of Noncompliance
Table 7A. Federal Reporting Requirements
Table 7B. Federal Reporting Requirements
Table 7C. Federal Reporting Requirements
Table 7D. Federal Reporting Requirements
Table 7E. Federal Reporting Requirements
The role of the Institutional Animal Care and Use Committee (IACUC) in ensuring the ethical and sensitive care and use of animals in research is pivotal. The Office for Protection from Research Risks (OPRR) of the National Institutes of health has made this the cornerstone of its approach to ensuring the highest standards for animal use. Similarly, the Animal and Plant Health Inspection Service of the U.S. Department of Agriculture's approach to regulation underscores this principle. It is with this fundamental premise that the idea and focus of this Guidebook was conceived by the Council of the Applied Research Ethics National Association (ARENA), a national organization for members of IACUCS, Institutional Review Boards, Hospital Ethics Committees, and similar groups concerned with ethical and practical issues related to the conduct of research. With the endorsement and support of its parent body, Public Responsibility in Medicine and Research, ARENA authorized the project and the appointment of an Editorial Board. Distinguished individuals related to the animal care and use field were identified, and accepted the responsibility for the development of A Guidebook for Institutional Animal Care and Use Committees. At the first meeting of the Editorial Board it was agreed that the primary goals of the project would be:
1. To develop a manual for guidance of IACUCS.
2. To incorporate relevant laws and regulations into that manual.
3. To organize this (reference) manual so as to lend itself to being kept current.
Thus, although it was felt that the Guidebook would be helpful to researchers, administrators, and others related to research involving the use of animals, the focus on, and importance of the IACUC member to the process was affirmed.
The many authors, listed on page iv, made this publication possible. The Editorial Board salutes them for their most generous contribution of time, knowledge, and experience. Their quick willingness to take on the responsibility was exemplary; and all users of the Guidebook are their beneficiaries.
The decision of the OPRR to issue this Guidebook to institutions with Animal Care and Use Assurances (paralleling The Official IRB Guidebook, which was produced some years ago) is a testament to the Office's basic postulate that control of animal care and use programs should be within the institution carrying on the research. This Office's partnership in the editing, publication, and issuance of the Guidebook has been invaluable, and the informed, hands-on help of the staff of its Division of Animal Welfare has been incalculable.
It is with deep personal feeling, and on behalf of ARENA, that I express appreciation to, and honor the members of the Editorial Board. Their personal investment and untold hours of work made the fruition of this project possible.
David B. Bernhardt, Chairman Editorial Board, Institutional Animal Care and Use Committee, Guidebook
David B. Bernhardt, Chairman Jenkintown, PA
Rebecca Dresser, J.D., Professor Case-Western Reserve School of Law Cleveland, OH
Karl J. Hittelman, Ph.D. Associate Vice Chancellor, Academic Affairs University of California San Francisco, CA
Michael D. Kastello, D.V.M., Ph.D., Executive Director Research Resources ∧ Engineering Services Merck, Sharp ∧ Dohme Research Laboratories Rahway, NJ
Andrew N. Rowan, D.Phil., Director Center for Animals and Public Policy University Tufts University North Grafton, MA
Harry Rozmiarek, D.V.M., Ph.D. Professor and Director University Laboratories animal Resources University of Pennsylvania Philadelphia, PA
Ada Sue Selwitz, M.A., Director Office of Research Subjects University of Kentucky Lexington, KY
John D. Strandberg, D.V.M., Ph.D., Director Division of Comparative Medicine Johns Hopkins School of Medicine Baltimore, MD
Charles R. McCarthy, Ph.D. Director Office for Protection from Research Risks
John G. Miller, D.V.M. Director Division of Animal Welfare
Nelson L. Garnett, D.V.M. Chief, Compliance Branch
Denis Doyle Chief, Assurance Branch
Roberta Sonneborn Executive Assistant for Animal Welfare Education
Agnes Richardson Secretary to Director
William Emmett Barkley, Ph.D., Director Laboratory Safety Howard Hughes Medical Institute Bethesda, MD
Ruth M. Beckner, A.H.T. Animal Care Facility University of California San Francisco, CA
Gary G. Berntson, Ph.D., Professor Psychology, Pediatrics, and Psychiatry The Ohio State University Columbus, OH
Gordon M. Burghardt, Ph.D., Professor Department of Psychology University of Tennessee Knoxville, TN
Ruth Clark, Associate Director Office of Research Administration University of Pennsylvania Philadelphia, PA
Jeffrey M. Cohen, Ph.D. Research Compliance Officer SUNY - Albany Albany, NY
Ralph B. Dell, M.D., Chairman Institutional Animal Care and Use Committee Columbia University New York, NY
John C. Donovan, D.V.M., Director Office of Laboratory Science National Cancer Institute National Institutes of Health Bethesda, MD
Rebecca Dresser, J.D., Professor School of Law Case-Western Reserve University Cleveland, OH
James Fox, D.V.M., Ph.D., Professor and Director Division of Comparative Medicine Massachusetts Institute of Technology Cambridge, MA
Lauretta W Gerrity, D.V.M.Associate Professor Division of Comparative Medicine The University of Texas Southwestern Medical Center Dallas, TX
Alan M. Goldberg, Ph.D., Director Center for Alternatives to Animal Testing School of Hygiene and Public Health The Johns Hopkins University Baltimore, MD
Richard J. Haines, D.V.M., M.S. Department of Laboratory Animal Resources The University of Texas Health Science Center San Antonio, TX
Peter L. Jepsen, D.V M., Manager Animal Resources Department Batelle Columbus, OH
Michael D. Kastello, D.V M., Ph.D., Executive Director Research Resources ∧ Engineering Services Merck, Sharp ∧ Dohme Research Laboratories Rahway, NJ
Catherine J. Knight, J.D., Assistant Counsel Merck, Sharp ∧ Dohme Research Laboratories Rahway, NJ
Dennis F. Kohn, D.V M., Ph.D., Professor and Director Institute of Comparative Medicine Columbia University New York, NY
Joseph LaDou, M.D., Chief Division of Occupational and Environmental Medicine University of California San Francisco, CA
Neil S. Lipman, D.V.M. Division of Comparative Medicine Massachusetts Institute of Technology Cambridge, MA
Ian McCance, Ph.D., Professor Department of Physiology Monash University Clayton Victoria, Australia
Charles R. McCarthy, Ph.D., Director Office for Protection from Research Risks National Institutes of Health Bethesda, MD
Joy A. Mench, D.Phil., Associate Professor Department of Poultry Science University of Maryland College Park, MD
Sharon K. Molander, M.A.S., Administrator Division of Comparative Medicine The Johns Hopkins University School of Medicine Baltimore, MD
Charles D. Newton, D.V.M., M.S., Professor of Orthopedic Surgery and Director of Continuing Education School of Veterinary Medicine University of Pennsylvania Philadelphia, PA
Steven P. Pakes, D.V.M., Ph.D., Professor and Chairman Division of Comparative Medicine University of Texas Southwestern Medical Center Dallas, TX
Andrew N. Rowan, D.Phil., Director Center for Animals and Public Policy School of Veterinary Medicine Tufts University North Grafton, MA
Ada Sue Selwitz, M.A., Director Office of Research Subjects University of Kentucky Lexington, KY
James R Seward, M.D., M.P.P., Chief Occupational Health Service University of California Berkeley, CA
Jerald Silverman, D.V.M., Director Laboratory Animal Center The Ohio State University Columbus, OH
Alison C. Smith, D.V.M. Assistant Professor Division of Laboratory Animal Resources Medical University of South Carolina Charleston, SC
Chris S. Smith, D.V.M., Associate Director Center for Laboratory Animal Medicine and Care The University of Texas Health Science Center Houston, TX
John D. Strandberg, D.V.M., Ph.D., Director Division of Comparative Medicine The Johns Hopkins School of Medicine Baltimore, MD
W. Ray Stlicklin, Ph.D., Associate Professor Department of Animal Sciences University of Maryland College Park, MD
M. Michael Swindle, D.V.M., Director Division of Laboratory Animal Resources Medical University of South Carolina Charleston, SC
Byron Tepper, Ph.D., Director Safety and Environmental Health The Johns Hopkins University Baltimore, MD
Philip C. Tillman D.V M. Campus Veterinarian University of California Davis, CA
Joseph E. Wagner, D.V.M., Ph.D., Professor and Chairman Veterinary Pathology College of Veterinary Medicine University of Missouri Columbia, MO
Julie Watson, M.R.C.V.S. University Laboratory Animal Resources University of Pennsylvania Philadelphia, PA
William J. White, V.M.D., M.S., Director Professional Services Charles River Laboratories Wilmington, MA
Jan Wyrick, D.V.M. University of California San Francisco, CA
The Editorial Board extends their appreciation and thanks to Stephen J. Braverman, Barbara Jermyn, Joan Rachlin and Gayle Daley for their personal help and superb administrative support in ensuring the publication of this Guidebook.
A-1. Background and History
Introduction
Historically, regulations concerning the care and use of animals in laboratory research have been derived from two main sources: the experimental scientists themselves and the local humane societies formed to protect pets, farm animals and work animals from abuse. Since 1896, when the National Institutes of Health (NIH) originated, it has taken an active role in encouraging proper care and use of laboratory animals.
In the 1920s, the Director of NIH held himself personally responsible for decisions on the use of animals in any given experiment and whether the requirement for anesthesia could be waived. During the Second World War, the Committee for Medical Research was created by Franklin D. Roosevelt and, in conjunction with the National Research Council of the National Academy of Sciences, it mounted a major effort to reduce the effects of war-related disease and injury. In 1946, fifty projects from the Wartime Office of Scientific Research were transferred to NIH and the budget for the latter increased, from $180,000 in 1945 to $8,000,000 in 1947. In 1958, NIH's Division of Research Grants was reorganized and the peer review system for selecting the most meritorious grant applications was developed.
In 1963, the first edition of the Guide for the Care and Use of Laboratory Animals (Guide) was issued by the Animal Care Panel (later renamed the American Association for Laboratory Animal Science). Subsequent editions of the Guide have been developed by the National Research Council (NRC). The fifth, and most recent, edition of the Guide was published in 1985 (NIH85-23, later numbered NIH86-23). This is the primary reference for research animal care and use in the United States.
In 1966, suggestions in the press that pets were being used in research caused a public outcry and led to Congress' enacting the Pet Protection Act of 1966, the first version of what is now called the Animal Welfare Act (AWA). The United States Department of Agriculture (USDA) was given responsibility for implementing the new law. The act applied only to dogs, cats, rabbits, monkeys, guinea pigs and hamsters. Although research facilities were required to be registered, to have their suppliers licensed, and to undergo inspection by Animal and Plant Health Inspection Service (APHIS) personnel, the Act did not apply directly to the conduct of research using animals. The AWA was revised in 1970 and 1976, and underwent a major revision in December 1985. Although NIH is not responsible for enforcing the Act, requirements for compliance with it have been incorporated into any research conducted or supported by any component of the Public Health Service.
In 1973, a new policy applying to all PHS awardee institutions was drafted. This PHS Policy required compliance with AWA and the recommendations of the Guide. It also required each institution to provide NIH with an assurance which gave a detailed plan for research, training, testing, education, experimentation, or demonstration purposes. An institution's failure to comply could lead to withdrawal of NIH approval and suspension or termination of all PHS-supported research at that institution. Individual investigators could be disqualified from receiving PHS awards. Thus, this Policy required that individual institutions assume responsibility for the quality of its animal research program and the conduct of its investigators and animal care personnel. However, the very general nature of the PHS Policy made its enforcement difficult.
In 1974, the Institutional Regulations Branch of the Division of Research Grants was transferred to the Office of the Director of NIH and renamed the Office for Protection from Research Risks (OPRR). The third PHS Policy was prepared jointly by OPRR and what is now called the National Center for Research Resources, and came into effect January 1979. It covered all vertebrates used in research and the emphasis was put on the responsibility of awardee institutions to train staff for the management of their animal programs. This Policy gave institutions three options for obtaining NIH approvals: 1) accreditation by the American Association for Accreditation of Laboratory Animal Care (AAALAC); 2) an assurance that the institution's own Animal Care Committee had found the institution in full compliance with the Guide; and 3) provisional assurance of plans for correction, if deficiencies found by the Committee's annual inspection were reported to OPRR.
In July 1981, NIH issued the first comprehensive Policy which required written assurance of accreditation, either by an appropriate professional body, or by an institutional committee which included at least one veterinarian, before NIH funding could be awarded for research or teaching. The standards for evaluation were those set forward in the Guide with annual insti- tutional committee inspections.
During the 1980s the incidents of vandalism, harassment and thefts of animals increased substantially. Subsequently, Congress has manifested an increasing interest in the care and use of laboratory animals, and powerful lobbying forces have maintained this interest. A new committee was formed to revise and update the PHS Policy, concurrently with the yInstitute for Laboratory Animal Research of the National Academy of Sciences being commissioned to update the Guide. The final version of the PHS Policy, THE PUBLIC HEALTH SERVICE POLICY ON HUMANE CARE AND USE OF LABORATORY ANIMALS, was made effective January 1, 1986. It extends to foreign institutions receiving PHS funding and to intramural institutions operated directly by NIH and other agencies of PHS. The process of revision of the Policy was closely followed by Congress. Subsequent to the revision, Congress enacted the Health Research Extension Act, which added several provisions to PHS Policy. The latter was revised to conform with the new law and reissued in September 1986. Key elements of PHS Policy include:
1. Negotiation of Animal Welfare Assurances which include commitments by the awardee Institution to its animal care and use program, to appropriate staff training, and to an occupational health program for employees;
2. establishment, according to specified criteria of an Institutional Animal Care and Use Committee with defined responsibilities;
3. detailed requirements for the submission of applications for awards;
4. recordkeeping requirements to ensure clear accountability for the quality of the program; and
5. reporting requirements to enable funding agencies and OPRR to exercise oversight of the entire system.
Each institution subject to the PHS Policy is expected to operate its program in accordance with the U.S. Government Principles for the Utilization and Care of Vertebrate Animals used in Research and Training. Recently, the USDA issued Parts 1 and 2 of final regulations implementing the 1985 amendments to the AWA. Part 2, subpart C pertains to research institutions. Most of the provisions included in this subpart of the USDA Regulations are similar or identical to those included in the PHS Policy. Part 3 of the implementing USDA Regulations describes the standards which must be met when using species of animals covered by the USDA Regulations. Many of the requirements specified in Part 3 are similar to the recommendations made in the Guide and establish standards for the care and maintenance of covered species.
A-2. Authority, Composition and Functions (See Table 1)
Each institution which falls under authority of the AWA and/or receives PHS support for research and teaching involving laboratory animals must operate a program with clear lines of authority and responsibility, a properly functioning Institutional Animal Care and Use Committee (IACUC), procedures for self monitoring, adequate veterinary care, a program of occupational health, sound animal husbandry practices, and appropriate maintenance of facilities for housing animals.
The IACUC also monitors the use of animals in teaching activities as specified in the USDA Regulations, but this does not come under the Policy, unless it is supported by PHS.
The IACUC must have at least five members, including a veterinarian with program responsibilities, a scientist experienced in laboratory animal research, a non-scientist and an individual who has no other affiliation with the Institution besides membership in the IACUC. The IACUC must have the full support of the Institutional Official responsible for the program; evaluate the entire program every six months; prepare a report on the evaluation and the inspection of the facilities which is to be filed with the Institutional Official; and make recommendations to this Official concerning deficiencies, with a proposed timetable for corrections. The IACUC has the authority to suspend PHS-supported research activities.
The IACUC has an obligation to review all research projects, proposed for PHS support, prior to their receiving funding. A written report of this review confirms that the project will be conducted in accordance with PHS Policy, the Guide and the AWA. At least one member of the Committee must review each proposal, but all members must have prior opportunity to request full Committee review. The IACUC has authority to approve, require modifications before approval, or withhold approval of proposals submitted to it for review. No activity involving animals can begin unless it is first approved by the IACUC.
The frequency of IACUC consideration of approved, ongoing activities is one of the few areas in which PHS and USDA have differing requirements, i.e., PHS requires it at least once every three years, whereas USDA requires it annually. Ideally, institutions should choose to establish a uniform mechanism which satisfies both federal requirements. In deliberating this issue it is helpful to refer to consideration of ongoing activities by the use of the term "annual review" as opposed to the function of the IACUC performed at the outset of a new activity and at the expiration of an approved activity, referred to as "review." OPRR has interpreted PHS Policy to require an institutional process which provides review of proposed activities, with committee approval for a specified period of time generally not to exceed three years. This "initial renewal review" and approval may be accomplished by either convened Committee action or by a "designated reviewer/expedited review" process which meets the PHS Policy requirements of Section IV.C.2. During this period of approval, annual review must be accomplished to meet USDA requirements. The purpose of annual review is to confirm that no changes have taken place in the approved activity which might require further consideration by the IACUC, and to ensure that any new requirements of PHS, USDA or the institution are transmitted to the investigator. Annual review need not require a convened IACUC or designated reviewer/expedited action but must be adequately documented. Planned modifications must be brought to the attention of the IACUC prior or to initiation. A relatively simple mechanism to meet USDA requirements is the annual circulation of a standard form giving current basic IACUC information, e.g., approval number, date, title, species, etc., to all investigators with IACUC-approved activities. The investigator then notes that either no changes have taken place, or he/she describes any changes which have occurred. The IACUC may then separate responses, filing those indicating no changes and passing along the remainder to an IACUC-designee for assessment of the changes reported. Any changes to the approved activity which are deemed of sufficient magnitude to merit further consideration may then be presented to the IACUC. All of these dispositions should be documented as official IACUC actions.
Table I. Federally Mandated IACUC Functions
1. Review, at least once every 6 months, the research facility's program, using USDA Regulation/Guide as basis.
2. Inspect, at least once every 6 months, all of the animal facilities, including animal study areas/satellite facilities, using USDA Regulations/Guide, as basis.
3. Prepare reports of IACUC evaluations and submit the reports to the Institutional Official.
4. Review and investigate legitimate concerns involving the care and use of animals at the research facility resulting from public complaints and from reports of non-compliance received from facility personnel or employees.
5. Make recommendations to the Institutional Official regarding any aspect of the research facility's animal program, facilities or personnel training.
6. Review and approve, require modifications in (to secure approval), or withhold approval of those components of proposed activities related to the care and use of animals.
7. Review and approve, require modifications in (to secure approval), or withhold approval of proposed significant changes regarding the care and use of animals in ongoing activities.
8. Suspend an activity involving animals when necessary; take corrective action and report to funding agency and USDA.
B. IACUC Review of Proposals
B-1. Fundamental issues, Criteria (See Table 2)
Introduction
Current federal regulatory policy, as well as generally accepted ethical principles, incorporate two general goals. The first is that scientific reliance on live animals should be minimized. The second is that pain, distress, and other harm to laboratory animals should be reduced to the minimum necessary to obtain valid scientific data. Federal policy directs the IACUC to review proposals for animal use to ensure that investigators incorporate these principle into their research.
Reducing and avoiding animal use
The number of experimental animals should be the minimum necessary to produce valid results. When possible and appropriate, a non-animal substitute should be used, or a species of lower phylogenetic order substituted if available. Unnecessarily duplicative research should be avoided for scientific and ethical reasons.
The level of IACUC involvement in overseeing this process is not clear cut. The decision to require the modification of proposals to take these considerations into account requires an assessment of the necessity of a given procedure. This can be a decision for which the IACUC is inadequately equipped insofar as the Committee cannot expect to cover all aspects of scientific and technical expertise for all proposals which they will be asked to review. However, certain general questions should be answered in the proposal: the onus should be on the investigators to justify and explain their experiments.
While neither PHS Policy nor USDA Regulations explicitly prescribe an institutional mechanism to "track" animal usage by investigators under IACUC approved activities, both require that investigators include in their applications to the IACUC, identification of the appropriate number of animals to be used, and a rationale for the appropriateness of the species and number of animals they propose to use. These provisions implicitly require that institutions establish mechanisms to monitor and document the number of animals acquired and used in approved activities.
OPRR is aware of many institutions at which this mechanism precludes an investigator from using a single animal in excess of the number approved by the IACUC. Other institutions have reported mechanisms by which the number of animals acquired under an approved activity may exceed the approved number by a small percentage, e.g., 5% (this generally applies only to rodents). These institutions require investigators using non-rodent mammalian species to acquire and use only the precise number of animals approved by the IACUC.
Administrative linkage of acquisition of animals to an IACUC approval number is the method of choice employed by many institutions to "track" the number of animals being acquired under an approved activity Many also have relatively simple computerized systems which will alert the system operator and generate a report when an investigator has reached a preset per- centage, e.g., 80-90%, of the number of animals approved for the activity. This report is then submitted to the investigator with a request for specific justification for acquisition of animals in excess of the number approved. Such systems often generate animal identification or cage card labels which provide a simple mechanism to meet the required project-specific assignment of animals to corresponding approved activities. Small institutions using limited numbers of animals may choose to maintain a "hard copy" log of each IACUC-approved activity, merely subtracting the number of animals acquired with each order from the number approved, with verbal notification of the investigator as the number of animals approved is approached. Whatever mechanism an institution chooses, it must satisfy PHS Policy requirement that the use of animals be limited to the minimum necessary to obtain valid results.
Any of the above mechanisms is satisfactory in meeting the requirements of PHS Policy, the Guide and USDA Regulations.
Minimizing pain and distress (see also Section B-2-1)
Minimizing pain and distress is a basic aim of the Animal Welfare Act and the PHS Policy. This entails defining, recognizing, and reducing or eliminating these states wherever possible. The inherently subjective nature of pain makes definition difficult, but several have been proposed. For example, the International Association for the Study of Pain defines pain as "an unpleasant sensory and emotional experience associated with actual or potential tissue damage, or described in terms of such damage." The American Veterinary Medical Association's Panel report on the Colloquium on Recognition and Alleviation of Animal Pain and Distress has also formulated working definitions of pain, anxiety, fear, stress, distress, comfort, discomfort and injury. These definitions provide useful references for IACUCS.
Human observers must be able to recognize pain and distress in order to minimize them when they occur. While this may be difficult, observable signs may include departures from normal behavior or appearance and physiological parameters of a particular species or individual animal. The institution, through the IACUC, must ensure that research personnel are appropriately educated on how to assess pain and distress in their laboratory animals. This is particularly crucial for chronic experiments.
Finally, minimizing pain and distress means adopting approaches to eliminate or reduce these states when they are observed. The IACUC must develop familiarity with the most relevant of these strategies and ensure that investigators are educated in the measures applicable to their proposals. The IACUC may prohibit certain procedures altogether if it feels that pain and distress cannot be reduced to an acceptable level.
Evaluating the justification for laboratory animal use
When the IACUC reviews animal use proposals, justification questions will often arise. The need for a given number of animals is probably the most common. Certain proposals will necessarily entail pain and distress, despite all measures taken for their minimization, if the scientific question being asked is to be answered. If there is no alternative to the use of the specified animals, an evaluation of the research must be attempted. The higher the level of the anticipated distress the stronger must be the justification of the value of the research.
Often the IACUC will lack the scientific expertise to perform the relevant evaluation. Also, considerations of social value will be difficult when society itself has not made such determinations. This limits the IACUC's ability to make absolute judgments, but the committee can still have a valuable impact by simply raising the consciousness of investigators to the need for careful preparation of the justifications they supply. If justifications supplied are too general, or in insufficient detail, further elaboration should be sought by the IACUC. Additional literature reviews describing the potential con- tribution of the work in terms comprehensible to nonscientist members of the IACUC can also be requested.
The IACUC is usually able to judge the adequacy of the training and skill of the investigators proposing the research. Information on the adequacy of the equipment and facilities should also be made available. Supplemental committee members can be appointed to evaluate specific aspects of proposals submitted to the IACUC, for example, a biostatistician will often be useful either as a consultant or as a full member.
Although certain aspects of proposal review are likely to remain difficult, due to their conceptual complexity and controversial political nature, IACUC members can still do a great deal to ensure that animal research is conducted humanely and ethically in their institution.
References
AVMA Colloquium on Recognition and Alleviation of Animal Pain and Distress. November 15, 1987. JAVMA 191(10).
Pain, Anesthesia and Analgesia in Common Laboratory animals (Bibliography). 1991. Current Bibliographies in Medicine, Number 91-9, National Library of Medicine.
B-1-1. Approval and Disapproval of Proposals
Processing by the IACUC
Most IACUCs require submission of proposals in a particular format, often reflecting requirements of the Animal Welfare Act or other federal regulations. Clear presentation of a proposal will obviously assist the Committee in its deliberations. Also, preliminary discussion between the investigators and Committee members can help the former develop a proposal containing all the requisite information. Often it is helpful for a specific Committee member to be assigned a given proposal for in-depth review and liaison with the investigator. This primary reviewer is responsible for communication with the investigator and for obtaining and relaying information to and from him/her.
Another factor affecting the IACUC response is its technical expertise in areas relevant to the proposal, particularly for very complicated and involved submissions. In some instances the committee may use additional consultants to assist with specific proposals.
As a result of their review, an IACUC may approve a proposal as submitted, may reject modification, or may reject a proposal outright.
Unqualified approval: The IACUC considers that all significant points have been addressed by the investigator and that no question has been raised by any elements of the proposed study. As a result of this approval, the investigator has permission to conduct the experiments on the number of animals described in the proposal.
Deferral: This may reflect insufficient information for the IACUC to make a judgment. Absence of Committee members or other procedural reasons make it appropriate to defer a decision.
Approval pending clarification: The proposal may have no major problems but require clarification on specified minor points, signatures of responsible investigators or other administrative paperwork. The approval will be issued when the information is supplied without necessity of further discussion by the full Committee.
Qualified or conditional approval: In these cases the concerns are not major but the IACUC considers specified areas of the proposal inappropriate or problematic. The changes sought may, for example, require the substitution of different drugs or documentation demonstrating that the personnel are appropriately qualified. The approval could also be conditional on further information being provided, such as preliminary data or substantiating references from the published literature.
Limited approval: This is issued when the IACUC has more serious concerns and feels the need to follow the study more closely. This could be due to complexity of the study or the fact that it entails new and untried procedures. Thus, approval may be granted for preliminary studies with only a limited number, of animals. The initial data will then be reviewed by the Committee and the remainder of the proposal decided upon. In these instances, it is important to remember that the primary concern of the Committee is to facilitate the performance of appropriate and productive scientific endeavor, consistent with applicable laws, regulations and policies.
Disapproval: This is rare due to the on-going discussions which usually occur between investigators and Committee members. However, if the investigators refuse to modify their proposal or fail to supply information showing that their laboratory has appropriate facilities, or their staff has the necessary training to cover the concerns of the Committee, this may be the only option. A more usual result is withdrawal of the proposal.
Appeal of Committee disapproval
Most committees have procedures for addressing the situation of an investigator who chooses to resubmit a proposal which has been previously disapproved. The appeal should include the provision of additional evidence by the investigator or the solicitation of experts able to assist the Committee in their concerns. In all cases in which there is lack of Committee unanimity, the reasons for disapproval by the majority or minority membership should be presented to the investigator in writing. The Committee records should reflect this diversity of opinion.
Ultimately the IACUC must take responsibility for using animals in the research conducted at its institution. The disapproved proposals cannot be administratively approved by a higher authority. However, the opposite is not true; an IACUC approved proposal can be administratively disapproved due to financial, facility-related or other considerations.
B-2. Special Issues
B-2-1. Minimization of Pain and Distress
Definitions
For purposes of this discussion, the terms analgesia and anesthesia are defined as follows: analgesia is a state of insensibility to pain without loss of consciousness; anesthesia is a state of lack of awareness or sensitivity, with or without loss of consciousness. In research using laboratory animals, the appropriate use of anesthetic and analgesic reagents is very important for both ethical and regulatory reasons. Tranquilizers or sedatives may also be used to alleviate distress.
Assessment
All aspects of a proposed study must be thoroughly examined to determine what manipulations may cause pain or distress in the animals to be used. A judgment of the level of pain and discomfort must be made, in conjunction with an assessment of any alterative methods for obtaining the data of interest. Should there be no adequate alternative the investigator must decide what reagents can be used to reduce, or eliminate pain and distress. This will include means to assess the effectiveness of the agents and the use of established criteria for re-dosing the animal. The main difficulty is an accurate assessment of pain and distress in an animal.
Table 2. Federal Criteria for Granting IACUC Approval
| Activities | Must be in accord with USDA Regulations/PHS Policy. |
| Pain/Distress | Must avoid/minimize discomfort/distress/pain. If pain/distress is caused, appropriate sedation, analgesia or anesthesia will be used. Attending veterinarian must be involved in planning. Use of paralytic without anesthesia is prohibited. Animals with chronic/severe unrelievable pain will be painlessly killed. |
| Surgery | Must meet requirements for sterile surgery and pre/post operative care. Cannot use one animal for several major operative procedures from which it will recover, without meeting specified conditions. |
| Euthanasia | Euthanasia method must be consistent with USDA Regulations/AVMA recommendations. |
| Housing/Health | Animal living conditions must be consistent with standards of housing, feeding and care directed by veterinarian or scientist with appropriate expertise. |
| Alternatives | There must be considered alternatives to painful procedures; also must document consideration of alternatives if animals experience pain or suffering. |
| Rationale and Methods | Must provide Written narrative of methods/sources. |
| Duplication | Must provide assurance that activities do not unnecessarily duplicate previous efforts. |
| Qualifications | Personnel must be appropriately qualified. |
| Deviations from Requirements | Must be justified for scientific reasons, in writing. |
Pain is usually defined in terms of human perception of noxious stimuli and the threshold is variable among humans. Animals react similarly to the prick of a needle or the cut of a scalpel, but there is no way to know precisely how and to what degree they perceive such stimuli without imposing human standards upon the process.
Distress is even more difficult to assess. There are no generally accepted criteria for distress, although there are a number of metabolic, physiologic and behavioral parameters which are thought to be altered by it. These include changes in reproductive performance. elevation in glucocorticoid levels, and elevation in catecholamine levels.
Some experimental conditions such as prolonged restraint, alterations in the environmental temperature without provisions for behavioral or physiological adaptations, prolonged food and water deprivation as well as other procedures whose harshness or duration could be perceived by any organism as perilous, may be assumed to cause distress. While almost any condition can be perceived to cause distress, only those whose magnitude or duration are severe enough to produce significant quantifiable changes should be considered for pharmacologic intervention, providing the use of drugs does not negate the purpose of the experiment. Despite the difficulty, an investigator must attempt to incorporate some objective assessment of the level of discomfort or distress produced.
Choosing an anesthetic or analgesic
The responses of different species to different anesthetics, analgesics or tranquilizers vary and are not fully defined. Often the effects of a given drug have only been examined in a single species and definitive information, for example, on cardiovascular and respiratory function or on the ability to relieve the perception of noxious stimuli, is missing. As a result, dosages have been developed on the basis of the amount required to produce cessation of movement when the animal is con- fronted by what is assumed to be a painful manipulation, in conjunction with an adequate recovery. Because of the imprecise nature of the studies, dosage ranges are often quite wide, requiring a very conservative approach to their use. The use of drug mixtures further complicates the choice of an adequate dose. Numerous reference texts exist and IACUCs may request their veterinarian to prepare current charts of recommended doses as an institutional resource for investigators.
Summary
While the use of anesthetics and analgesics is far from simple, careful consideration to their use must be given by IACUCs and investigators. In some instances pilot studies may be necessary to assess the compatibility of drugs with the particular investigation proposed. Particularly careful justification must be required of any procedures for which alleviation of pain or distress cannot be reasonably assured. It is the responsibility of the investigator to justify the proposed study and to show that he/she has considered all the options for minimizing pain and distress.
References
AVMA Colloquium on Recognition and Alleviation of Animal Pain and Distress. November 15, 1987. JAVMA, 191(l0).
Pain, Anesthesia and Analgesia in Common Laboratory animals (Bibliography). 1991. Current Bibliographies in Medicine, Number 91-9, National Library of Medicine.
White, W. J. and K J. Field, September 1987. Anesthesia and Surgery of Laboratory Animals. Veterinary Clinics of North America: Small Animal Practice, 17(5):989-1014.
B-2-2. Surgery
Introduction
Surgery on animals is commonplace in biomedical research. It is a complex issue for IACUCs to address and for institutional veterinarians to monitor. The issue is further complicated by the multiple regulatory requirements, including qualification of personnel performing the surgery, species-specific facility and anesthetic requirements, requirements for postoperative care depending on species and operative procedure, varying requirements for major and minor surgery, and survival and non-survival procedures.
Surgery and postoperative care are addressed in the Guide, PHS Policy and USDA Regulations. These regulatory documents form the basis on which the IACUC must operate. The regulations and guidelines in the Guide and USDA Regulations are the most stringent requirements and form the basis for this discussion. Some procedures are banned or discouraged by these regulations. For example, multiple major surgical procedures may not be performed on the same animal for cost considerations, but may be performed if it is a scientifically necessary part of the proposal, has been approved by the IACUC, or if it is necessary for the health of the animal; paralytic agents may not be used without analgesia.
Facilities
The Guide provides the most stringent guidelines for surgical facilities. Aseptic facilities for non-rodent mammals should include the following: a surgical support area; a preparation area; an operating room(s); a dressing area for surgeons; and an area for animal intensive care and supportive treatment. The PHS and the AAALAC have interpreted these recommendations to mean a minimum of three rooms; an operating room used solely for that purpose, an animal preparation room and a surgeons'preparation room. The surgical support area for storage and cleaning of instruments and supplies may be combined with another area. Operating rooms should not include sinks and should contain only movable equipment such as anesthetic machines and monitoring equipment. Room surfaces should be impervious to moisture and sanitizable. If volatile anesthetics are utilized, a gas scavenging system should be provided.
With the approval of the IACUC, minor surgical procedures on non-rodent mammals may be performed without the use of a fully aseptic facility. These have been defined as procedures which do not invade a body cavity or produce permanent physiological or physical impairment. Such procedures must still be performed utilizing aseptic technique in accordance with standard veterinary procedures.
Separate facilities are not necessary for rodent surgery, and it may be performed in a laboratory or portion of a laboratory which has been sanitized and provided for that purpose. Non-survival surgery may also be performed in common laboratories. This is defined as surgery in which the animal is not allowed to recover from anesthesia.
Aseptic technique
The use of appropriate facilities must be accompanied by proper techniques to ensure the maintenance of asepsis. This includes the preparation of animals, surgeons, operating rooms, instruments and supplies, and the maintenance of asepsis during the procedures. The requirements for non-rodent mammals are more stringent than for rodents. However, adherence to aseptic techniques is required for all survival surgery regardless of species.
Intra-operative monitoring
An anesthetic proposal appropriate for the species and the procedure forms part of the submission to the IACUC. It must include the methods to be used to maintain an appropriate plane of anesthesia. Monitoring will depend on the procedure but may include body temperature, heart rate, blood pressure, blood gases and electrocardiographic activity. The personnel must be suitably qualified to cope with anesthetic and surgical complications.
For major surgical procedures on non-rodent mammals an intraoperative anesthetic monitoring record should be kept, and included with the surgeon's report as part of the animal's records. This record should be available to the personnel providing postoperative care. Some institutions provide a centralized surgical facility which is staffed by appropriately trained veterinarians and technical staff. While costly, this is probably the ideal method for ensuring that surgical complications are minimized and all regulatory requirements are met.
Postoperative care
The surgeon is responsible for ensuring that care is provided which is both appropriate to the species and to the procedure performed. The institutional veterinarian must oversee the postoperative care programs, but does not necessarily provide care unless complications arise, or his/her consultation is sought.
Components of a postoperative care program include appropriate analgesia, monitoring of surgical wounds, observation of animals for normal behavior, monitoring physiological function and recordkeeping. Special facilities may be required, depending on the procedures performed. For example, complex cardiac surgery may require the use of cages equipped with heat and oxygen, ventilation equipment, and cardiac and blood gas monitoring equipment. Other procedures may need only a technician to observe that the animal has recovered from anesthesia. Supportive fluids, analgesics and other drugs must be provided as needed. An external heat source should be available since animals often become hypothermic while under anesthesia. Postoperative records should reflect that the animal was observed until it was extubated and had recovered the ability to stand.
The postoperative period is generally considered at an end when the skin sutures are removed or the wound has healed. Until this point a minimum of recorded daily observations is needed. Consultation with a veterinarian is encouraged to ensure that adequate analgesia is provided, if necessary, throughout the postoperative period.
Personnel qualifications
The IACUC must decide whether the personnel proposing and supporting surgical proposals are adequately qualified to conduct a given procedure; and if not, how to ensure adequate instruction. In general, individuals without formal surgical training may perform surgery when qualified by experience. Various requirements may be made as to the professional background of the investigator, and documentation showing ability to perform the specific surgery may be required. Physicians trained in a surgical specialty are expected to be qualified to perform surgery on animals within the same area of surgical expertise they hold as physicians. The same assumption can be made for individuals with dental degrees. All other types of surgery will probably require additional, albeit not necessarily formal, instruction. Performing a pilot study under a supervision of an experienced person or as a part of multidisciplinary team should be sufficient. Persons with other graduate degrees, students or technicians, should only perform surgery after formal instruction in surgical techniques or with specific documentation that the proposed technique has been previously performed without complications by the person.
It is impossible to design a standard surgical instruction course which is applicable to all situations. Regulations require that each institution provide instruction in aseptic surgery applicable to their research programs. As a general rule, these courses should include instruction in aseptic technique, anesthesia and analgesia, and proper surgical technique, and may focus on a specific procedure. Cooperative multidisciplinary approaches to the instruction and conduct of the surgery are encouraged.
References
Academy of Surgical Research: Guidelines for training in surgical research in animals. 1989. J. Invest. Surg., 2(3):263-268.
Romatowski, J. 1989. Prevention and control of surgical wound infection. JAVMA, 194(l):107-114.
Swindle, M. M. and R. J. Adams (Ed.) 1988. Experimental Surgery and Physiology: Induced Animal Models of Human Disease. Baltimore: Williams and Wilkins.
White, W. J. and K. J. Field. 1987. Anesthesia and surgery of laboratory animals. Veterinary Clinics of North America: Small Animal Practice, 17:989-1017.
B-2-3. Euthanasia
Introduction
PHS Policy and USDA Regulations require that an IACUC review and approve the methods of euthanasia which are proposed. These must be consistent with the recommendations of the 1986 Report of the AVMA Panel on Euthanasia, or succeeding revised editions, unless there are scientific justifications for alterative methods.
Definition
Euthanasia means the humane killing of an animal accomplished by a method which produces rapid unconsciousness and subsequent death without evidence of pain or distress, or a method which utilizes anesthesia produced by an agent that causes painless loss of consciousness and subsequent death. Other criteria used to evaluate the appropriateness of a given method include compatibility with the requirements of the research, reliability, irreversibility, the minimization of distress to animals and persons performing euthanasia, and safety to the latter. The species of animal being used and the qualifications of the investigators are also important considerations.
Methods
Three categories of methods exist: inhalant and noninhalant pharmacologic agents, and physical methods.
Inhalant Agents:
Carbon Dioxide (CO2): Carbon dioxide is an effective and widely used agent to euthanasia rodents. This method does not cause asphyxiation; depressant and anesthetic activity occur prior to death. Since the effects of carbon dioxide are reversible it is important to ensure that the animals are dead. This can be done by performing a thoracotomy prior to discarding the carcass. Commercially filled tanks of carbon dioxide are the preferred source for C02, although with appropriate precautions against possible contact by the animals, dry ice may be used to generate the gas.
Inhalant anesthetics: Halothane, methoxyflurane and isoflurane are often used for small numbers of rodents but their expense precludes their use for larger numbers. As with carbon dioxide, death of the animal must be ensured. It is important to minimize exposure of personnel to these potentially toxic agents; therefore fume hoods must be used. Ether was formerly used extensively but its use is now dis- couraged because it is a fire hazard and potentially explosive. Similarly, the use of chloroform is discouraged as it is a potential human carcinogen.
Non-inhalant agents:
Barbiturates: These can be used to euthanasia virtually any species, but are most commonly used for non-rodents, as equally humane and less time-consuming methods are available for the latter. Most commonly, the barbiturate pentobarbital is administered by intravenous injection. With a dose of twice that required for anesthesia, unconsciousness occurs in several seconds, followed by death. In animals which are difficult to restrain, a sedative or tranquilizer may be given prior to the barbiturate. Also, the barbiturate can be administered by intraperitoneal injection, but this requires a larger and more variable dose.
T-61: This is a commercial product which has been withdrawn and should no longer be used.
Potassium Chloride (KCI): KCI induces immediate cardiac arrest without any significant depression of the central nervous system. Hence, it must be used only after the animal is deeply anesthetized.
Paralytic agents (succinyl choline, curare, etc.): These drugs induce muscular paralysis and death by suffocation. They have no direct effect on the central nervous system and they must be used only on unconscious animals.
Physical methods:
Physical methods are sometimes necessary to obtain scientifically valid data and, while aesthetically displeasing to some individuals, are entirely humane when administered under controlled conditions.
Stunning: Stunning of rodents and rabbits is accomplished by a blow to the skull of sufficient force to induce immediate unconsciousness. This is followed by decapitation or thoracotomy, with severance of a major blood vessel, to ensure death. An IACUC must be assured that only well-qualified personnel will perform this technique.
Captive Bolt Pistol: This method is used for ruminants and swine when chemical agents are scientifically contraindicated. Penetrating captive bolt pistols are more effective for inducing unconsciousness than non-penetrating pistols and should be used. To ensure death, animals should be exsanguinated subsequent to stunning.
Cervical Dislocation: This is frequently used for mice, birds, immature rats and rabbits weighing less than one kilogram. It is assumed that cervical dislocation induces immediate unconsciousness but it is preferable to sedate or lightly anesthetize the animals first. This method can also be used for hamsters and guinea pigs but is more difficult in these species, due to their muscular necks. The IACUC must be assured that the personnel are appropriately qualified in the use of this method for the specific species involved.
Decapitation: A guillotine may be used to decapitate rodents and occasionally larger species. The section should be through the atlanto-occipital joint. The 1986 AVMA Report recommended that decapitation not be done on conscious animals until further information became available. A 1988 report concluded that appropriately performed decapitation of conscious animals does not produce neurological evidence of pain or distress. This additional information may be taken into account by IACUCs, while alterative methods should always be considered.
Microwave irradiation: This method is used when a project requires fixation of brain metabolites without the loss of the anatomic integrity of the brain. Commercial microwave chambers are available for rodents which will render an animal unconscious in less than a second. These instruments differ from household units in that they direct most of the rays at the head. The kilowattage needed to induce immediate unconsciousness is proportional to the animal's size.
Exsanguination: This must not be performed on conscious animals.
Euthanasia of poikilothermic animals
The 1986 Report of the AVMA Panel on Euthanasia does not address poikilothermic animals beyond stating that methods for mammals may be inappropriate. Two useful guides for euthanasia of such species are Humane Killing of Laboratory Animals and the Canadian Guide for the Care and Use of Experimental Animals.
Chemical Agents
Intraperitoneal administration of pentobarbital is an effective method of euthanasia in amphibians, turtles and snakes. Tricaine methane sulphonate (MS222), commonly used to anesthetize frogs and toads, may be used for euthanasia. Inhalant anesthetics such as halothane may be used for amphibians and reptiles. Due to the low oxygen requirements for reptiles, the onset of unconsciousness and death will be significantly lengthened.
Physical Methods
Frogs and toads may be euthanized by stunning or pithing. Turtles may be euthanized by stunning. Decapitation is not humane due to the long period needed to induce unconsciousness by hypoxia.
References
Humane Killing of Laboratory Animals, 4th Edition. 1988. Universities Federation for Animal Welfare, Potters Bar, Herts, England.
Kitchell, R. L. 1983. Animal Pain: Perception and Alleviation. E. Carsten, et al., Eds. Bethesda, MD. Am. Physiol. Soc.
Smith, A.W., K.A. Houpt, R.L. Kitchell, D.F. Kohn, L.E. McDonald, M. Passaglia, J.C. Thurmon, and E.R. Ames. 1986. Report of the AVMA Panel on Euthanasia. JAVMA 188:252-268.
Vanderwolf, C.H., G. Buszsaki, D.P. Cain, and B. Robertson. 1988. Neocortical and hippocampal electrical activity following decapitation in the rat. Brain Research 451:340-344.
Guide to the Care and Use of Experimental Animals. 1980 and 1984. Canadian Council on Animal Care. Vols. 1 and 2.
B-2-4. Methodology
Animal preparation
All animals should exhibit normal behavior prior to entering a study. If restraint or altered conditions are needed for the study, this should be planned ahead of time so that the animal will be comfortable with the new conditions prior to actual conduct of the study.
All animals should receive a physical examination prior to being used in a study. For acute studies this need be only cursory; for more involved procedures, more extensive examinations should be conducted. It is important to determine the presence of preexisting abnormalities or conditions which would impact on the subsequent interpretation of study results.
Fluid and tissue collection
All personnel must be qualified to handle the species being used and to perform procedures necessary to collect the fluid or tissue. The method and volume of collection must be appropriate to the species, age and physical condition of the animal. Specific guidance on acceptable volumes and frequency of collection should be sought from the attending veterinarian. In instances where multiple samples are needed over a long period of time, surgical placement of indwelling catheters can facilitate sample collection.
Tissue collection via biopsy generally requires local, regional or general anesthesia, as well as appropriate aseptic techniques. The principles and procedures described in B-2-2 of this Guidebook should be adhered to under these circumstances.
Dosing and handling
Dosing with test chemicals is a routine use of laboratory animals which may result in distress due either to the nature of the chemical or the procedures involved in its administration, e.g., gavage. The IACUC may request that the veterinarian develop a chart of appropriate volumes which may be routinely administered by specified routes, and make this available to investigators preparing proposals.
Data collection
Some data collection can be accomplished by non-invasive techniques, such as arterial blood pressure monitoring by means of ultrasonic Doppler Flow Detection. Fetal development and EEG can be followed by the same techniques developed for humans.
In some instances chronic indwelling catheters or electrodes are used to monitor specific physiological parameters. The use of implantable telemetry can be preferable, as it reduces the risk of infection and damage from exposed wires.
Antibody production
Polyclonal antibody production
Of principal interest to the LACUC is the nature of the adjuvant, especially if it is Complete Freunds Adjuvant (CFA), injected with the antigen to augment the antibody response. CFA is commonly used, but it can cause severe inflammation, and can occasionally result in ulceration at the site of injection (CCAC, 1984). CFA should be used only for the initial immunization, with Incomplete Freunds Adjuvant (IFA) used for subsequent booster injections. Other adjuvants should also be considered, and CFA and IFA used only if no appropriate alternative are available. Footpad injections are strongly discouraged and frequently prohibited by institutions.
Monoclonal antibodies
The production of monoclonal antibodies is a two step process. First, an animal (usually a mouse) is immunized to generate antibody producing cells which are fused with a tumor cell line. The second step is to perpetuate the antibody secreting cells either in culture, or by injection into the peritoneum of mice to yield ascites. The yield for in vivo production of antibody is as much as two thousand fold greater per unit volume compared to in vitroproduction.
To produce ascites in mice, the animals must be primed with pristane. The volume of oil used is an important consideration. Pistane causes irritation of the peritoneum, secretion of fluid into the peritoneal cavity and suppression of the immune response to the growing tumor. The mouse is injected with approximately three million hybridoma cells and the fluid accumulating in the peritoneum collected. The pristane, the ascites tumor and the removal of the fluid are all potential sources of distress.
Chronic pathological states/disease induction
The justification for inducing chronic states must be carefully thought through and limits of allowable disease condition for the animal predetermined. A full schedule of monitoring, particularly for potentially painful conditions, must be included in any proposal presented to the IACUC. The study of naturally occurring pathological models is preferable to inducing such states artificially.
Antibody References
McGuill, M. W., and A. N. Rowan. 1989. Refinement of monoclonal antibody production and animal well-being. ILAR News 31(1):7-10.
Niemi, S. 1988. Use of immune stimulants. Pp. 119-121 in Science and animals: Addressing Contemporary Issues, H. N. Guttman, J. A. Mench, and R. C. Simmonds, Eds. Bethesda, MD: Scientists Center for Animal Welfare.
Osebold, J. W. 1982. Mechanisms of action by immunologic adjuvant. J. Am. Vet. Med. Assoc. 181:983-987.
Amyx, H. L. 1987. Control of animal pain and distress in antibody production and infectious disease studies. J. Am. Vet. Med. Assoc. 191(10):1287-1289.
B-2-5. Role of the Attending Veterinarian
Introduction
Institutions using animals for teaching and research are required by law to have an attending veterinarian associated with their animal care and use program, unless they either use only rats and mice or receive no federal funds. Institutions with small programs often opt for a veterinarian as a part-time consultant. The veterinarians overall responsibilities remain the same in all cases.
Qualifications
The veterinarian participating in a laboratory animal care and use program should have training or experience in attending to the species of animals maintained by the institution. Formal courses are now available for graduate veterinarians at a number of government, academic and commercial institutions which prepare the veterinarians for certification by the American College of Laboratory Animal Medicine (ACLAM). Alteratively, veterinarians may qualify for ACLAM certification by working in a laboratory animal resource program.
Responsibilities
The chief responsibility of the veterinarian is to provide for the health and welfare of the animals. The veterinarians must coordinate with the technical staff to ensure adequate daily animal husbandry. The details will depend on the species of animals employed and the nature of the activities in which they are used, but in all cases the care must comply with USDA Regulations and PHS Policy.
One of the prime mechanisms for ensuring high-quality laboratory animals is to purchase them from reputable vendors who produce pathogen-free stock. Generally, rodents and rabbits are purpose-bred. Certain states have passed legislation requiring that cats and dogs to be used in research be bred specifically for that purpose. Random source or wild-caught animals are not bred by the supplier but are obtained from a variety of sources, including pounds, shelters or farms which are not subject to the same standards. Before their use, clinical evaluation and conditioning of these animals is required to ensure that they are not carrying diseases which can be transmitted to other animals, including humans.
Although selection of high-quality laboratory animals has reduced the prevalence of infectious diseases in research institutions, additional preventive medical programs, conducted under the guidance of the attending veterinarian continue to be important for maintenance of healthy animals. These programs include immunization against disease-causing agents; surveillance of colonies for specific infectious microbial agents; prophylaxis utilizing pharmaceutical agents; isolation and quarantine of incoming animals; and separate housing of animals according to species and source.
While such programs are successful in reducing the incidence of disease, illness and injury still occur in laboratory animal colonies. The veterinarian is responsible for monitoring animal health and providing adequate diagnosis and treatment of animals when illness or injury dictates veterinary medical care. The veterinarian may delegate responsibility for care to trained technical staff, but he/she must always be available to provide for rapid diagnosis and treatment.
Specific areas requiring the veterinarian's attention are the utilization of suitable anesthetic and anlagesic agents; appropriate selection of species for research projects; ensuring that surgical procedures are performed properly and that pre- and post-surgical care are adequate. The veterinarian should discuss with investigators the design and implementation of their study proposals using animals and may provide written guidelines dealing with these and other issues. Consultation between the investigator and the veterinarian before submission of a proposal to the IACUC may address many of the committee's concerns and expedite the review process.
USDA Regulations require institutions utilizing animals in research and teaching to provide training and instruction to personnel on humane methods of animal maintenance and experimentation. The veterinarian and the animal resource program staff, in conjunction with the IACUC, are usually responsible for providing such training.
At some institutions, the veterinarian or his/her staff may become directly involved in activities involving animals as a coinvestigator providing clinical or surgical expertise to the study. Veterinarians may also be principal investigators and be responsible for their own research programs. In such situations, the committee has the same obligation to review and approve the proposed activities.
The Policy requires institutional occupational health programs to include personnel who work in the animal resource facilities or whose activities include substantial animal contact. The veterinarian, in cooperation with appropriate health and safety officials at the institution, is responsible for the implementation and execution of the aspects of the program which are concerned with animal health and safety issues.
The Veterinarian and the IACUC
The veterinarian's role on the IACUC is mandated by USDA Regulations and the PHS Policy. Institutions employing several veterinarians may appoint more than one to the IACUC, but all institutions regardless of size must have at least one veterinarian as a member of the IACUC.
The veterinarian should keep abreast of current literature on comparative medicine and laboratory animal science. The knowledge gained often leads to suggestions for alterative techniques, models, or species which may augment the study design and help ensure completion of the proposed study
B-2-6. Personnel Qualifications
Investigator qualifications
If valid results are to be obtained, it is important that the investigators concerned have adequate knowledge and experience in the techniques used. USDA Regulations and PHS Policy put responsibility on the research institution to ensure that all personnel involved in animal care and use are appropriately qual- ified to conduct the proposed activities.
These federal requirements have led to the need for definitive outlines and documentation of instructional methods employed by institutions. The specific programs established are dependent upon a number of factors including outside requirements, institutional philosophy, and types of studies being performed.
Development of an educational program
Program design: The institution is the legally responsible entity, but the practical development and implementation of an education program will be done by the IACUC, the veterinary staff and the investigators using animals. Program coordination is best achieved by either of the first two, but specialized procedures will rely on the latter. Outside consultants can also contribute to specific areas. The program must be flexible in design so that a heterogeneous group of investigators, technicians, stu- dents, IACUC members and veterinarians can be accommodated.
Program content: [See Table 3] The content of educational programs will be governed by legal requirements and by the specific scientific activities conducted at the institution. Certain basic procedures will be common to most programs, for example, blood sampling, injection methods, anesthesia and analgesic use. The investigators must also be familiar with the means to correct perceived deficiencies of animal care and treatment. They must be aware of information sources for optimal methods and methods to avoid unnecessary duplication of studies. Instructional methods should be designed to heighten the users' sensitivities to their animals and any potential adverse effects to these animals as a result of the procedures used. Review of proposals by the IACUC will highlight areas for inclusion in instructional programs.
Instructional methods: These can include a variety of approaches ranging from individual instruction, the provision of self-instructional materials, institutional handbooks containing basic general information, and library support including audiovisual materials and organized courses. A newsletter is also an effective way of communicating to the variety of people involved in animal care and use.
Table 3 Required Contents for an Institutional Training Program
Training and instruction of personnel must include guidance in several areas.
(1) Humane methods of animal maintenance and experimentation, including: [i] the basic needs of each species of animal; [ii] proper handling and care for the various species of animals used by the facility; [iii] proper pre- and post-procedural care of animals; and [iv] aseptic surgical methods and procedures.
(2) The concept, availability, and use of research or testing methods which limit the use of animals or minimize animal distress.
(3) Proper use of anesthetics, analgesics, and tranquilizers for any species of animal used by the facility.
(4) Methods whereby deficiencies in animal care and treatment are reported, including deficiencies reported by any employee of the facility. No facility employee, Committee member, or laboratory personnel shall be discriminated against or be subject to any reprisal for reporting violations of regulations or standards under the [Animal Welfare] Act.
(5) Utilization of services (e.g., National Agricultural Library, National Library of Medicine) available to provide information: [i] on appropriate methods of animal care and use; [ii] on alternatives to the use of live animals in research; [iii] unintended and unnecessary duplication of research involving animals; and [iv] the intent and requirements of the Act.
Source: USDA Regulations, 9 CFR Part 2, Subpart C. Section 2.31. Federal Register, August 31,1989.
Evaluation of the program: To remain useful, the program must be revised and updated to fit the needs of the individuals involved, and the legal requirements of the institution. The IACUC should monitor the instruction available and assess the capabilities of investigators and staff. The opinions of the individuals completing different components of the educational program should be solicited.
Additional instructional resources: Instruction can be assisted by programs developed for technicians at local community colleges. However, it is important to have one unit at the institution responsible for coordination and oversight of all aspects of the educational program. AALAS conducts formal training and certification programs which can form an important component of the institutional effort.
References
AALAS Training Manual Series and Instructional Guide for Technician Training. Cordova, Tennessee.
Bennett, B. T. 1990. Essentials for Animal Research: A Primer for Research Personnel. National Agricultural Library.
Education and Training in the Care and Use of Laboratory Animals. 1991. Institute of Laboratory Animal Resources. National Academic Press.
B-2-7. Hazardous Materials
Introduction
The IACUC must pay particular attention to proposals employing potentially hazardous materials, including radioactive substances, infectious microorganisms and hazardous chemicals. These all have the potential of causing harm to animals in the facility and the personnel caring for them.
Some hazardous materials are strictly controlled by federal, state and local regulations and often an institution has a specific committee to deal With all instances of their use. Radiation Safety Committees(RSCs) and Institutional Biosafety Committees(IBCs) have been mandated by the federal government to ensure that certain radioactive materials and recombinant DNA materials, respectively, are handled safely. The role of these committees is often extended to consider research involving human and animal pathogens. The IACUC should be generally familiar with the responsibilities of the various safety committees and organizations at their institution and the institution should ensure that the functions of the committees are coordinated.
In addition to the various safety committees, institutions should have professional staff or resources available with expertise in handling chemical, biological and radiological agents.
Radioactive materials
The U.S. Nuclear Regulatory Commission (USNRC) directly, or by its State designee, issues licenses permitting institutions to procure, use and dispose of specified radioactive materials. These licenses do not cover X-ray machines, high voltage accelerators, electron microscopes and radioactive materials from sources other than reactor by-products, although these are all sources of ionizing radiation. Radiation safety committees usually cover all radioactive sources. They also monitor all procurement, use and disposal of these materials; therefore, their approval should be coordinated with IACUC review of any proposal involving radioactivity. General information on potential health risks from exposure to ionizing radiation can be found in the USNRC Regulatory Guide.
Biohazardous materials
Infectious diseases may arise in many animal studies, due to natural infections as well as those specifically induced as parts of experiments. Consensus biosafety guidelines have been established for the use of animals in research involving infectious agents (Biosafety in Microbiological/and Biomedical Laboratories). These guidelines provide a concept for assessing risks and selecting appropriate safeguards. Four combinations of practices, safety equipment and facilities are described.
Recombinant DNA experiments involving animals also require approval from the IBC. The IACUC should read the report from the IBC review of facilities, procedures and practices, and expertise and training of personnel for any given proposal subsequently submitted to the IACUC.
Hazardous chemicals
In addition to animal care concerns, activities using hazardous chemicals must also deal with chemical storage and disbursement procedures, dosage preparation and challenge procedures, and waste management and disposal practices. It is also necessary to determine whether the chemicals will be present in feed, feces or urine. A rigorous review to assure appropriate safety practices, containment equipment and facility safeguards is essential for animal experiments involving chemical inhalation. A strategy for assessing risks and selecting safeguards for experiments involving chemical carcinogens has been developed by NIH.
Proposals submitted to the IACUC may, not include sufficient documentation to assess the adequacy of precautions to control exposure of personnel to the hazardous agents involved in animal experiments. The identification by the IACUC of protocols involving hazardous chemicals, e.g., the use of known carcinogens to induce tumors in animal models, determination of car- cinogenicity, mutagenicity, or teratagenicity, or acute toxicity studies, may be essential for institutional compliance with safety and health standards. The Occupational Safety and Health Administration (OSHA) laboratory standard, "Occupational Exposure to Hazardous Chemicals in the Laboratory," is of particular importance. The IACUC should be familiar with the requirement in this standard for a chemical hygiene plan for controlling exposures to hazardous chemicals. Written protocols may be required describing appropriate safety precautions and specific "designated areas" where hazardous chemicals will be used or stored.
One health and safety issue common to most IACUCs concerns the use of the inhalation agent ether for anesthesia and euthanasia. Ether forms explosive peroxide when stored in metal containers and must be used with special precautions because of its volatility and flammability. Ether must be used with special ventilation and kept away from flames or electrical ignition sources. Carcasses of ether-killed animals should be stored in well-ventilated areas and not incinerated until the ether is volatilized. Other inhalation anesthetics, e.g., halothane, methoxyflurane, nitrous oxide, although not without some degree of toxicity in an occupational setting, are less hazardous when used with proper precautions and a waste gas scavagening system. Another hazardous chemical routinely encountered in the laboratory environment is formaldehyde. Specific OSHA guidelines are available for handling formaldehyde and other chemicals. Material Safety Data Sheets, which provide useful information on specific hazardous chemicals, must be maintained on site for each hazardous agent present.
Hazardous waste
Animal wastes contaminated with radioactive materials, infectious agents or hazardous chemicals must be carefully managed to avoid human exposure or damage to the environment. Special efforts should be made in experimental design to minimize the generation of wastes containing hazardous chemicals. Those containing radioactivity in addition to hazardous chemicals are particularly difficult to deal with. Wastes containing infectious agents should be decontaminated, preferably in a steam autoclave, before disposal. Incineration is the recommended treatment for contaminated feed and bedding.
Institutional policies should be reviewed, and assistance sought from the professional health and safety staff, who have responsibility for hazardous waste management at the institution, when animal care proposals involving hazardous materials are received.
References
Biosafety in Microbiological and Biomedical Laboratories, 2nd Edition, May 1988. U.S. Department of Health and Human Services Publication No. (NIH) 88-8395.
Guidelines for Research Involving Recombinant DNA Molecules. 1985. Federal Register, Vol. 51:16958-16985.
NIH Guidelines for the Laboratory Use of Chemical Carcinogens. May 1981. U.S. Department of Health and Human Services Publication No. (NIH) 81-2385.
Occupational Exposure to Hazardous Chemicals in Laboratories. January 31, 1990. Federal Register, Vol. 55:3327-3335 29CFRI910.1450.
U.S. Nuclear Regulatory Commission Regulatory Guide. December 1980. Regulatory Guide 8.29, Instruction concerning risks from occupational radiation exposure.
B-2-8. Field Studies
Review of proposals involving field studies poses special problems for the IACUC since the federal requirements and standards focus primarily on laboratory animal care and use. Professional field biologists in organizations devoted to the study of fish, amphibians, reptiles, birds, and mammals have recently prepared guidelines for field work with these animals, and they form a useful reference. The report of a recent work- shop sponsored by the National Science Foundation is also of value.
Unique concerns in the field include the impact of a given proposal on the native populations of animals at the experimental site. This can be due to the use of enclosures, or, more indirectly, the effect of changed behavior of the study animals on other populations in the vicinity.
The proposed study can be assessed by the IACUC according to a number of priority questions which are similar to those used for a laboratory-based proposal:
Species selection: The most common and least sensitive species should be studied in preference to rare or endangered ones, or ones known to be particularly timid and susceptible to distress. Consideration should be given to whether the population is stable, growing, declining or marginal. If either of the latter is evident, careful attention must be given to the potential impact of the proposal.
Site selection: Many animals live in a variety of habitats and the choice of a study site conducive to obtaining maximal results with minimal disruption should be given priority. Also, the impact of other human enterprises on the site should be considered, for example, agriculture, tourists, hunting, and fishing may all disrupt a study. The ownership of the site, and whether permits are required to gain access or to conduct experiments, are also crucial considerations. The investigator must be able to assure the IACUC that all the necessary permits have been obtained.
Methodologies to be employed: If animals are to be captured, methods and numbers should be detailed in the proposal. Measures taken to alleviate distress and injuries should be described. Some discussion of the potential impact of capture on the subsequent behavior of the animals should also be included. If the animals are to be monitored individually, the investigators must indicate whether they plan to follow the animals by their natural markings or whether they will be artificially marked. If the latter, a clarification of methodology and potential trauma is necessary; e.g., paint markings may increase visibility to predators. Often the fact that a method has been used traditionally is presented as a justification; for non-endangered populations and if minimal distress is involved this is usually adequate.
Site manipulations may include the removal of prey, predators, or the addition of either. The latter is rarely justified if non-native species are involved. It may be proposed to erect fences to limit the movement of populations. Individual animals may be treated to alter their behavior by surgery or drugs, or their markings changed to assess responses by the community. Individual animals may be periodically removed to take tissue samples. When these individuals are meant to survive, aseptic practices should always be employed for such surgical procedures. In all of these instances any potential pain or distress to an individual animal must be assessed and the investigator's justification evaluated in the context of the value of the data to be obtained. Consideration should also be given to additional data that might be obtained at no further cost. Potential long term effects on individual animals, their community, and other species in the vicinity must also be evaluated.
Clearly many of these questions are difficult to answer definitively, but their consideration will help the IACUC judge the potential impact, as well as the potential value, of the study proposed. Such questions can be expected to assist the investigator in obtaining maximum information from his study with minimum negative environmental impact.
References
Acceptable field methods in mammalogy: Preliminary guidelines approved by the American Society of Mammalogists. 1987. Journal of Mammalogy, 68(4, Suppl.): 1-18.
Bowman, P. 1989. Institutional animal care and use committee review of wildlife field research. Lab Animal, 18(3):28-30.
Burghardt, Z. M. and H. A. Herzog, Jr. 1980. Beyond conspecifics: Is Brer Rabbit our brother? Bioscience, 30:763-768.
Guidelines for the use of fish in field research. 1987. American Society of Ichthyologists and Herpetologists (SIH), American Fisheries Society (AFS), and the American Institute of Fisheries Research Biologists (AIFRB). Copeia, (Suppl.):1-12.
Guidelines for the use of live amphibians and reptiles in field research. 1987. American Society of Ichthyologists and Herpetologists (ASIH), The Herpetologists' League (HL), and the Society for the Study of Amphibians and Reptiles (SSAR). Journal of Herpetology, 4(Suppl.): 1-14.
Orlans, F. B., Ed. 1988. Field research guidelines: Impact on animal care and use committee. Bethesda, MD. Scientists Center for Animal Welfare.
Report of committee on the use of wild birds in research. 1988. American Ornithologist's Union. Auk, 105(1, Suppl.):1A-41A.
B-2-9. Death as an Endpoint (See Table 4)
Background
Since it provides an objective and unequivocal data point, death has been used as an endpoint in cancer, infectious disease and other animal studies, especially for regulatory purposes (e.g., drug safety/efficacy studies). Increased public interest and regulation have led to a reevaluation of its appropriateness. Much of the current concern arises from the use of LD50 tests for chemicals and drugs to determine acute toxicity measures. In the main, euthanasia provides tissues more appropriate for subsequent study and alleviates potential suffering by the animal. Hence, euthanasia is often feasible scientifically, as well as ethically
The routine use of death as an endpoint should be discouraged. Endpoints other than death must always be considered and should be used whenever the research objective makes it possible. Use of death as an endpoint must be justified in writing in the proposal and its use must be approved by the IACUC prior to beginning a study. Drugs or techniques to alleviate pain or distress preceding death must be used unless they would interfere with the scientific objectives of the study. A proposal foregoing the use of anesthetics, analgesics or tranquilizing drugs must be extensively justified to the IACUC.
Alternative endpoints to death for studies with potentially lethal outcomes
In many types of studies where lethality was previously considered essential, alterative endpoints are now proving to be more scientifically sound and ethically acceptable. Factors to consider include percentage loss of body weight, respiratory changes, cardiovascular alterations, neural indicators, inflammation, etc.
Where survival data are required, euthanizing animals when they become moribund has proven useful. This requires the investigators to define a moribund state prior to initiating the study They must also ensure that throughout the study the personnel monitoring the animals have appropriate qualifications and choose sufficiently frequent observation points. The latter will be determined by the species, the onset, severity and duration of expected clinical signs, and other specific characteristics of the agent being tested.
Examples of studies involving potential lethality
Anti-neoplastic drugs frequently have toxic side effects. This poses a particular problem, as the drugs are normally administered close to their toxic dose in order to demonstrate their effectiveness. Whether imminent death is due to the cancer or the drug can be difficult to determine and can make euthanasia inappropriate.
Lethal, whole body irradiation, used to render the immune system incapable of rejecting grafts, seriously reduces an animal's ability to deal with infection, and causes dose-dependent radiation injury. Defined predeath endpoints can be impossible when only subtle differences exist between control and experimental groups.
Similar considerations should be given to acute toxicity tests for drugs and chemicals and to studies of infectious diseases which may cause a variety of lethal outcomes. They do not invariably require death, and sampling prior to death may actually reveal more information. In all cases alterative endpoints should be considered, as well as a careful evaluation of the minimum number of animals necessary for the study.
References
American College of Toxicology, Policy Statement: Care and Use of animals in Toxicology. Summer 1988. American College of Toxicology Newsletter, Vol. 8, No. 2.
Amyx, H. L. Winter 1987. Alternatives to the LD50 in Infectious Disease Studies. Scientists Center for Animal Welfare Newsletter.
Canadian Council on Animal Care, Ethics and Animal Experimentation. Revised, May 1988.
Hayes, A. W., Ed. 1989. Principles and Methods of Toxicology. Raven Press, New York, 2nd Ed.
Kallman, R. F., Ed. 1987. Rodent Tumor Models in Experimental Cancer Therapy. Pergamon Press.
Montgomery, C. A. November 15, 1987. Control of Animal Pain and Distress in Cancer and Toxicologic Research. JAVMA, Vol. 191, No. 10.
Tomasovic, S. P., et al. January/February 1988. IACUC Evaluation of Experiments Requiring Death as an End Point: A Cancer Center's Recommendations. Lab Animal.
U.S. Environmental Protection Agency, Revised Policy for Acute Toxicity Testing. 1988.
Workman, P., et al. 1988. UKCCCR Guidelines for the Welfare of Animals in Experimental Neoplasia. Laboratory Animals, 22:195--201.
Table 4 Example Alternative Endpoints for Studies with Potential Lethality
Alterative Endpoint: Tumor Characteristics
Example: 10% of normal body weight, necrosis, infection
Application: Subcutaneous or intraperitoneal tumors and hybridomas
Alternative Endpoint: Peripheral Blood Cell Counts
Example: Depends on cell type
Application: Leukemias, infectious disease, anemia
Alternative Endpoint: Prolonged Inappetence/Cachexia
Example: Loss of weight (20% of normal body weight) and/or condition
Application: Metastatic disease, chronic infectious disease
Alternative Endpoint: Inability to Obtain Feed and Water
Example: Paralysis, oro-facial or cervical lesions, other non-ambulatory condition
Application: Many
Alternative Endpoint: Signs of Severe Organ or System Involvement
Example: Respiratory: rapid or labored breathing, coughing, rales
Cardiovascular: shock, hemorrhage, anaphylaxis
Gastrointestinal: severe diarrhea or vomiting
Peripheral Nervous System: flaccid or spastic paralysis
CNS Signs: circling, blindness, dementia, convulsion
Integument: extensive hair loss, inflammation
Application: Toxicity testing
Alternative Endpoint: Moribund or Pre-moribund State
Example: Define with specific clinical signs and euthanize when reached
Application: Many
C. IACUC Oversight of Animal Care and Use Program
C-1. Policies, Procedures and Responsibilities
C-1-1. Facility and Program Review
Introduction
Under PHS Policy and USDA Regulations, the IACUC must inspect all institutional animal facilities every six months. These inspections provide an ongoing mechanism for ensuring that the institution maintains compliance with the applicable animal care and use policies guidelines and laws. They can also benefit programs for animal care by serving an educational function for the animal care personnel, research staff and IACUC members. Also, by giving the facility personnel a prior warning, the IACUC can assist an institution to prepare for subsequent visits by outside inspectors. The interaction of an IACUC and the animal care personnel at their institution should be constructive, and not adversarial, as both ultimately share the same goals of good animal care.
Staffing and scheduling inspection
The IACUC must schedule the inspections of facilities. This may be accomplished by assigning specific facilities to subcommittees which must contain at least two members as required by the USDA Regulations. No IACUC member should be excluded should he/she wish to attend a particular inspection, and additional ad hoc consultants may be used. The inspection team must have a working knowledge of the Guide and USDA Regulations in order to fully evaluate the facilities which are being inspected. Section C-2 of this Guidebook also provides general guidance in this regard. It is helpful for the team to have a preprepared list of the categories to be inspected, such as sanitation, food and water provisions, animal identification, waste disposal, animal health records, environmental control, staff training, etc.
The IACUC may determine whether the supervisory personnel of various facilities should be notified of the date and time of an inspection. Advance notification allows individuals to be available to answer questions, but an unexpected visit shows the facility during usual operations.
Performing inspections
An updated list of all facilities to be inspected should be maintained by the IACUC. All proposals submitted to the IACUC must contain details of all locations at which animal research is to be performed. The USDA Regulations require inspection of the centrally designated or managed animal resource facilities as well as any other animal containment facilities in which animals are kept for more than twelve hours. PHS Policy requires inspection of all surgical facilities and areas in which animals are maintained longer than 24 hours. It is helpful to keep a list of all facilities by room number, use, species and deficiencies noted in the last inspection. For satellite areas a contact person is useful. For facilities with multiple rooms, a map will assist the inspectors.
Notes should be taken throughout the visit to assist in preparation of the final report. Apparent deficiencies should be discussed with the person in charge of the facility to ensure that the team's perception of the situation is correct. In some cases an apparent deviation will be due to the experimental proposal in process, for example, withholding of food prior to surgery.
Documentation
After the visit a formal report is prepared. Any deficiencies must be categorized as minor or significant. The latter is defined, by USDA Regulations and PHS Policy, as one of significant threat to animal health or safety. A plan and timetable for correction of all deficiencies must be included in the final report. All individuals to be involved in the corrections should be consulted to ensure that the plan is realistic. If the institution is unable to meet the plan, the IACUC through the Institutional Official must inform Animal and Plant Health Inspection Service (APHIS) officials within fifteen working days of the lapsed deadline. If the activity is federally funded, the relevant agency also must be informed.
The report must be reviewed and approved by a quorum of the IACUC, and in cases involving USDA Regulations, be signed by all those who accept the report. Minority views should be included in the final document. A copy is then sent to the Institutional Official and must be kept on file for a minimum of three years. It is often useful for the report to be delivered in person in order to emphasize the findings and plans for action. Annually, the institution must notify OPRR of the dates of the semiannual inspections and the dates the report was submitted to the Institutional Official.
C-1-2. Program Evaluation
Both the PHS Policy and USDA Regulations include a requirement that semiannually the IACUC conduct an evaluation of the animal care and use program. Neither of these documents includes specific guidance regarding the mechanisms or procedures to employ in conducting this evaluation. OPRR has recommended that institutions use the Table of Contents of the Guide, exclusive of the facility and physical plant chapters, as an outline for program evaluation. The USDA Regulations refer institutions to other portions of those Regulations as a basis on which to conduct this program evaluation.
Key aspects of an animal care and use program that should be emphasized in the semiannual evaluation include IACUC functions and procedures, including proposal review practices, provisions for dealing with whistle blower" or other concerns regarding animal care and use, and the procedures employed to meet reporting requirements. In addition, the institution's occupational health program, veterinary care procedures and personnel qualification review process should be evaluated. Specific procedures to accomplish program evaluation may include presentations by appropriate individuals, e.g., the institutional veterinarian, occupational health personnel, etc. Written institutional policies such as standard operating procedures may be reviewed and modified if necessary.
Program evaluation deals principally with administrative aspects of the animal care and use program. In most instances these aspects will not change nor need to be modified with the same aspects of the facility or physical plant. Thus, when large changes are made in program aspects, a comprehensive evaluation by the committee should be conducted, while the review of that aspect six months later may be merely a brief evaluation of its implementation to date. Ongoing review of established practices allows the opportunity for institutions to detect a gradual change in practices from written procedures, thereby allowing modification of one or the other as appropriate. Institutions that are AAALAC accredited will find their pre-site visit package helpful in identifying areas for inclusion in the semiannual evaluation.
C-2. Animal Health and Husbandry
C-2-1. General
The Guide states "Proper management of animal facilities is essential to the welfare of animals, validity of research data, and health and safety of the animal care staff. A good husbandry program provides a system of housing and care that permits animals to grow, mature, reproduce, and maintain good health. Good husbandry minimizes variations that can modify an animal's response to experimentation. Specific operating practices depend on many subjective and objective factors unique to individual institutions. Well-trained and motivated personnel can often ensure high quality animal care, even in institutions with less than optimal physical plants or equipment."
Adequate animal husbandry practices and health maintenance are facilitated by well-constructed and maintained caging or housing systems. Animal housing must not only confine the animal(s), but also should promote animal comfort and safety by providing sufficient space and other accommodations for normal postural and behavioral activities. The USDA regulations and the Guide provide minimum cage size requirements/recommendations for most common laboratory animal species.
Cages should allow for adequate ventilation and enable ready access to food and water receptacles. They should be constructed of materials that can be easily cleaned and sanitized, with common materials including polycarbonate plastic, stainless steel and fiberglass. Unsealed wood generally is not acceptable in animal cages or pens, as it can not be satisfactorily cleaned or sanitized.
Many animal species are social in their natural state. Some, such as the dog and cat, are readily socialized to humans. Encouragement of intra- and interspecies socialization is recommended by USDA Regulations and the Guide and is widely recognized as advantageous to animal well-being and the research endeavor. A sound husbandry program will include provision of the opportunity for animals to establish and/or reinforce social activities, including physical exercise.
Environmental factors can have a profound effect on the health and well-being of animals as well as the outcome of experimental manipulation. Temperature, humidity, air pressure and rate of turnover and noise levels all may affect animal well-being and research results. A review of an animal care and use program should include consideration of environmental standards adopted for the facilities with adequate justification available for significant deviations. While environmental control in outdoor facilities is much less stringent, acceptable ranges in temperature for several species are available in USDA Regulations. Reliable methods for monitoring environmental control systems should be in place. Redundancy in heating, ventilation, air conditioning, and lighting systems is highly desirable. Protocols for caring for animals and personnel during failures in environmental control systems should be established with personnel informed of proper procedures.
It is imperative that research animals be adequately and appropriately identified and that records pertaining to individual or groups of animals be maintained. A wide range of acceptable identification methods can be employed, ranging from cage cards to individual animal tatoos. The formerly widespread use of toeclipping to identify individual rodents is now generally discouraged, with tall tattooing or ear-tagging in more common use. Animal records may consist of only a cage card or may involve detailed individual animal information, depending principally on research requirements. Of paramount importance for research facility animal records is prominent display of the investigator's name, location and the IACUC-approved protocol number.
C-2-2. Animal Care
a. Feeding
All animals should receive food that is palatable, free from contamination and of sufficient quantity and nutritive value to maintain their good health. Specific diets should be selected based on the needs of each species, with special consideration of the requirements for vitamin C by guinea pigs and some species of non-human primates. Animals should be fed at least once a day except under conditions of hibernation, veterinary treatment, pre-procedural fasts, or other justified circumstances.
It is known that standard commercial dry bulk foods, when stored properly, retain their nutritional value for six months (three months for those containing Vitamin C). To assure that age deterioration of food does not occur, the milling date should be known (it is usually stamped on each bag), and bags should be stored so that the oldest is used first. Large amounts of food should not be stored in animal rooms. Small quantities may be kept in animal rooms if stored in tightly covered, leak- and verminproof containers; these should not be moved from room to room.
Food should be provided in receptacles that are accessible to all animals in a cage or pen and placed so as to minimize contamination. Food receptacles should be easily cleaned and sanitized, and those functions performed on a schedule that meets Guide and USDA Regulation requirements. With limited exceptions, e.g., germ-free animals in microisolator cages, food should not be placed on the bottom of the cage. Although some species may prefer this presentation, it results in waste and contamination of the food.
b. Watering
Potable drinking water should be available continuously or provided as often as necessary for the health and well-being of the animal, considering the animal's species, age, condition and any research requirements. Water may be provided in receptacles, e.g., bowls, bottles or via automatic watering systems. Whatever method is used, care should be taken to ensure that water does not become contaminated and is actually available. Sipper tubes and automatic watering devices should be checked routinely for patency. Water bottles generally should be replaced rather than refilled.
c. Bedding
Bedding may be used in the housing of a variety of commonly used laboratory animals. Bedding material should be absorbent and free of any substances that might harm the animals or alter research data. Cedar and pine products can affect liver enzymes which may in turn affect immunologic or other physiologic parameters.
Animals may be placed directly on bedding material, a common practice with many rodent species, or it may be placed under a wire or slat-bottom cage. This latter method is used occasionally for rabbits, dogs, nonhuman primates and farm animal species. Bedding should be changed as often as necessary to keep the animals clean and dry and the animal room relatively odor free.
d. Animal Activity
There is widespread disagreement among experts and the general public regarding laboratory animals' needs for supplementary physical and mental stimulation. Current USDA Regulations reflect the will of Congress in the 1985 Amendments to the Animal Welfare Act in that they require institutions to develop plans for: (1) providing dogs the opportunity to exercise and (2) enhancing the environment of nonhuman primates so as to promote their psychological well-being. Conditions and requirements for these plans are included in the Regulations. Where no regulatory requirements exist, the decision to supplement activity in animals should be made by the institution, in most cases through the IACUC and based on recommendations of the veterinarian in consultation with the investigator. Factors to be considered in this decision include the animal's species-specific characteristics, temperament, physical condition, previous housing conditions, nature of the research and anticipated duration of housing. There is widespread agreement that the best enhancement of environment for an animal that is social in nature is another animal. Group housing of most species generally is encouraged, recognizing that research requirements may preclude it and some negative effects (e.g., fighting, potential for disease transmission) may be seen.
e. Emergency, Week-end and Holiday Care
Laboratory animals must be observed by qualified personnel every day in order to ensure their health and well-being, as well as to promote sound research practices. Skilled assistance, including veterinary care, must be readily available at all times. Names and telephone or pager numbers of those assigned these responsibilities should be prominently displayed in the facility.
C-2-3. Facility Maintenance
a. Cleaning and Sanitation
Cleanliness and sanitation are essential to the operation of an animal facility. The Guide and USDA Regulation set forth recommended frequencies and methods for cleaning and sanitation of facilities, equipment and accessories. In general, the frequency and methods should ensure that animals are maintained in a clean, dry environment, free from exposure to harmful contamination and excessive animal odors. Cleaning equipment such as mops and pails should not be moved from room to room.
The most efficient and effective method of cleaning and sanitizing cages and accessories (e.g., feeders, water bottles, sipper tubes) is the use of a mechanical washing machine that provides rinse water temperature of at least 83 C (180 F). Alternatively, portable high pressure spray washing and disinfection may be used. Least efficient and effective is hand washing and disinfection of such equipment. The supply lines of automatic watering systems should be flushed and disinfected on a regular basis.
b. Waste Disposal
A research animal facility generates a significant amount of waste that must be removed and disposed of on a regular, frequent basis. Disposal methods, including incineration and removal to land-fill must conform to federal, state and local requirements. Some jurisdictions consider all soiled animal bedding from a research facility to be "medical waste," with consequently more stringent disposal requirements.
If waste must be stored while awaiting disposal, the storage area should be outside the animal holding and clean equipment areas. Animal carcasses and tissues require a separate cold storage area and regularly scheduled removal. Hazardous waste, including carcasses of animals exposed to radioactive or biohazardous agents, must be adequately stealized and/or contained prior to removal and disposal.
c. Vermin Control
The research animal facility is an active place, with frequent movement of personnel, animals, equipment, containers, food and bedding. This creates ideal conditions for the introduction of vermin from arthropods to wild rodents. Vermin control programs are complicated by the potential for harm to animals and personnel, as well as interference with research data by many commonly used pesticides. The most effective program combines elimination of all entry and harborage sites, with good waste disposal and personnel training.
C-3. Occupational Health (See Table 5)
Purpose of occupational health programs
The health of individuals working in animal care Programs is an area of institutional concern. PHS Policy and the Guide identify the need for an occupational health program for all personnel who work in laboratory animal facilities or who have substantial animal contact. The emphasis of such a program is the prevention of illness, but it also includes provisions for early diagnosis and treatment when such illnesses occur.
Elements of an occupational health program
An effective program will have the following components: 1) replacement medical evaluation; 2) periodic medical surveillance; 3) educational component; 4) provisions for treating illness or injury; and 5) provisions for consultation with other professional staff. The specific elements will be dictated by the extent and nature of the employee's exposure [see table].
Replacement and periodic medical evaluations: Replacement evaluations are conducted to ensure that the individual is capable of the demands and exposure of the job, and also to provide a medical reference baseline. The evaluation may include: clinical history, physical examination, spirometry, baseline tests such as TB test and serum sample collection, appropriate immunizations, educational/instructional component and appropriate feedback to the employee on all test results. Specific tests will depend on the species of animals and the nature of the procedures employed.
Periodic evaluations allow detection of early stages of disease, updating of immunizations and a re-evaluation of medical restrictions.
A uniformity in the evaluation of different individuals and the same person at different times is important to enable accurate comparisons to be made. These comparisons may allow a possible problem to be identified and corrected before it becomes a major health hazard.
Education: There are ethical and legal requirements to inform individuals of health risks and precautions which affect them. This must be part of an employee's overall orientation and job training. Some institutions rely on formal courses.
Medical care and treatment: In addition to established mechanisms for reporting and treating accidents and injuries, the institution should have access to medical expertise in zoonotic diseases and other health risks associated with laboratory animal care. Such expertise will greatly assist in the recognition of diseases, allergies or musculoskeletal overuse syndromes associated with animal care. Good communication with medical staff will also facilitate better management of the health of animal care personnel and minimize repeat injuries and infections.
Specific medical concerns for individuals working with laboratory animals
Allergy and musculoskeletal injury constitute the primary health risks to individuals using and caring for laboratory animals. Allergies are a significant problem, and can be reduced by the provision of protective equipment to affected personnel. Musculoskeletal injuries can be minimized by good laboratory planning, use of transport equipment such as carts, and training in lifting and equipment use.
Infectious diseases also form a significant risk depending on the species and health status of animals involved and the level of exposure to them by animal care personnel. Infectious diseases to which animal care personnel may be exposed include a number of viral infections, some of which can be extremely serious, for example rabies. In addition, infections acquired from live animals, animal tissues and excreta can serve as a source of zoonoses. Careful monitoring and quarantine of any animals with potential viral or bacterial infections is a crucial part of any animal care program. Particular care must be taken in all facilities handling primates as they are likely to carry infections which can be transferred to humans, for example, Herpes Virus Simiae (Herpes B) and tuberculosis. Routine TB testing is essential.
Animal bites and scratches are hazards common to animal facility personnel. All cases should be documented. Tetanus prophylaxis should always be considered and, depending on the species, rabies prophylaxis and antibiotics arranged.
References
Bowman, P. N. 1991. A flexible occupational health and safety program for laboratory animal care and use programs. AALAS Bulletin 306(6).
Centers for Disease Control, Update: Ebola-related filovirus infection in nonhuman primates and interim guidelines for handling nonhuman primates during transit and quarantine. 1990. MMWR 39(2), 22-23.
Montrey, R. D., et al. September 1989. An automated occupational health program for laboratory animal handlers. Lab Animal, 27-30.
Richmond, J. Y. February 1991. Hazard reduction in animal research facilities. Lab Animal 20(2), 23-29.
Soave, 0. and C. D. Brand. February 1991. Employer responsibility for employee health in the animal environment. Lab Animal 20(2), 41-44.
Table 5. Occupational Health Program for Animal Handlers
| Species Used/ Other Factors | Direct Regular Contact (8hrs/week or more) | Direct Contact Limited Exposure (less than 8hrs/week or more) | No Direct Contact, Occassional Exposure |
| Small Animals (Rabbits, Rodents, Birds) | 1 2 5 | 1 2 5 | |
| Dogs, Cats and Feral Animals | 1 2 4 | 1 2 4 | 3 |
| Primates | 1 2 3 8 | 1 2 3 8 | |
| Farm Animals | 1 2 6 | 1 2 6 | |
| Amphibians, Reptiles, Fish and Other Cold-Blooded Animals | 2 | 2 | |
| Infectious Disease Studies (Class III or Higher) | 1 2 7 8 9 | 1 2 7 8 9 | 1 2 7 8 9 |
| Work with Animal Tissues | 7 | 7 | 7 |
| Code Number | Procedure |
| 1 | Pre-Employment Physical Exam (including serum for banking) |
| 2 | Tetanus Immunization (every ten years or following known injury or advice of physician) |
| 3 | TB test (every six months) |
| 4 | Rabies Immunization |
| 5 | Pre-Employment Allergy Evaluation and Education |
| 6 | Special Education on Large Animal Diseases |
| 7 | Special Consideration by Infectious Disease Committee |
| 8 | Post-Employment Physical (including serum for banking) |
| 9 | Annual Physical Exam |
D. IACUC Evaluation of Animal Welfare Concerns
D-1. Policies, Procedures and Reporting (See Table 6)
Introduction
One of the basic functions of the IACUC, as specified in the USDA Regulations is to "review and, if warranted, investigate concerns involving the care and use of animals at the facility resulting from public complaints received and from reports of noncompliance received from laboratory or research facility personnel or employees" 9 CFR Part 2, Subpart C, Section 2.31 (c)(4). Also required under 9 CFR Part 2, Subpart C, is training of personnel in the "methods whereby deficiencies in animal care and treatment are reported including deficiencies in animal care and treatment reported by any employee of the facility." The institution should have established procedures for reporting, receiving and handling allegations of animal mistreatment or other noncompliance.
Definition of mistreatment and non-compliance
Mistreatment is physical or psychological, wrongful or abusive treatment of an animal. This is a broad definition and clear gray areas exist which the IACUC must interpret carefully. Fortunately, it is rare that mistreatment is intentional. Noncompliance means that procedures or policies are not being followed, and this may stem from confusion or misunderstanding. There are areas of overlap between mistreatment and noncompliance, and the procedures for handling both are very similar.
Institutional policies
The highest administrative levels must advocate the finest animal care and by so doing assure the public, researchers, employees or students that there is a true desire to investigate allegations of mistreatment or noncompliance. The IACUC and veterinarians must perpetuate the same attitude. There must be no implication that reporting such instances could be detrimental to an individual's standing within the organization. Indeed, the USDA Regulations provide specific protection under the law. [9 CFR, Part 2, Subpart C 2.32 (c)(4)]
It is prudent to have established procedures for handling allegations of mistreatment or noncompliance before any such allegations are raised. Addressing complaints against the appropriate use and care of animals is very difficult and only with a clear set of guidelines can this be performed in a thoughtful and systematic manner. A direct statement to the IACUC, or its designated subcommittee, helps empower the IACUC to deal with such allegations.
Reporting allegations: It is not always obvious at what level of alleged mistreatment or noncompliance the IACUC should become involved. Frequently the attending veterinarian animal care personnel and investigators can work together to prevent or correct problems. However, serious or repeated problems always demand the involvement of the IACUC. If in doubt, it is better to report, as this may well protect the institution, the complainant, the alleged violator(s) and, of course, the animals.
A variety of ways can be provided in which to report allegations. These may include conversations with, or letters to, members of the IACUC, the veterinary staff or the Institutional Official. As dictated by the USDA Regulations, there must be no restrictions on who can report an alleged incident and no threat of reprisals against those reporting perceived mistreatment or noncompliance. Whatever the route, the information should be quickly relayed to the chairman of th