of Ethics Online Collection:None
A.The Institutional Animal Care and Use Committee (IACUC)
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:
- 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;
- Establishment, according to specified criteria of an Institutional Animal Care and Use Committee with defined responsibilities;
- Detailed requirements for the submission of applications for awards;
- Recordkeeping requirements to ensure clear accountability for the quality of the program; and
- 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
- Review, at least once every 6 months, the research facility's program, using USDA Regulation/Guide as basis.
- Inspect, at least once every 6 months, all of the animal facilities, including animal study areas/satellite facili- ties, using USDA Regulations/Guide, as basis.
- Prepare reports of IACUC evaluations and submit the reports to the Institutional Official.
- 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.
- Make recommendations to the Institutional Official regarding any aspect of the research facility's animal program, facilities or personnel training.
- 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.
- 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.
- 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 IACUCapproved 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 oxy- gen, 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-2-2.
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 discouraged
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 qualified 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 repor