Welcome to the course Writing an Animal Protocol for Research on Rats.

This is the rat module in a course series on the preparation of an animal use protocol. Each course in this series refers to a different animal species. Every course offers information that is both pertinent to all research animals and specific to the one animal species presented.

• The first 7 lessons are similar in all courses of this series. Differences in these lessons relate mainly to the regulatory coverage, housing requirements, and the zoonotic hazards of each animal species.

• The remaining lessons present information that is more specific to each animal species, such as biological features, anesthetic doses, and biomethodologies.

The goal of this course is to cover important information about using rats in biomedical research settings. If you are responsible for handling rats or if you must write an animal use protocol, this course will be useful by providing you with:

• Summaries of key regulatory issues.
• Guidance on searches for alternatives in the care and use of animals.
• Highlights of unique biological features of these animals.
• Overviews of acceptable basic methodologies.
• Requirements for supportive care procedures.

Hypertext links in this course provide you with supporting information, such as regulatory sources, drug doses, practical tips, etc.

This course will not provide you with detailed information on how to conduct the methods and procedures described. For this, you should use other courses offering in-depth information and hands-on instruction from your institution's animal facility staff.

Click to view the credits for this course.

To ensure the humane treatment of laboratory animals, animal research is regulated by two federal agencies:

• The United States Department of Agriculture (USDA) / Animal Care; and
• The Public Health Service / Office of Laboratory Animal Welfare.

The USDA and PHS mandates on animal welfare differ greatly with respect to the laboratory strains of mice and rats. These species are not covered by the USDA but are included in PHS regulations and policy. However, the USDA may eventually regulate these species as well.

If your institution receives any funding from the PHS or is accreditated by the Association for Assessment and Accreditation of Laboratory Animal Care, International (AAALAC), then your research must also comply with the National Research Council publication, the Guide for the Care and Use of Laboratory Animals. This document will simply be referred to as the Guide in this course.

The Public Health Service Policy requires institutions to have an occupational health and safety program for individuals working with laboratory animals. This requirement is also reiterated in the Guide.

It is the responsibility of principal investigators to assure that their laboratory staff are informed of and participate in their institution's occupational health and safety program.

Elements of an occupational health and safety program, including institutional responsibilities, are described in the guideline, Occupational Health and Safety in the Care and Use of Research Animals (shown at right), published by the National Research Council.

Working with rats is associated with the following hazards:

Injuries
Personnel handling rats can be injured by bites from the incisor teeth. Generally, these are caused by a lack of knowledge in how to handle, transport and restrain a rat. Generally, a rat will bite only when frightened or in pain. Likewise, poor technique in handling, etc., can cause injury to the rat. Training staff to work effectively and humanely with rats is essential to prevent injuries to people and rats.

Allergies
People can develop an allergy to rats after having contact with them for some time. Persons who develop allergy symptoms should seek medical counseling and may have to discontinue working with this species.

Zoonoses

In general, transmission of zoonotic disease from naturally infected laboratory animals is uncommon because of the ongoing efforts to improve the health status of animals by vendors of specific pathogen free animals and by facility staff. Experimentally infected animals are a source of zoonotic transmission to humans. Also, contact with wild rats in field research may expose humans to zoonotic agents carried by this species. Health surveillance programs, routine sanitation, and personal protective equipment have important roles in preventing zoonoses.

Rats can be a reservoir of the following infectious agents that are transmissable to people:

Viruses

• Hantavirus




Hantavirus is a bunyavirus carried by wild mice and rats. The virus is transmitted to man by excretions and aerosols from the lungs, saliva, and urine of infected animals.

Humans are at risk for Hantavirus infection (Korean Hemorrhagic Fever) primarily from wild rodents (e.g., the wharf rat or cotton rat). Strains vary in symptoms based on geographical origin (US, Asia, Scandinavian, and Europe). Hantavirus occurring in the southwestern U.S. causes a severe pulmonary syndrome. Viral strains orginating in Asia produce a hemorrhagic fever and nephropathy. Strains originating in northern Europe generally produce renal symptoms of less severity. Researchers performing field studies on wild-caught rodents are at particular risk. To see recommendations for preventing exposure to hantavirus infection in field study environments, click here.

Bacteria

• Leptospira spp.




Rats may be a reservoir for Leptospira spp. bacteria, which are shed in the urine. Transmission occurs by contact with urine and tissues, or inhalation or ingestion of aerosol droplets.

Humans with leptospirosis may have influenza-like symptoms, orchitis, rash, skin and mucosal hemorrhage, hemolytic anemia, hepatorenal failure, jaundice, encephalitis, and pneumonia. Click here for information on human leptospirosis from the CDC website.

• Salmonella spp.
  
  

  Rats may carry Salmonella spp., which are ubiquitous in nature. These bacteria are transmitted via the fecal-oral route.

  Humans infected with Salmonella may have inapparent clinical signs (and be carriers) or may have a febrile enterocolitis, septicemia and focal infections in diverse tissues. Increased severity of the disease occurs due to reduced immunocompetence, e.g., in persons with AIDS, neoplasia, immunosupression therapy, etc., and due to treatment of antibiotics.

• Streptobacillus moniliformis and Spirillum minus
  
  

  These organisms cause rat bite fever. Rats may carry Streptobacillus moniliformis and Spirillum minus as commensal organisms living in the nasopharynx. Transmission to humans occurs by rat bite. Symptoms may include wound inflammation, lymphadenopathy, fever, headache, malaise, myalgia, joint pain, and arthritis. Click here for a recent case study of rat bite fever in a boy.

Fungi

• Sporothrix schenckii
  
  

  Sporothrix schenckii is a fungus associated with multiple species, including rats.

  Humans acquire Sporothrix schenckii via bites or direct contact. Disease manifests as a nodule on the hand or arm, which may progress with additional nodules forming along the path of lymphatic vessels. Deep visceral infections may also occur.

• Trichophyton spp.
  
  

  Dermatophytic fungi grow in the skin and hair follicles and cause a condition of reddened skin and patchy hair loss known as ringworm. The symptoms are the same in animals and humans. Infection may be inapparent in individual animals. Dermatophytes are spread by direct contact. Fungal spores are long-lived and may become widely dispersed in the environment. Extended therapy with antifungal medication may be needed to eliminate infection in animals and humans.

For more information, refer to Occupational Health and Safety in the Care and Use of Research Animals, published by the National Research Council.

Your protocol form should ask you for an assurance that you have considered alternatives to the use of animals if painful or distressing procedures are proposed. This is to satisfy a mandate by the Animal Welfare Act and PHS Policy to avoid or minimize discomfort, pain, and distress consistent with sound scientific practices. Alternative procedures are those which may replace animals with nonanimal methods, reduce the number of animals used, or refine the methodology to minimize animal pain or distress. For more information on what is meant by alternatives to the use of animals, please refer to the course Working with the IACUC, which is part of this series.

The assurance often takes the form of a written narrative that describes which sources were used to determine that alternatives were not available. Typically, you may be asked to provide the results of a database search including information on:

1. The databases searched.
2. The date the search was performed.
3. The years of citations covered by database searches.
4. The key words and/or search strategy used when searching a database.

It is strongly recommended that this information be sought during development of a protocol.

Organizations that can assist you in performing an alternatives search are:

• ALTWEB, Center for Alternatives to Animal Testing, Johns Hopkins University
• Animal Welfare Information Center, National Agricultural Library
• Center for Animal Alternatives, University of California - Davis

The following is a case study of alternatives searches that may guide you in the development of a search strategy that is pertinent to your own research.

Click on each database below for a sample search on key terms for this example.
You can follow the links to the database for a search in real time.

Example Search:
Use of gas anesthesia instead of injectable anesthetics for survival surgeries.

• PubMed
• Agricola

All of the federal laws, regulations, policies and guidelines applicable to animal research have a core intent to ensure the humane treatment of the animals involved in a study. Accordingly, your IACUC will have requirements for the proper care of your animals prior to, during, and after a research procedure.

What is a procedure? A procedure is any activity carried out on the animal, such as controlled behavioral observation (e.g., use of a maze), venipuncture, or surgery. Peri-procedural care requirements include:

• Properly preparing the animal to humanely undergo the procedure;
• Supporting the animal's physiological function during the procedure; and
• Providing additional supportive care to aid the animal in recovering from the procedure.

The investigator has the responsibility to see that staff working with the animals are properly trained not only to perform the procedure humanely but also to provide the necessary supportive care to the animals.

When performing any procedure, such as a blood collection, you should think through the steps that are necessary to protect the animal's welfare. For example, for blood collection, you should limit the volume to the minimum that may be taken safely. That amount may vary depending upon whether a single sample is taken or multiple, chronic collections are done And, with any venipuncture, you should be prepared to care for the animal in the event of trauma to the vein or excess hemorrhage.

Refer to your institution's IACUC and attending veterinarian for specific guidelines.

Your protocol form may ask you which type of housing you may need for your rats. There are important considerations in the selection of animal housing that affect the welfare of your animals. Rodent caging has two types of flooring: solid and wire mesh.

• The solid flooring of shoebox cages are covered with a bedding material that absorbs liquid wastes. Bedding has been shown to be preferred by rodents for resting, and it is considered to provide them with comfort, warmth, and the opportunity to burrow. This type of flooring is well suited to breeding because pups are better protected from chilling.
• Wire mesh flooring has long been used for rodent caging because of advantages in sanitation. Typical rodent cage mesh has two to four wires per inch (2.5 cm). This type of flooring has been associated with foot injury in rats. Inserting a solid resting board into the cage provides the rat with a comfortable resting area and may reduce foot injuries. The use of wire bottom cages is discouraged for rodents, especially on long-term studies. Use of wire bottom cages should be scientifically justified and approved by the IACUC

Because of data on rodent preferences for solid flooring and the risks for animal injury on wire mesh flooring, the use of wire bottom cages should be scientifically justified and approved by your institution's IACUC. Related guidelines are:

Guide for the Care and Use of Laboratory Animals

 

ARENA/OLAW IACUC Guidebook

Example Alternatives Search:
For additional information on types of rat caging and the impact of caging type on rats, please refer to the examples of "alternatives" searches on rat housing.

Upon arrival at your facility, your rats should have an acclimation period before they are used in research studies. This period of time allows animals to adapt to a new environment. Effects of transportation stress include alterations in various blood parameters, immune cell function and animal behavior. The period of time necessary for biological stabilization will depend on the parameters to be studied. Refer to your institution's attending veterinarian for recommendations that are appropriate for your project. Typically, acclimation periods range from 4 days to 1 week.

Example Alternatives Search:
For citations on the impact of transportation stress and acclimation on rats, please refer to the example of "alternatives" searches provided.

Routine quarantine procedures may prolong the holding of your animals in special facilities. Quarantine aims to prevent transmission of diseases between new animals and established colonies.

Acclimation and quarantine periods run concurrently, although they serve different purposes. Most institutions do not allow experiments on animals while quarantined.

If your proposed study involves a painful procedure, the protocol form may ask for a method of assessing if the rats are experiencing pain or distress.

Assessing pain and distress in rats is difficult at times because rats, like many other species, commonly conceal outward signs of moderate pain and distress. In this case, the behavioral changes that reveal a rat's pain and distress may be subtle and detectable only with specialized behavioral methods.

Severe pain and distress causes overt clinical signs in rats. Laboratory staff working with rats should be trained to recognize these abnormalities in:

• Activity level – e.g., hypoactivity, hyperactivity, restlessness.
• Behavior – e.g., vocalization, self-trauma, isolation from cage mates, aggressiveness, ataxia.
• Appearance – e.g., unkempt greasy fur, porphyrin staining around eyes and nostrils, hunched posture, cyanosis, pale mucous membranes, soiled anogenital area.
• Vital Signs – e.g., respiratory distress.
• Body Condition – e.g., weight loss, emaciation, dehydration.
• Intake – e.g., reduced intake of food and water.

The image below shows porphyrin staining and encrustation on a rat's nose. This is a nonspecific sign of pain or distress.

A chronic state of pain or distress may be more subtle and difficult to detect. A good knowledge of the animal’s normal appearance and behavior is especially important to recognize chronic pain or distress.

For methods on assessing and alleviating pain and distress in rodents, refer to another course in this series, Post Procedure Care of Mice and Rats in Research: Minimizing Pain and Distress.

Example Alternatives Search:
For citations on detecting pain and distress, please refer to the example of "alternatives" searches provided.

Inbred strains and outbred stocks of rats produce animals that are used for different purposes. The decision to use isogenic inbred strains or non-isogenic outbred stocks is determined by the experimental strategy.

• Inbred strains are used for genetic engineering and finely controlled studies that capitalize on genetic isogenicity. Inbred strains with characteristics of human diseases or physiological conditions are generally preferred models for biomedical research.
  Example: Lewis rats, LEW/CrlBR
• Outbred rats are used when outbred vigor is desirable, e.g.,when genetic heterogeneity and phenotypic variability are not a concern.
  Example: Wistar rats, Hsd:WI

Please check with your animal resource department for information on vendor choices as animal source affects animal health status.

Though rats share many anatomical and physiological features with humans, rats have many unique biological characteristics. A knowledge of species-specific characteristics is helpful to effectively manage these animals and to plan experimental procedures for their use. Researchers should be aware of the following practical features of rat anatomy and biology.

Click on the following items for a brief description and some practical tips.
The photo shows a rat with overgrowth of the incisors (upper and lower) and jaw malalignment.

Anatomy

• Ocular
• Teeth

Gastrointestinal

• Inability to vomit. Note: click this ink to find find out why there is no justification for routinely fasting rats prior to surgery!
• Gall Bladder
• Coprophagy

Metabolism

• Albinism
• High rate of metabolism – impact on drug clearance
• High surface area – impact on hypothermia

The following are volume recommendations for intravenous fluid administration and blood collection in adult rats:

IV Fluid Volume (ml) max. acute admin.	Total Blood Volume (ml)	Safe Bleed Volume (ml)a	Bleed-out Volume (ml)b
0.5 - 1.0 ml	64 ml/kg	5.5 ml/kg	6-14 ml

^aRemoving greater quantities of blood (exceeding 10% of total blood volume) can produce hypovolemic shock. Repeated collections of smaller amounts of blood will have the same effect. In such procedures, animals should receive warmed, physiological fluids to replace the volume of blood collected. In addition, monitor the animal’s hematocrit for anemia.

^bAnimals should be exsanguinated only under anesthesia; volumes shown refer to adult rats.

From:

1. Hawk and Leary, Formulary for Laboratory Animals, 2nd Edn., Iowa State University Press, 1999.
2. Wolfensohn and Lloyd, Handbook of Laboratory Animal Management and Welfare, 2nd Edn., Blackwell Science. 1998.

Below are peripheral vessels that are commonly accessed for blood collection or fluid administration. Recommended needle sizes are 23 to 27 gauge. Larger needles may be necessary for injecting large volumes or viscous materials.

Below are the nonvascular routes of injection that are commonly used in rats. Included are volume recommendations for the safe administration of fluids acutely in adults (average 200 g). Recommended needle sizes are 23 to 27 gauge; larger needles may be necessary for injecting viscous materials.

Subcutaneous (SQ or SC) - 25 ml/kg

Intraperitoneal (IP) - 25 ml/kg

Oral (PO) - 10 ml

Intradermal (ID) - 0.05 ml/site

Intramuscular (IM) - 0.1 ml per site

From:

1. Hawk and Leary, Formulary for Laboratory Animals, 2nd Edn., Iowa State University Press, 1999.
2. Wolfensohn and Lloyd, Handbook of Laboratory Animal Management and Welfare, 2nd Edn., Blackwell Science. 1998.

Example Alternatives Search:
For citations of blood collection procedures in rats, please refer to the examples of "alternatives" searches provided.

When using any animal species for polyclonal antibody production, the issues below and on the following screen should be addressed within the animal protocol. For more detail, refer to the Institute for Laboratory Animal Research publication, ILAR Journal Volume 37(3) Adjuvants and Antibody Production, 1995.

Antigen Preparation
The antigen preparation should be free of extraneous microbial contamination and byproducts such as polyacrylamide gel. The protocol should describe how the antigen-adjuvant emulsion will be prepared.

Adjuvant Used
If the use of Freund's complete adjuvant (FCA) is proposed, your IACUC may require justification of this choice of adjuvant. FCA has been associated with granulomatous inflammation, focal necrosis, ulceration of skin, fistulous tracts, muscle atrophy, self-induced trauma, hypersensitivity reactions, and weight loss. The USDA states that the injection of FCA may cause more than momentary or slight pain. This means that FCA injections might put an animal into USDA pain category D (painful/stressful but relieved), requiring the use of post-injection analgesics. Check with your IACUC to determine your institution's policy.

Recommendations for FCA from the Institute of Laboratory Animal Research are as follows:

1. FCA should be used only once, usually for the initial immunization.
2. Formulations of FCA should not exceed 0.1 mg dry mycobacterial cell mass/ml.
3. Less inflammatory alternatives to Freund's adjuvant are available and should be considered.

Booster Frequency
In common booster schedules, the initial and subsequent immunizations are spaced at intervals of two to three weeks (minimum). Booster immunizations may be delayed if significant inflammatory reactions are still present from the initial immunization.

Injection Site Selection and Preparation
Excerpted from the ILAR Journal 37(3) (Institutional Policies and Guidelines on Adjuvants and Antibody Production): "Anatomic sites used for grasping, handling, or restraint….should be avoided when possible. Extension of granulomatous inflammation into the spinal cord following inadvertent injection of a FCA-antigen mixture into the paraspinal musculature has been associated with posterior paresis in guinea pigs (Kleinman et al., 1993). Care therefore should be taken when making injections near the dorsal spinal column. Granulomas can also be noted in other organs after injections with FCA (Schiefer and Stunzi, 1979)."

Post-injection Observations
Your animal protocol should describe how animals will be monitored for post-injection lesions and how affected animals will be treated. Refer to the ILAR Journal for recommendations on post-injection observations.

Because rats have a high rate of metabolism, drugs are rapidly eliminated from their bodies. Dose rates appropriate for larger species produce ineffective drug concentrations when used in rats.

This section includes dose rates for the common drugs and drug regimens used in rats. If you need to use other drug agents, check with your institution's veterinary staff for assistance in determining a dose rate appropriate for use in rats.

Click on the drug types for doses of common agents and drug regimens that may be used in rats:

Analgesics:
Available in two drug types – the opioids and the nonsteroidal anti-inflammatory drugs (NSAIDs). The rapid clearance of many of these drugs in rats results in the need for an increased frequency of administration.

Sedatives:
Sedatives may obtund consciousness but in normal doses do not do so sufficiently to ablate the perception of pain or other sensations. When combined with general anesthetics, they may be used to induce a "balanced" anesthesia where muscle relaxation, unconsciousness, and analgesia are enhanced.

Sedatives + Analgesia:
Some sedatives also have analgesic effects. When combined with general anesthetics, a balanced anesthesia is attained, and these sedatives enhance analgesia through specific effects.

Anesthetics:
Because rats metabolize drugs so rapidly, many anesthetic agents have brief durations of effect. An anesthetic regimen should be chosen to match the duration of drug effects with the length of the procedure. In particular, short acting agents (and regimens) should be not be used for long procedures because repeat drug administrations, necessary to prolong anesthesia, will produce uneven blood concentrations and therefore periodically inadequate anesthesia. For long procedures, gaseous anesthesia is often the most practical method to sustain uniformly adequate levels of anesthesia. Explosive agents such as ether should not be used.

The practice of using hypothermia as an anesthetic for neonates is generally discouraged.

It is not clear whether the depression of neural function by hypothermia is sufficient to prevent the sensation of pain related to a surgical procedure. Also, the recovery from hypothermia may be a painful experience in animals, as it is known to be in humans.

An acceptable alternative to hypothermia for neonates may be a non-explosive inhalational agent such as halothan delivered using a non- rebreathing system.

Example Alternatives Search:

For additional information on anesthesia in rats, including the physiological impact of anesthetic agents on rats, please refer to the examples of "alternatives" searches provided.

Aseptic technique should be used when performing surgery on rats. The standards described here are consistent with the Guide for the Care and Use of Laboratory Animals.

Surgery on rats should be performed in a location that allows for a physical separation of the operative field from other functions of the procedure (such as animal preparation and anesthetic recovery) and other laboratory activities. The isolation of the operative field avoids contaminating sterile areas with animal fur, bedding, nonsterile supplies, etc. The location used for the operative field should be cleaned and sanitized before use. Materials and supplies used in support of the procedure should be positioned and managed to avoid contaminating sterile areas.

Surgical procedures in rats should be conducted using aseptic technique.

Nonaseptic methods are not acceptable. Rodents have been shown to develop subclinical infections, a consequence which has led to an outdated belief that rodents tolerate nonaseptic technique without developing postoperative infections. The Guide recommends methods for adapting aseptic technique to the scale of rodent surgery. In this way, efficiencies and economies can be realized without sacrificing asepsis.

Supportive care aims to:

• Maintain the animal's physiological status as nearly normal as possible.
• Minimize animal pain and distress.

Supportive care includes monitoring both physiological parameters and analgesia during anesthetic and surgical procedures. Monitoring of vital signs and pain should be conducted throughout the procedure and the recovery period.

The photo shows the monitoring of analgesia in a rat (assessing the toe pinch reflex).

Keep in mind that:

• General anesthesia causes disturbances of thermoregulation and other physiological functions. Maintaining body temperature, e.g., via insulating materials and heating sources that do not pose a risk of thermal injury, is an important objective of supportive care.
• During surgery, the animal may experience pain if anesthesia is inadequate at any time during the procedure.
• Postoperatively, the animal may experience pain, discomfort, and distress unless treated with analgesics and appropriate supportive care measures.

Due to the interaction of metabolic factors and drug effects that can cause animal mortality, rats should receive good supportive care and monitoring during anesthesia, whether or not the procedure involves surgery.

During anesthesia and surgery, the following procedures are recommended.

Supportive Care:

• Provide a source of warmth to rats from the onset of anesthesia to the end of anesthetic recovery.
• Inject sterile physiological fluid (warmed to body temperature) to compensate for blood loss during a procedure and depressed fluid intake post-procedure.

Monitoring during Anesthesia:

• Analgesia - toe pinch.
• Respiration -gross changes in rate, character of breathing.
• Color of mucous membrane and skin – blue (poor oxygenation), pale (poor blood perfusion).

The photo shows a rat with the trachea intubated, ready to be connected to a ventilator.

After anesthesia and surgery, the following procedures are recommended.

Monitoring post Anesthesia:

• Rats must be monitored until fully recovered from anesthesia as indicated by the ability to ambulate and maintain core body temperature.

Monitoring Post Procedure:

• Assess appearance, activity, and behavior as indications of pain and discomfort (see screen Detecting Pain and Distress).
• Assess food and water intake.
• Provide floor-level access of food and water post procedure if stretching overhead for these items (in the cage wirelid) may be painful.
• Assess wound repair. Routine use of antibiotics is not indicated after uncomplicated, aseptic surgery.

The term euthanasia is derived from Greek and means "good death." Animals should be euthanatized when killed for any purpose, including research. To euthanatize a rat, you must be trained in the concepts of euthanasia, the method to be used, and the proper handling of rats.

Methods are classified as acceptable or conditionally acceptable, as set by the American Veterinary Medical Association. The inclusion of conditionally acceptable methods in your protocol may require scientific justification and IACUC approval.

The photo shows a CO₂ chamber for euthanatizing rodents.

Click on each method below for recommendations on its use.

Acceptable Methods:

• Barbiturates
• Inhalant anesthetics
• Carbon dioxide (compressed tanks only)
• Carbon monoxide
• Potassium chloride in conjunction with general anesthesia
• Microwave irradiation

Conditionally Acceptable Methods*:

• Ether
• Nitrogen
• Argon
• Cervical dislocation (rats < 200 g)
• Decapitation

*The inclusion of conditionally acceptable methods in your protocol may require scientific justification and IACUC approval.

Very Important!

Before placing euthantized rodents in a bag and placing the bag in a necropsy refrigerator or freezer, you must make sure the rodents are really dead! Rodents can stop breathing for a minute or more then regain respiratory function and survive. This is particularly true of younger rodents, which are somewhat resistant to carbon dioxide asphyxiation and take longer to succumb than adult rodents.

To ensure death in rodents euthanatized with carbon dioxide, the chest cavity may be opened with scissors, or the rodents may be observed for an extended period of time to make sure they are dead. Your institution may have specific requirements.

The Office of Laboratory Animal Welfare (responsible for enforcing PHS policy) has made it clear that rodents remaining alive in bags after ineffective euthanasia is a serious breach of PHS policy, and must be reported to regulatory officials.

Example Alternatives Search:
For additional information on euthanasia of rodents, including the impact of euthanasia agents on tissues, please refer to the From:PHS POLICY ON HUMANE CARE AND USE OF LABORATORY ANIMALS CLARIFICATION REGARDING USE OF CARBON DIOXIDE FOR EUTHANASIA OF SMALL LABORATORY ANIMALS; Release Date: July 17, 2002; NOTICE: NOT-OD-02-062

Unintended recovery of animals after apparent death from CO2 (e.g., in necropsy coolers) is a documented occurrence. Institutions are reminded that such incidents constitute serious noncompliance with the PHS Policy and serious deviation from the provisions of the Guide for the Care and Use of Laboratory Animals (http://www.nap.edu/readingroom/books/labrats/). As such, the IACUC, through the Institutional Official, must promptly provide OLAW with a full explanation of the circumstances and actions taken. Prompt reporting in accordance with PHS Policy requirements (PHS Policy IV.F.3) is an essential component of the formal relationship between OLAW and PHS-Assured institutions."

Click here to see the entire source document on the OLAW website.

Federal Laws, Regulations, Policies:

1. Animal Welfare Act, as Amended (7 USC, 2131-2156).
2. Animal Welfare Act Regulations and Standards, Code of Federal Regulations, Title 9, Part 2-Regulations, Sections 2.31-2.33, 1998.
3. Health Research Extension Act of 1985, Public Law 99-158, November 20, 1985, "Animals in Research".
4. Public Health Service Policy on Humane Care and Use of Laboratory Animals, Revised September, 1986, Reprinted March, 1996.
5. USDA Animal and Plant Health Inspection Animal Care Policy Manual.
   Policy #11 – Painful/Distressful Procedures.
6. U.S. Government Principles For The Utilization And Care Of Vertebrate Animals Used In Testing, Research, And Training, Interagency Research Animal Committee.

Guidelines:

1. Guide for the Care and Use of Laboratory Animals, National Research Council, 1998.

Texts:

1. Hawk and Leary, Formulary for Laboratory Animals. 2nd Edn., Iowa State University Press, 1999.
2. Hrapkiewicz, Medina, and Holmes, Clinical Laboratory Animal Medicine: An Introduction, 2nd Edn., Iowa State University Press, 1998.
3. Sharp and LaRegina. The Laboratory Rat. CRC Press, 1998.
4. Wolfensohn and Lloyd, Handbook of Laboratory Animal Management and Welfare. 2nd Edn., Blackwell Science. 1998.