The euthanasia method chosen must be appropriate for the species
and research use described by the protocol, and must be consistent with
the current recommendations of the
American Veterinary Medical Association
Guidelines on Euthanasia.
The NIH Guide defines euthanasia as "the procedure of killing
animals rapidly and painlessly". University of California, Riverside
euthanasia guidelines, summarized in the
Office of the Campus
Veterinarian
follow those established by the American Veterinary Medical Association
Panel on Euthanasia. Euthanasia techniques must be reviewed and approved
by the Institutional Animal Care and Use Committee (IACUC) during review
and approval of the Animal Use Protocol Form.
Euthanasia must be carried out by personnel properly trained in the
procedure being used. While decapitation and cervical dislocation may be
humane when administered by properly trained personnel, animal use
protocols proposing these techniques without sedation or anesthesia must
include the scientific rationale justifying this exclusion. Measures
should be taken to ensure that euthanasia is performed in a way that
minimizes reactions among other animals that may be present.
Gentle, careful handling of subject animals is of the utmost importance
during the procedure in order to minimize distress to the animal, as well
as to the operator. An unconscious animal does not perceive pain.
Appropriately conducted procedures which render the cerebral cortex
nonfunctional by means such as hypoxia or drug induced anesthesia
eliminate perception of pain. Reflex motor activity may be present in an
animal with a nonfunctional cerebral cortex, but pain is not perceived.
Proper euthanasia technique includes a follow-up exam to confirm the
absence of a heartbeat, which is a reliable indicator of death. Monitoring
respiration is not considered sufficient since with some euthanasia
techniques heartbeat may be maintained after visible respiration has
ceased. Decapitation, cervical dislocation or thoracotomy should be used
after administration of euthanatizing drugs to insure that animals do not
revive.
The need to minimize fear and apprehension must be considered in
determining the method of euthanasia. Distress vocalizations, fearful
behavior, and release of certain odors or pheromones by a frightened
animal may cause anxiety and apprehension in other animals. Therefore,
whenever possible, animals should not be exposed to euthanasia of others,
especially of their own species. The resultant distress may lead to
physiologic changes in other animals, such as a release of hormones, which
may effect research results. The acceptable methods of euthanasia vary
aesthetically. Personnel's perception must often be considered in addition
to experimental requirements when a method of euthanasia is chosen.
The following are methods and procedures which are accepted by the
IACUC for humane killing of animals. In general these are the same as
recommendations of the American Veterinary Medical Association (AVMA)
Panel on Euthanasia, 1986. Methods other than those generally approved may
be approved with adequate justification based on scientific need and
demonstration that the method chosen produces the minimum amount of animal
pain and distress consistent with experimental requirements. Also included
is a list of unacceptable methods deemed inhumane or dangerous to
personnel and to other animals.
Many of the approved methods of euthanasia require technical
proficiency for proper conduct and should not be attempted without prior
training (e.g., intravascular injections and physical methods). Other
methods, such as carbon dioxide inhalation and electrocution, may only be
used with properly designed equipment. Trained personnel in the ACU are
available to perform or assist in the performance of animal euthanasia.
Carbon Dioxide
Carbon dioxide inhalation can be
suitable for all species, provided acceptable equipment is used.
Practically, its use is limited to rodents and other mammals weighing less
than about 500 grams. Compressed CO2 from cylinders is the only acceptable
source. Dry ice is no longer permitted as a CO2 source.
Chamber method - This method employs a transparent top-opening chamber which is
charged with CO2 prior to introducing the animal(s).
Please note:
An opaque chamber is not acceptable as
this prevents the visual assessment of the animal and may prolong the
suffering of the animal.
After the animal(s)
are placed in the chamber, a slow flow of CO2 should be continued for a
few minutes to maintain a high concentration at the bottom of the chamber.
After breathing has stopped and the animal(s) are unconscious,
euthanasia may be completed by any of the following procedures:
- continued exposure to CO2 for 10-30 minutes after breathing has
stopped (newborn of most species are more resistant than adults to CO2
and will require exposure times 2-3 times as long to assure death.);
- exsanguination;
- cervical dislocation;
- administration of injectable anesthetic or euthanasia preparation.
Induction of general anesthesia followed by death without regaining
consciousness is acceptable. Inhalation of anesthetic gas or injection of
parenteral general anesthetic agents are suitable for most species
including pig, dog, cat, rodents, rabbits, birds and primates. Ether is
extremely flammable and not acceptable unless used with adequate
protection against the risk of explosion. After anesthesia is induced by
usual means, euthanasia may be completed by any of the following
procedures:
- continued exposure to CO2 for 10-30 minutes after breathing has
stopped (newborn of most species are more resistant than adults to CO2
and will require exposure times 2-3 times as long to assure death.);
- exsanguination;
- cervical dislocation (in small animals less than 200 gm);
- administration of injectable anesthetic or euthanasia preparation.
Barbiturates such as pentobarbital or barbiturate combinations
formulated for animal euthanasia are suitable for most species. Such
agents include Beuthanasia-D (Burns-Biotec) and Sleepaway (Fort Dodge
Laboratories). Detailed records of use of these combinations must be
maintained because these are controlled substances. Typically the dosage
for euthanasia is three times the dose required for anesthesia.
Cervical dislocation (dislocation of the neck) is a simple and humane method of killing mice and
small rats (<125 gms). Although not required, the use of sedation or
anesthesia prior to euthanasia is encouraged. If sedation or anesthesia
prior to cervical dislocation is not used the investigator must provide
scientific justification for its exclusion. In the case of mice and small
rats, the animal is held by its tail and placed on a surface that it can
grip, then it will stretch itself out so that a pencil or similar object
can be placed firmly across the back of the neck. A sharp pull on the base
of the tail will then dislocate the neck.
Decapitation is acceptable for
rodents, rabbits and birds, and other animals of similar size. This
requires special equipment and is aesthetically unacceptable to many.
Because of electroencephalographic evidence suggesting continued cortical
activity after decapitation the Report of the American Veterinary Medical
Association Panel on Euthanasia recommends that animals be anesthetized
when decapitated or that the investigator provide scientific justification
for the use of this technique without anesthesia or sedation. While the
use of anesthetics prior to decapitation is preferred, the scientific
justification for decapitation may precludes their use. In lieu of the use
of anesthetics, prior recommendations have required the immediate
placement of the decapitated head into liquid nitrogen. This is still an
acceptable alternative however it is not required in part because of the
human safety risk.
| Method |
Rabbits & Rodents <125gm |
Rabbits & Rodents >125 gm and < 1 kg |
| Carbon dioxide |
Method of choice |
Method of choice |
| Barbiturate overdose (iv) |
Method of choice |
Method of choice |
| Barbiturate overdose (ip) |
Method of choice |
Method of choice |
| Anesthetic overdose |
Method of choice |
Method of choice |
| Exsanguination in anesthetized animal |
Other acceptable method |
Other acceptable method |
| KCL (iv) in anesthetized animal |
Not recommended due to technical difficulty |
Not recommended due to technical difficulty |
| Decapitation in sedated or anesthetized
animal |
Other acceptable method |
Other acceptable method |
| Cervical dislocation in anesthetized or
sedated animal |
Other acceptable method |
Other acceptable method |
| Decapitation in awake animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
| Cervical dislocation in awake animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
| Stunning in an awake animal |
Unacceptable method of euthanasia |
Unacceptable method of euthanasia |
| Method |
Rabbits/Rodents > 1 kg but < 5 kg |
| Carbon dioxide |
Other acceptable method |
| Barbiturate overdose (iv) |
Method of choice |
| Barbiturate overdose (ip) |
Other acceptable method |
| Anesthetic overdose |
Method of choice |
| Exsanguination in anesthetized animal |
Other acceptable method |
| KCL (iv) in anesthetized animal |
Other acceptable method |
| Decapitation in sedated or anesthetized
animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
| Cervical dislocation in anesthetized or
sedated animal |
Unacceptable method of euthanasia |
| Decapitation in awake animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
| Cervical dislocation in awake animal |
Unacceptable method of euthanasia |
| Stunning in an awake animal |
Unacceptable method of euthanasia |
| Method |
Avian species |
| Carbon dioxide |
Other acceptable method |
| Barbiturate overdose (iv) |
Method of choice |
| Barbiturate overdose (ip) |
Method of choice |
| Anesthetic overdose |
Method of choice |
| Exsanguination in anesthetized animal |
Other acceptable method |
| KCL (iv) in anesthetized animal |
Other acceptable method |
| Decapitation in sedated or anesthetized
animal |
Other acceptable method |
| Cervical dislocation in anesthetized or
sedated animal |
Other acceptable method |
| Decapitation in awake animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
| Cervical dislocation in awake animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
| Stunning in an awake animal |
Acceptable only with scientific justification in
writing on the Animal Care and Use Review Form |
Amphibians
- Inhalant anesthetics
- CO2
- Barbiturates
- Tricaine methane sulfonate (MS222)
- Double pithing
- Benzocaine
- Conditionally acceptable - Single pithing; stunning and
decapitation; decapitation and pithing
Fish
- Tricaine methane sulfonate (MS222)
- Benzocaine
- Barbiturates
- Inhalant anesthetics
- CO2
- 2-phenoxyethanol
- Conditionally acceptable - stunning followed by decapitation/pithing;
decapitation and pithing
Reptiles
- Barbiturates
- Inhalant anesthetics (in appropriate species)
- CO2 (in appropriate species)
- Conditionally acceptable - stunning and decapitation; decapitation
and pithing
| Species |
IV route (mg/kg) |
IP route (mg/kg) |
| Mouse |
150 |
150 |
| Rat |
150 |
150 |
| Rabbit |
100 |
150 |
| Guinea Pig |
120 |
150 |
| Hamster |
150 |
150 |
| Sheep |
90 |
|
| Goat |
90 |
|
| Chicken |
150 |
150 |
| Swine |
90 |
|
| Ferret |
120 |
12035 |
NOTE: Euthanasia with barbiturates usually requires approximately 3X
the anesthetic dosage.
Adapted from the report of the American Veterinary Medical Association
Panel on Euthanasia (J. Am. Vet. Med. Assn. 202:229-249, 1993). The JAVMA
article provides the rationale for these recommendations. The University
Animal Care and Use Committee reviewed approved this table.
These methods are in accordance with humane euthanasia as defined by
the Federal Animal Welfare Act (54 FR 36112-36163).
Methods always unacceptable in an awake animal include: potassium
chloride, magnesium sulfate, strychnine, neuromuscular blocking agents,
exsanguination, air embolism, and chloroform.
Comments on Recommended Agents and Methods of Euthanasia
INHALANT ANESTHETICS - Because in the liquid state most inhalant
anesthetics act as topical irritants, animals should be exposed to the
vapors of the anesthetic only. Air or oxygen must be provided during the
induction period. All agents are given "to effect" until
respiratory and cardiac arrest occurs.
Halothane and isoflurane have the most rapid action, and since
halothane is better tolerated, it is preferred. Methoxyflurane is
less suitable, due to its slow effect and poor market availability.
Care should be taken to minimize personnel exposure to vapors.
Ether is acceptable but not recommended because it poses an explosive
hazard and is a respiratory irritant that is considered stressful to
animals. It cannot be used in the UCR centralized facilities, and special
precautions must be taken when used elsewhere. Administration should be
performed in a fume hood, and signs indicating that ether is present or in
use should be posted conspicuously. To avoid explosions, the carcasses of
ether-killed animals should be stored in explosion-safe refrigerators or
freezers, and should not be incinerated until the ether is removed by
aeration in a hood. Methoxyflurane is a similar but non-explosive and less
irritating agent which is recommended as a substitute for ether.
NON-ANESTHETIC GASES - Most agents in this category require the use of
special equipment.
CO2 - Carbon dioxide is the preferred technique for
euthanizing rodents and other small laboratory animals. Use of a sealed
chamber filled by a compressed gas cylinder is required. CO2
generated by other methods, such as from dry ice, is unacceptable because
gas flow can't be regulated precisely. Chambers should not be
overcrowded. CO2 concentration of 70% or more should be
utilized for euthanasia. Because CO2 can act as a reversible
anesthetic, it is imperative that the animals be kept in the chamber for
several minutes after respiratory arrest. Where possible, death
should be verified by absence of a palpable heart beat. Due to physiologic
characteristics, neonates require prolonged exposure to the gas. For
more information, see the IACUC
guidelines on using CO2 for rodent euthanasia.
Nitrogen or carbon monoxide may be acceptable but are not recommended
and require special equipment.
PHARMACOLOGICAL AGENTS - Use of these agents requires adequate
restraint and mastery of appropriate injection techniques.
Barbiturates such as pentobarbital are acceptable for mammalian species
and birds. These drugs should be administered intravenously (IV) except in
rodents where intraperitoneal (IP) administration is an acceptable
alternative. Sodium pentobarbital (Nembutal) is the most common
barbiturate agent for euthanasia. The dosage is usually at least twice
that required for anesthesia, and ranges from 85 mg/kg for larger species
to 200 mg/kg for some rodents. A dosage of 120 mg/kg is sufficient for
most species, but more should be given if death does not ensue. Commercial
barbiturate euthanasia formulations as are also appropriate, and should be
used following label directions (e.g., 1 ml/lb for Beuthanasia-D*). Sodium
pentobarbital is a Class II drug which is regulated by the Drug
Enforcement Agency. Personnel using this agent are required to store it in
a locked location and maintain records which include the date and amount
of use.
Chloral hydrate is not recommended, but may be used in ruminants and
swine when administered I.V. at 900 mg/kg, but only after sedation with
another drug.
Neither magnesium sulfate nor potassium chloride (KCl) can be used as a
sole agent of euthanasia. Overdose with KCl is permissible in an
anesthetized animal. Concentrated KCl should be given rapidly IV until
rising serum potassium levels result in cardiac arrest.
Tricaine methane sulfonate (MS222) can be used either as an injectable
agent (200-300 mg/kg of a 1% buffered solution) or as an immersion bath (2
mg/ml in H2O) for amphibians and fish. The immersion time needed to assure
death can range from 20 minutes to three hours, so it may be advantageous
to use MS222 as an anesthetic followed by a physical method of euthanasia.
Benzocaine immersion (100-200 mg/liter H2O) is also acceptable.
Neuromuscular blocking drugs are absolutely condemned for use as
euthanasia agents.
PHYSICAL METHODS - These methods require that the user have experience
and skill in the techniques to be used.
Exsanguination is acceptable for all species if animal is first
rendered unconscious by another agent.
Cervical dislocation is acceptable for mice, birds, rats (< 200 gm)
and rabbits (< 1 Kg), but proper technique is essential. It is
therefore recommended that animals be first sedated with another agent
(carbon dioxide, pentobarbital or halothane are suggested). Its use as a
sole means of euthanasia requires scientific justification and IACUC
approval for more information, see the IACUC
guidelines on the use of cervical dislocation for euthanasia of
rodents. Decapitation with proper equipment may be performed on small mammals or
birds after the animal has been sedated or lightly anesthetized (carbon
dioxide, pentobarbital or halothane are suggested). Decapitation of fish,
amphibians and reptiles should be followed by pithing. Use as a sole means
of euthanasia in any species requires scientific justification and IACUC
approval. Decapitation should generally be used only when study design
requires it due to the potential hazard to personnel. Many species react
adversely to the smell of blood, so animals should not be decapitated in
the presence of other animals and the person performing decapitation
should change gloves and/or wash hands between animals.
Pithing of both the brain and the spinal cord (double pithing)
may be used as the sole means of euthanasia in frogs of the genus Rana
or other amphibians with anatomic features that facilitate easy access to
the central nervous system. In all other amphibian and reptile species
pithing should be followed by decapitation.
Under very specialized circumstances, stunning, rapid freezing or air
embolism (under anesthesia) may be allowed in small species if research
needs make it necessary and there are no available alternatives.
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