Public Health Service Policy states, “Methods of euthanasia will be consistent with the recommendations of the American Veterinary Medical Association (AVMA) Panel on Euthanasia, unless a deviation is justified for scientific reasons in writing by the investigator.”

Policy:

Investigators must use methods of euthanasia for mice and rats which are consistent with the AVMA Panel on Euthanasia.  Mice and rats must be euthanized by trained personnel using appropriate technique, equipment and agents. Euthanasia must not be performed in an animal room.

Do not place more than a single layer of adults in a cage to be euthanized at one time.  For example, in a small shoebox cage, the maximum number of mice would be approximately ten.

Chambers must not be overcrowded.  In this regard, it is important to also consider that mixing unfamiliar animals in the same container may be distressful.  All animals in the chamber must be able to make normal postural adjustments.

If using the open drop technique with an inhalant anesthesia such as Isoflurane or Halothane (not CO2), you must conduct the anesthetic overdose in a fume hood that vents to the outside of the building.

Animals must be left in the container until clinical death has been assured by observation of at least two of the following signs: movement cessation, glazed appearance of the eye and membranes, and lack of heartbeat.

Remove each group of deceased mice from the euthanasia chamber before introducing a group of live mice.

Death must be confirmed.  Adjunctive methods, including decapitation, cervical dislocation, and/or pneumothorax are recommended for all species. 

Rats must receive a bilateral thoracotomy or decapitation to ensure death.

Applicability:

This policy applies to all mice and rats euthanized.

Instructions:

Fetuses:

A.        Fetuses up to 14 days in gestation:  Neural development at this stage is minimal and pain perception is considered unlikely.  Euthanasia of the mother or removal of the fetus ensures rapid death of the fetus due to loss of blood supply.

B.         Fetuses 15 days in gestation to birth:  The literature on the development of pain pathways suggests the possibility of pain perception at this time.  Acceptable methods of euthanasia are listed below.

•         Decapitation with sharp, surgical scissors.

•         Rapid freezing by immersion in liquid nitrogen.

•         Removal of vital organs and chemical fixation by immersion or perfusion requires prior anesthesia by hypothermia, injection of chemical anesthetic, or deep anesthesia of the mother with a chemical agent that crosses the placenta (e.g. pentobarbital). Consult with ARC veterinarian staff for fetal sensitivity to specific anesthetic agents not listed above.

When fetuses are not required for study, the method chosen for euthanasia of a pregnant mother 15 or more days post coitum must ensure rapid death of the fetus.  Acceptable methods of euthanasia are:

•         Pentobarbital overdose of the mother.

•         CO2 hypoxia euthanasia of the mother followed by exteriorization of the fetus and rapid euthanasia by one of the methods outlined above.

Neonates:

A.        Up to 14 days of age: CO2 hypoxia must be followed by a second physical method of euthanasia.  Acceptable methods include:

•         Decapitation, bilateral thoracotomy, removal of vital organ (requires prior anesthesia).

•         Immersion or perfusion with chemical fixatives (requires prior anesthesia).

•         Rapid freezing by immersion in liquid nitrogen (requires prior anesthesia).

•         Overdose injection of pentobarbital.

Acceptable anesthetics include:

•         Hypothermia (requires a barrier between the body of the animal and the wet ice, do not use dry ice.)

•         CO2 narcosis  (100% for two minutes.)

•         Inhalant Isoflurane/Halothane  (4% concentration or higher for two minutes.)

•         Injection of pentobarbital (120 mg/kg.)

B.       Older than 14 days:  Follow guidelines for adults.

Adults:

A.     Methods requiring prior anesthesia:

•         Decapitation

•         Cervical dislocation

•         Bilateral thoracotomy

•         Removal of vital organ

•         Perfusion with chemical fixatives

B.     Acceptable Methods

•         Inhalant Isoflurane/Halothane overdose

•         Overdose injection of pentobarbital

•         Carbon dioxide

Carbon dioxide requirements:

•         DRY ICE GENERATION OF CO2 GAS IS NOT ALLOWED FOR EUTHANASIA.

•         Compressed CO2 gas in cylinders is the only approved source of carbon dioxide as it allows the inflow of gas to the induction chamber to be controlled.

•         Carbon dioxide narcosis must not be used for anesthesia for survival procedures. 

•         The euthanasia chamber must allow visibility of the animals.

•         Do not pre-charge the chamber.  Follow the ARC approved specific instructions that must be clearly posted near the chamber.

•         Animals should be left in the container until clinical death has been assured by observation of at least two of the following signs: movement cessation, glazed appearance of the eye and membranes, and lack of heartbeat.

It is recommended that you stay with the animals during the euthanasia procedure.

Exceptions:

Any exceptions to this policy must be scientifically justified and approved by the IACUC in an animal protocol.

Contact Information:

For any additional questions concerning this policy, please contact the IACUC Manager at 5-6420.