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cannot be relieved be painlessly euthanized at the end of the procedure or, if appropriate, during the procedure” (AWR 2.31 (d)(1)(v)). The authoring committee notes that requirement does not preclude the development and study of animal models of chronic or persistent pain (AWR 2.31 (d)(1)(iv)(A)); however, animals in severe or intolerable pain should be euthanized. Additionally, animals in studies in which severe pain develops as an unintended consequence should be euthanized or the manipulation causing the unintended pain should be stopped if that would eliminate the pain.
Training staff members to properly perform euthanasia is essential. Training must include instruction both in the specific technique that will be used and in the recognition and confirmation of death (Close et al., 1996). For example, exposure to carbon dioxide can cause deep narcosis that can appear to be, but is not, death. In such cases, animals that superficially appear to be dead may eventually awaken; this arousal can occur after the disposal of carcasses into refrigerators or freezers. The occurrence of death after exposure to carbon dioxide must be confirmed based on careful assessment of the animal for unambiguous signs of death, such as cardiac arrest or fixed, dilated pupils. If an animal is removed from a CO2 chamber before death occurs, the animal either can be returned to the chamber for additional exposure, or, if it is unconscious and nonresponsive, can be humanely euthanized via a physical method (e.g., decapitation or cervical dislocation).
In some species, fear induces animals to become immobile; such immobility must be distinguished from loss of consciousness or death (Close et al., 1996). Some animals release pheromones indicative of fear or distress, which may in turn stress or otherwise disturb other animals (NRC, 1996). Therefore, euthanasia should ideally be performed in an area separate from other animals. However, a recent study suggests that witnessing decapitation may be no more disruptive to Sprague-Dawley rats than other common procedures, such as cage changing, restraint, and injections (Sharp et al., 2003).
Methods of euthanasia that are commonly used in neuroscience research include decapitation, cervical dislocation, carbon dioxide inhalation, and barbiturate overdose. Focused high-intensity microwave irradiation is also used in some cases for measurement of highly labile substances or metabolites (for example, Delaney and Geiger, 1996; Ikarashi et al., 1985; Mayne et al., 1999; Nylander et al., 1997; Theodorsson et al., 1990; Todd et al., 1993). The recommendations of the AVMA Panel on Euthanasia (AVMA, 2001) should be followed unless deviation is justified for scientific or medical reasons (PHS Policy IV(C)(1)(g); APHIS/ AC Policy 3; Guide, p. 65). However, the AVMA Panel consensus concerning the need for anesthetization prior to decapitation is controversial and is based largely on one publication (Mikeska and Klemm, 1975). Other authors dispute the conclusions drawn from that study, concluding instead that hippocampal and cortical responses to decapitation do not reflect consciousness or resemble the response to painful stimuli (Allred and Berntson, 1986; Vanderwolf et al., 1988),