period (Hofer and Shair, 1992). The significance of the vocalizations is not clear, but they may indicate distress. To reduce possible unintended pain associated with cooling, the technique for inducing hypothermia should include partial insulation of the pup (for example, by wrapping in a latex blanket) (Danneman and Mandrell, 1997).
As with adult animals, assessing the effectiveness of anesthesia in neonates is important before beginning a potentially painful procedure. Adequately anesthetized rat pups will not respond to a light pinch of the foot or tail. Similarly, adequately anesthetized adult rats will not respond to a pinch of the toe or tail.
Opioid drugs provide effective analgesia against thermal, inflammatory, and mechanical pain in neonatal rodents as young as P1 (Barr, 1999; Barr et al., 1992; Helmstetter et al., 1988; Marsh et al., 1999; McLaughlin and Dewey, 1994) and should be considered for use whenever analgesia would be provided for an adult animal. Fentanyl is a recommended analgesic for neonatal dogs and humans because it has less of a respiratory depressant effect than morphine (Luks et al., 1998a).
Neonatal exposure to pain, especially when pain is an unintended outcome, may have developmental effects on the central and peripheral nervous systems and alter behavior and the threshold for pain in adulthood (Anand et al., 1999; Bhutta et al., 2001; Fitzgerald and Beggs, 2001).
Aside from the technical difficulties associated with using very small animals, surgical procedures involving neonatal rodents present such challenges as maternal neglect and cannibalism. As with adult rodents, pups should be kept warm, dry, and well hydrated postoperatively. They should be placed in a warm— not hot—environment until they have regained the ability to right themselves when placed on their backs or sides, after which they should be returned to their mothers. Some rodent mothers (particularly in some strains such as BALB/c mice) will reject or kill their pups under these circumstances. Some steps can be taken to reduce that problem. First, pups should be sufficiently recovered from anesthesia that they are able to right themselves and respond to stimulation. Smearing a pup with bedding and urine from littermates that remained with the mother can be helpful, as can placing the pup in the middle of the litter and allowing it to settle in for a minute or two before reintroducing the mother. Other methods that may work include masking olfactory cues by sprinkling baby powder on mother, pups, and bedding and smearing the pups and the mother’s nose with an aromatic agent, such as Vicks Vapo Rub®.
The following method is cumbersome, but it can greatly improve the rate of successful reunion of mouse pups with their mothers and might be considered when maternal neglect of pups is substantially inhibiting progress of a study: