Hypothermia During General Anesthesia Interferes with Pain Assessment in Laboratory Rats (Rattus norvegicus)

Accurate pain assessment methods are necessary to ensure animal welfare and reliable data collection in animal research. The Rat Grimace Scale (RGS), a facial expression pain scale, allows effective identification of pain. However, the potential confounds of this method remain mostly unexplored. General anesthesia, which is used in many laboratory procedures, suppresses thermoregulation and results in hypothermia. We investigated the effects of isoflurane-induced hypothermia on RGS scores. Twenty (10 male and 10 female) Sprague-Dawley rats each received 30 min of anesthesia, followed by 30 min of observation after the return of sternal recumbency. Rats were randomized to receive warming with an electric heating pad or no warming during both periods. Unwarmed rats became hypothermic within 15 min after isoflurane exposure began and returned to normothermia within 15 min after returning to sternal recumbency. Warmed rats did not deviate from the normothermic range. The RGS scores of unwarmed rats were significantly higher than baseline levels for 3 h after anesthesia and were higher than those of warmed rats at 5 and 180 min after anesthesia. Hypothermia resulted in a larger proportion of rats crossing a predetermined analgesic intervention threshold. Our findings show that hypothermia induced by isoflurane anesthesia presents a confound to accurate RGS scoring. These results emphasize the importance of maintaining normothermia to avoid inflated pain scores and to obtain accurate pain assessment.