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OBJECTIVE To evaluate efficacy and safety of anesthesia with dexmedetomidine-ketamine-midazolam (DKM) in five-striped palm squirrels (Funambulus pennantii). ANIMALS 8 male squirrels. PROCEDURES Squirrels were anesthetized with DKM (dexmedetomidine, 0.1 mg/kg; ketamine hydrochloride, 30 mg/kg; and midazolam, 0.75 mg/kg) administered IM. Atipamezole (0.15 mg/kg) and flumazenil (0.1 mg/kg) were administered IM 40 minutes after induction of anesthesia. Vital signs and responses were recorded every 5 minutes during anesthesia. RESULTS Anesthetic induction and recovery from anesthesia were rapid and without complications in all squirrels. Median anesthetic induction time was 67.5 seconds (interquartile [25th to 75th percentile] range, 5.5 seconds), and mean ± SD recovery time after drug reversal was 147 ± 79 seconds. Heart rate, respiratory rate, and rectal temperature significantly decreased during the anesthetic period. All squirrels became hypothermic by 40 minutes after induction. The righting reflex was absent during the 40-minute anesthetic period in all squirrels, with variable responses for the palpebral reflex, jaw tone, forelimb withdrawal reflex, and hind limb withdrawal reflex. Only 2 of 8 squirrels had loss of the limb withdrawal reflex in both the forelimbs and hind limbs from anesthetic induction to 25 minutes after induction. CONCLUSIONS AND CLINICAL RELEVANCE DKM appeared to provide safe and effective anesthesia in five-striped palm squirrels, but oxygen and thermal support were indicated. At the doses administered, deep surgical anesthesia was not consistently achieved, and anesthetic depth of individual squirrels must be determined before surgical procedures are performed in palm squirrels anesthetized with this drug combination.