ObjectiveTo evaluate the sedative effects of two doses of alfaxalone when added to a combination of dexmedetomidine and methadone injected intramuscularly (IM) in healthy Beagles. Study designRandomized, blinded, crossover, experimental study. AnimalsA group of six adult Beagles. MethodsDogs were sedated on three different occasions with IM dexmedetomidine (3 μg kg–1) and methadone (0.3 mg kg–1) combined with two doses of alfaxalone (0.5 and 1 mg kg–1; A0.5 and A1, respectively) or saline (A0). Quality of sedation, response to tail clamping and rectal temperature were recorded at baseline, 5, 15, 25, 35 and 45 minutes. Pulse and respiratory rates, oxygen saturation of haemoglobin (SpO2) and noninvasive blood pressure (NIBP) were recorded every 5 minutes. Onset of sedation and duration of recumbency, response to venous catheterization and recovery quality were assessed. Physiological variables (analysis of variance) were analysed between treatments and within treatments compared with baseline (Student t test). Nonparametric data were analysed using Friedman and Cochran’s Q tests. Significance was p < 0.05. ResultsSedation scores were significantly higher when alfaxalone was co-administered (area under the curve; p = 0.024, A0.5; p = 0.019, A1), with no differences between doses. Onset of sedation was similar, but duration of recumbency was longer in A0.5 than in A0 [median (minimum–maximum), 43 (35–54) versus 30 (20–47) minutes, p = 0.018], but not in A1. Response to venous catheterization and tail clamping, and quality of recovery (acceptable) presented no differences between treatments. A decrease in all physiological variables (compared with baseline) was observed, except for NIBP, with no differences between treatments. All dogs required oxygen supplementation due to reduced SpO2. Conclusions and clinical relevanceAdding alfaxalone to methadone and dexmedetomidine enhanced sedation and duration of recumbency. Although cardiopulmonary depression was limited, oxygen supplementation is advisable.