Fear, anxiety and stress (FAS) are significant factors in human-animal interactions in veterinary hospital settings. The objective of this study was to determine the effect of aqua-acupuncture (Aqua-AP) compared with trazodone on the mitigation of FAS indicators in healthy, stressed dogs during routine veterinary examination and procedures. Fifty-five dogs exhibiting signs of FAS or history of significant FAS in veterinary hospital settings completed the study. Except for those premedicated with trazodone (Group 4), dogs were randomly assigned to 3 treatment groups. Groups 1 (An-shen) and 2 (GB-21) were treated with Aqua-AP bilaterally, while Group 3 served as untreated controls. Stress data, including heart rate, panting respiratory rate, behavior and saliva cortisol concentration were taken at T₀ (baseline) and T₃₀ (30 minutes post-treatment). Comparison of group outcome data found that Aqua-AP groups had significant decreases for heart rate (p = 0.004 and p = 0.0005, respectively) and behavior score (p = 0.0002 and p = 0.008, respectively) from T₀ to T₃₀, and posttreatment values were not higher than the pharmaceutical-treated subjects (noninferiority). In addition, Group 1 had significant reduction (p = 0.04) in incidence of panting respiratory rate. No significant stress parameter changes were observed in controls. The cortisol concentration change was not significant in any group, but the post-treatment cortisol concentration in both Aqua-AP groups was not greater than the trazodone group (p = 0.004 and p = 0.018, respectively). These results suggest that Aqua-AP at An-shen or GB-21 is effective in mitigating FAS in healthy dogs during routine veterinary examination and demonstrates noninferiority to premedication with trazadone.