Rehabilitation exercises play a crucial role in veterinary medicine. Among these exercises, sit-to-stand and stand-to-sit transitions are fundamental tasks. However, the activation patterns of hindlimb muscles during these transitions remain inadequately characterized. This study aimed to evaluate the activity patterns of the gluteus medius (GM), vastus lateralis (VL), and gastrocnemius (GC) in six healthy Beagle dogs. After treadmill acclimatization at 0.7 m/s, each dog performed 10 gait cycles at a walk, followed by five repetitions each of sit-to-stand and stand-to-sit transitions. Surface electromyography (sEMG) data were processed to obtain normalized peak amplitudes (peak %EMG), defined as the maximum amplitude during each transition, expressed as a percentage of the mean peak amplitude from ten walking gait cycles per dog. The VL exhibited significantly greater peak %EMG during sit-to-stand (337 ± 165 %) than stand-to-sit (215 ± 71 %) (p < 0.05), whereas the GC showed the opposite pattern, with significantly higher peak %EMG during stand-to-sit (206 ± 46 %) than sit-to-stand (125 ± 29 %). The GM showed no significant difference between transitions. Additionally, all three muscles functioned in a coordinated manner during the stand-to-sit transition. These findings highlight the critical role of the VL in generating stifle extension during the early phase of sit-to-stand, particularly relevant for rehabilitation strategies targeting stifle joint function. Conversely, the cooperative activation during stand-to-sit suggests this movement may serve as an effective rehabilitation exercise for improving coordinated hindlimb control. These insights provide objective data to refine targeted rehabilitation protocols and optimize functional recovery in canine patients.