We examined the effect of an acute bout of treadmill running with rubber tube (RT) and without rubber tube (NT) elastic constraints on electromyographic (EMG), 3D kinematics variability, and blood lactate concentration (LA). In the RT test, the constraints were attached to the hips and ankles. The selected variables were compared between 30 min of NT running and 30 minutes of RT running in 13 healthy recreationally trained male runners who had no prior exposure to RT. Statistical analysis revealed significantly higher EMG variability (p < 0.01) and muscle activity (p < 0.05) during RT compared to NT that decreased over time approaching NT, indicating movement pattern adaptation. 3D-kinematics and their variability remained generally unaltered. Changes occurred predominantly in the sagittal plane, specifically to the knee and the swing. A significant increase in LA was measured at the end of RT (p < 0.05). These findings suggest that RT running influences muscle recruitment and variability, but has only a minor influence on kinematics. Changes in LA were significant, although relatively small. The observed adaptations in EMG and kinematics suggest that the RTs provide a possibility to create within movement variability in various sports, and thus, variable training conditions may foster strategies to increase the ability to flexibly adapt to different and new situations.