The neuromodulatory endocannabinoid system is a promising target for therapeutic interventions. One of well-known behavioral effects of cannabinoid CB1 receptor activation with exogenous ligands such as THC is the inhibition of locomotor activity. However, the behavioral effects of endogenous cannabinoids have not been extensively studied. Despite broad medicinal and recreational use of cannabinoids, little is known about their effects on fine movements, critical for therapeutic applications. Such fine motor function is only resolvable with the precision of marker-based motion capture, which so far has not been available in freely moving mice. Therefore, we have adapted a high-speed, high-resolution marker-based 3D motion capture system to track movement (3D trajectories and speed of markers) during voluntary locomotor tasks in mice. Here we show that enhancing endocannabinoid signaling produces bidirectional effects on locomotor behavior that differ from those caused by exogenous cannabinoid receptor agonist, in unrestricted mice. Given our expectation of subtle rather than dramatic changes with the low and potentially therapeutic doses, we employed a sensitive and accurate marker-based 3D motion capture system to resolve the finest differences in full-body kinematic signatures of mice exploring horizontal and vertical environments. We found that selective upregulation of either of the two main endocannabinoids 2-arachidonoylglycerol (2-AG) or anandamide (AEA) with selective inhibitors of their degradation (MJN110 and PF3845, respectively), produced opposite effects: PF3845 suppressed locomotor activity whereas MJN110 enhanced it. Furthermore, while low doses of the synthetic cannabinoid agonist CP55,940 decreased locomotion in a an open field exploration task as expected, it surprisingly did not affect performance in a vertical climbing task. The results show that the effects of cannabinoid signaling on behavior are not predominantly inhibitory as commonly assumed. Furthermore, we found that the invariant microstructure of locomotory behavior remains unchanged under all treatments, pointing towards motivational rather than motor-related mechanisms of action.