Introduction: The purpose of this study was to determine the effects of vibration (VIB) on motor unit (MU) behavior of the vastus lateralis (VL) muscle during a 40% maximal voluntary contraction (MVC). Methods: Eleven healthy (age 21.3 ± 2.6 years) individuals participated in the study. Surface electromyography (EMG) data were recorded from the VL during isometric trapezoidal muscle contractions at 40% MVC. Firing events of single MUs and EMG amplitude were reported for the first, middle, and final seconds of a 12‐second steady force segment at 40% MVC. VIB was applied at 55 Hz to the patellar tendon for 15 minutes before and continued throughout the remainder of testing (VIB) or remained off (CON). Results: There were significant increases in MU firing rates during VIB in comparison to CON and no differences in EMG amplitude between VIB and CON. Conclusion: The VIB‐mediated reduction in muscle spindle function altered MU behavior at 40% MVC. Muscle Nerve 55: 500–507, 2017