This experiment investigated the fatigue induced by a 24-h running exercise (24TR) and particularly aimed at testing the hypothesis that the central component would be the main mechanism responsible for neuromuscular fatigue. Neuromuscular function evaluation was performed before, every 4 h during, and at the end of the 24TR on 12 experienced ultramarathon runners. It consisted of a determination of the maximal voluntary contractions (MVC) of the knee extensors (KE) and plantar flexors (PF), the maximal voluntary activation (%VA) of the KE and PF, and the maximal compound muscle action potential amplitude (M_max) on the soleus and vastus lateralis. Tetanic stimulations also were delivered to evaluate the presence of low-frequency fatigue and the KE maximal muscle force production ability. Strength loss occurred throughout the exercise, with large changes observed after 24TR in MVC for both the KE and PF muscles (−40.9 ± 17.0 and −30.3 ± 12.5%, respectively; P < 0.001) together with marked reductions of %VA (−33.0 ± 21.8 and −14.8 ± 18.9%, respectively; P < 0.001). A reduction of M_max amplitude was observed only on soleus, and no low-frequency fatigue was observed for any muscle group. Finally, KE maximal force production ability was reduced to a moderate extent at the end of the 24TR (−10.2%; P < 0.001), but these alterations were highly variable ( ± 15.7%). These results suggest that central factors are mainly responsible for the large maximal muscle torque reduction after ultraendurance running, especially on the KE muscles. Neural drive reduction may have contributed to the relative preservation of peripheral function and also affected the evolution of the running speed during the 24TR.