Microbial strains and indigenous microbiota in soil slurries have been reported to use electrons from electrodes for nitrate (NO3-) reduction. However, few studies have confirmed this in a soil matrix hitherto. This study investigated if, and how, an electric potential affected NO3- reduction in a soil matrix. The results showed that, compared to a control treatment, applying an electric potential of -0.5 V versus the standard hydrogen electrode (SHE) significantly increased the relative abundance of NO3--reducing microbes (e.g., Alcaligenaceae and Pseudomonadaceae) and the abundances of the nrfA, nirK, nirS, and nosZ genes in soil matrices. Meanwhile, the electric potential treatment doubled the NO3- reduction rate and significantly increased the rates of production of ammonium (NH4+), dinitrogen (N2), and nitrous oxide (N2O). The amount of NO3--N reduced under the electric potential treatment was comparable to the sum of the amounts of N observed in the increased N2O, N2, NH4+, and nitrite (NO2-) pools. An open-air experiment showed that the electric potential treatment promoted soil NO3- reduction with a spatial scale of at least 38 cm. These results demonstrated that an electric potential treatment could enhance NO3- reduction via both denitrification and dissimilatory NO3- reduction to ammonium (DNRA) in the soil matrix. The mechanisms revealed in this study have implications for the future development of potential techniques for enhancing NO3- reduction in the vadose zone and consequently reducing the risk of NO3- leaching.