Water electrolysis using anion exchange membranes ispromisingfor hydrogen production, and Ni-Mo catalysts have shown highactivity for alkaline hydrogen evolution reaction (HER). However,their performance has been mostly tested in a half-cell setup andrarely studied in a single-cell setup with a membrane electrode assembly(MEA) structure, which is used for practical applications. With Ni3Mo as the cathode, a single cell was fabricated using non-noblemetal catalysts exclusively. Interestingly, the activation proceduresignificantly affected the cell performance. The single cell performedbetter than that with the Pt/C catalyst when the Ni(3)Mocatalyst was mildly activated. The distribution of Mo in electrodes,membrane, and electrolytes was estimated, confirming Mo dissolutionfrom the cathode. Once the cell was activated, the cell performancewas stable without degradation in long-term chronopotentiometry operation,but the performance was degraded by sudden voltage change such asimposing open circuit voltage (OCV). The surface structure and reactionmechanism were studied with density functional theory: the Mo-dissolvedNi(3)Mo(101) surface could promote H2O dissociation,while MoO3 stably adsorbed on the surface weakened H* adsorption,promoting HER. This study provides important insights into the developmentof efficient catalysts for large-scale hydrogen production.