Ketamine, an open-channel blocker of the N-methyl-D-aspartate receptor (NMDAR), has been shown to have rapid and sustained antidepressant effects after a single sub-anesthetic dose. Among its two enantiomers, (S)-ketamine acts more potently at NMDARs than (R)-ketamine, and its use as an antidepressant was approved by the FDA in 2019. Given that the mechanism underlying ketamine’s sustained antidepressant action remains unclear, we investigated if (S)-ketamine has any long-term effect on NMDAR-mediated synaptic transmission. In extracellular recordings from the CA1 region of mouse hippocampal slices, we found that a 20 min application of 10 µM (S)-ketamine resulted in a long-term depression (LTD), lasting >2 h, of NMDAR-mediated synaptic responses recorded in low (0.2 mM) magnesium conditions. This LTD effect was eliminated when (S)-ketamine was applied in the presence of 1 mM magnesium, suggesting that binding of (S)-ketamine to the pore of NMDARs is required for its induction. In contrast, (S)-ketamine’s effects were not dependent on the electrical stimulation protocol, since neither the acute nor long-term inhibition of NMDAR-mediated responses were affected by an increase or pause in test-frequency stimulation. Finally, we found that the LTD effect of (S)-ketamine was not reversed by a subsequent application of 1 mM magnesium, whereas the slow wash-out of memantine (another NMDAR channel blocker) was accelerated by magnesium. Together, our results suggest that brief exposure to (S)-ketamine can lead to long-lasting changes in NMDAR-dependent synaptic function, which may be relevant to ketamine’s antidepressant action.