O-GlcNAcylation, a post-translational modification involving O-linkage of β-N-acetylglucosamine to Ser/Thr residues on target proteins, is increasingly recognized as a critical regulator of synaptic function. Enzymes that catalyze O-GlcNAcylation are found at both presynaptic and postsynaptic sites, and O-GlcNAcylated proteins localize to synaptosomes. An acute increase in O-GlcNAcylation can affect neuronal communication by inducing long-term depression (LTD) of excitatory transmission at hippocampal CA3-CA1 synapses, as well as suppressing hyperexcitable circuits in vitro and in vivo. Despite these findings, to date, no studies have directly examined how O-GlcNAcylation modulates the efficacy of inhibitory neurotransmission. Here we show an acute increase in O-GlcNAc dampens GABAergic currents onto principal cells in rodent hippocampus likely through a postsynaptic mechanism, and has a variable effect on the excitation/inhibition balance. The overall effect of increased O-GlcNAc is reduced synaptically-driven spike probability via synaptic depression and decreased intrinsic excitability. Our results position O-GlcNAcylation as a novel regulator of the overall excitation/inhibition balance and neuronal output. Synaptic integration and spike initiation in neurons is controlled by synaptic inhibition, which strongly influences neuronal output and information processing 1. Importantly, the balance of excitation to inhibition (E/I) is crucial to the proper functioning of circuits, and E/I imbalances have been implicated in a number of neurodevelopmental disorders and neurodegenerative diseases including schizophrenia, autism spectrum disorders, and Alzheimer's disease 2-5. Thus, understanding the mechanisms that modulate the strength of inhibitory transmission is fundamental to unraveling how neuronal circuits function in normal and disease states. Fast inhibitory transmission in the central nervous system is mediated by GABA A receptors (GABA A Rs), which are pentameric ligand-gated ion channels. The strength of this inhibition can be rapidly up-or down-regulated by post-translational modifications including ubiquitination 6 , palmitoylation 7,8 , and phosphorylation 9 of GABA A R subunits and/or associated proteins, which can alter channel function, trafficking, or stability at the membrane. While there is a vast body of literature on these post-translational modifications, O-GlcNAcylation, involving the O-linkage of β-N-acetylglucosamine (O-GlcNAc) to Ser/Thr residues on target proteins, remains severely understudied, with no examination to date of the effect of protein O-GlcNAcylation on inhibitory synapse physiology. Addition and removal of O-GlcNAc are catalyzed by the single pair of enzymes O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). O-GlcNAcylation is essential, as genetic deletion of OGT and OGA are lethal 10,11. O-GlcNAcylation is metabolically-regulated and highly dynamic; global changes reversibly occur within minutes, and are dictated by availability of UDP-GlcNAc, which is synthesized from glucos...