Dysfunction of prefrontal cortex (PFC), a critical brain region for emotion and cognition, is implicated in many stress-related disorders such as schizophrenia and depression. However, the mechanisms by which emotional stress influences PFC circuitry are poorly understood. By introducing various acute stress paradigms in intact animals, synaptic changes in PFC were detected using combined electrophysiological, biochemical, molecular, immunocytochemical and behavioral approaches. Acute stress produced a long-lasting potentiation of glutamatergic transmission in PFC pyramidal neurons, which was accompanied by increased surface glutamate receptor expression and dendritic spine growth. Molecular knockdown of the stress-activated immediate early genes, serum glucocorticoid kinase (SGK) 1 and 3, blocked the stress modulation of glutamate currents. The activity of small GTPase Rab 4, which mediates receptor recycling between early endosome and plasma membrane, was increased in stressed animals, and underlined the stress increase of glutamate receptor surface delivery. Consistent with the cellular modulation, stressed animals showed enhanced performance in PFC-mediated working memory tasks in a SGK-dependent manner. These findings unravel the cellular basis of acute stress in synaptic and structural plasticity of PFC, which may shed light on better understanding of stress contributes to PFC function under normal and pathological conditions.