Transient genetic manipulation of human neurons without chromosomal integration of the transgene would be valuable but has been challenging due to the quiescent nature of these postmitotic cells. In this study, we developed a set of baculoviral vectors for transient transduction in nondividing neurons derived from human embryonic stem cells (hESCs). Using a baculoviral vector equipped with the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE), we observed a quick onset of transgene expression as early as day 1 after baculoviral transduction and a high efficiency of up to 80%. Strong transgene expression in the cultured human neurons was observed for more than 1 month and the signal was easily detectable even after 3 months. Using two baculoviral vectors carrying different transgenes, we found that co-transduction at a single neuron level was possible. After transplantation into the brain of nude mice, the baculovirus-transduced human neurons were integrated into the mouse brain and maintained transgene expression for at least 4 weeks, portending the usefulness of this technique in assisting neural transplantation. Therefore, by mediating efficient transient gene expression, baculoviral vectors can provide useful tools for both basic gene function studies in human neurons and therapeutic applications of these cells.