The pyrrolidinone derivative nefiracetam facilitates hippocampal synaptic transmission in a PKC-dependent manner. The present study was conducted to understand the mechanism underlying the action of nefiracetam on the PKC activity. In the cell-free system, nefiracetam enhanced the activity of PKCε, activated by linoleic acid, in an ATP concentration (2-500 μM)-dependent manner, and a huge enhancement was found at the concentrations higher than 100 μM. Notably, a similar effect was also obtained with uridine. ATP dissolved in water was spontaneously degraded into ADP, AMP, and adenosine in an incubation time (1-15 days)-dependent manner. The ATP degradation was accelerated by adding nefiracetam or uridine. Taken together, the results of the present study show that nefiracetam as well as uridine enhance the activated PKCε activity possibly by assisting ATP hydrolysis and lowering the kinase activation energy.