Activation of prospective memory (PM) representations following awakening has been proposed to modulate expression of the cortisol awakening response (CAR). However, experimental testing of this hypothesis is still missing. This study examined the effect of a naturalistic PM-related manipulation on the CAR in a sample of 35 preschool-aged children. The CAR was assessed on two study days (0 and 30min post-awakening) using objective verification of awakening and sampling times. Children had to remember to perform a naturalistic PM-related task (reminding their parents about a gift) on the experimental day while there was no intervention on the control day (counterbalanced order). Results revealed an increased CAR on the experimental day (mean±SD increase: 9.97±7.05nmol/L) compared to the control day (mean±SD increase: 5.45±8.14nmol/L; p=.022). Our findings concur with the notion that expression of the CAR is modulated by post-awakening anticipatory processes involving activation of PM representations.