ObjectiveWe aimed to analyze whether or not fear conditioning exerts an effect on prepulse inhibition (PPI) of blink reflex (BR). To create fear conditioning, we used fearful faces. Since fearful faces lead to a specific set of fear conditioning, we hypothesized PPI of BR would change under the observation of fearful faces.MethodWe included 17 healthy subjects with a mean age of 30.86.9 years and seven healthy subjects with a mean age of 57.77.3 years between January 2018 and June 2018 and recorded PPI of BR. The recordings were done before observation of any image, during observation of images, and immediately after observation of images. Observation of images included observation of fearful faces for 30 s and a neutral image of a white screen for 30 s (in a randomized order).Results There was a R2-PPI deficit during observation of fearful faces in each group whereas R2-PPI fully developed at other time points. R1 amplitude and R2 magnitude were lower during observation of any image compared with baseline and post-observation time points.Conclusion In conclusion, a deficit of R2-PPI develops during observation of fearful faces in humans which is probably related to activation of the amygdala.