The purpose of this study was to identify the feasibility of using the pupil dilation and heart rate variability (HRV), to diagnose learners' changing cognitive load in a video learning setting. The pupil dilation and HRV of 23 participants were recorded during their learning progress, and the learning materials, including differences in task complexity, were presented to the groups of distinct prior knowledge. The results showed that the high prior knowledge group had lower pupil size than the low prior knowledge group, but the difference was not statistically significant. However, the ratio of low frequency to high frequency power (LF/HF ratio) was higher in the high prior knowledge group than in the low prior knowledge group throughout the entire learning process, and the two groups showed different arousal patterns. Overall, learners' changing cognitive load could be measured and visualised based on psychophysiological responses, the types of cognitive load could be assumed by using the two indices in parallel, and this study raised practical issues to the use of pupil size as an indicator of cognitive load in video learning.