Researchers hypothesized that entertainment via threedimensional virtual reality (3D-VR) provides greater enjoyment than the same media experienced in a twodimensional (2D) environment. This result can improve the quality of life of patients, particularly those who are hospitalized for long periods. To test this hypothesis, the instantaneous heart rates (IHRs) of eight subjects were monitored by electrocardiography using a Holter ECG as a measure of the autonomic nervous system activity levels when viewing a two-minute roller coaster video in 2D and 3D-VR (using an Oculus Rift). Each subject viewed the video in both 2D and 3D-VR in varying order with at least a 10 min interval between them; after viewing the video, they responded to a questionnaire that included questions about the subject's preference for 2D versus 3D-VR. In addition, subjective estimation of enjoyment was conducted according to the visual analog scale (VAS). Results show that the 3D-VR environment, which is closer to a real environment, elicited more excitement and enjoyment, as evidenced by the difference in the sympathetic nervous system activity (P < 0.01) when viewing the 2D and 3D-VT videos. The subjects particularly enjoyed the 3D-VR experience when it was observed before the 2D video, as demonstrated by larger IHR accelerations. However, the difference depended on whether the subjects liked roller coasters or not. In addition, two subjects experienced motion sickness, which is associated with visual fatigue due to parallax. One subject reported simultaneously feeling enjoyment and motion sickness; therefore, categorizing the cause of this subject's sympathetic nervous system activity was difficult. Additionally, for some of the subjects, the questionnaire responses (including the VAS estimation) were inconsistent with the sympathetic nervous system activity. Therefore, additional bio-signals (e.g., brain activity or salivary amylase activity) should be analyzed in parallel, to further evaluate the subject's physical response. In future studies, a video of a lighter activity that is more broadly liked by people, such as hiking or a riding a Ferris wheel with a moderate altitude, should be used.