Spontaneous eyeblink rates greatly vary among individuals from several blinks to a few dozen blinks per minute. Because dopamine agonists immediately increase the blink rate, individual differences in blink rate are used as a behavioral index of central dopamine functioning. However, an association of the blink rate with polymorphisms in dopamine-related genes has yet not been found. In this study, we demonstrated that a genetic variation of the nicotinic acetylcholine receptor CHRNA4 (rs1044396) increased the blink rate while watching a video. A receiver operating characteristic analysis revealed that the blink rate predicts a genetic variation in the nicotinic receptor gene with a significant discrimination level (0.66, p < 0.004). The present study suggests that differences in sensitivity to acetylcholine because of the genetic variation of the nicotinic receptor are associated with individual differences in spontaneous eye blink rate.