BACKGROUND AND AIMS: Nicotine, a widely used toxic substance, has various receptors scattered throughout the body that have shown opposite effects on inflammatory disorders. However, the effects of nicotine on heart rate variability in ulcerative colitis are unclear. Therefore, the present study aimed to determine the effect of acute nicotine injection on heart rate variability in a rat model of ulcerative colitis. METHODS: Six male Wistar rat groups, containing vehicle, UC (induction of ulcerative colitis without treatment), and nicotine (0.5, 1, 1.5, and 2 mg/kg), were assessed. First, the rats were anesthetized and the initial electrocardiogram was recorded. Twenty-four hours after the induction of ulcerative colitis with 4 % acetic acid by rectal injection, a second electrocardiogram was recorded. Finally, 15 min after nicotine injection in each group, the last electrocardiogram was recorded. Linear and nonlinear indices of heart rate variability were extracted from the recorded R-R intervals. RESULTS: A single injection of nicotine at high doses increased the standard deviation of R-R intervals, root mean square of successive differences between normal heartbeats, ratio of the short-term deviation to the long-term deviation of R-R intervals, and entropy of R-R intervals in ulcerative colitis animals (at least P < 0.05). CONCLUSIONS: Acute injection of nicotine at doses 1.5 and 2 mg/kg can improve R-R interval linear indices, balance the ratio of short-term deviation to long-term deviation, and modify the entropy in the induced ulcerative colitis rats. However, further research is needed for the clinical use of acute nicotine injection in ulcerative colitis.