Background There is an association between sports participation and substance use. However, there is some evidence that intramural sports in high school may not have the same effect. Therefore, the objective of this research was to examine the longitudinal associations between intramural participation in high school and substance use. Methods: This study used a three-year linked sample (2016-2018) of grade 9 and 10 (ages 13-17) Canadian high school students in the COMPASS (Cannabis use, Obesity, Mental health, Physical activity, Alcohol use, Smoking, Sedentary behavior) study (n = 7,845). Students reported their participation in intramurals over time (consistent, none, initiate, intermittent, and quit) and their substance use behaviors (binge drinking, cannabis use, cigarette use, and e-cigarette use). Mixed effects models were used. Results: 42% of students did not participate in intramurals. For binge drinking, male students who never participated had lower odds (0.66 [0.47-0.93]) compared to consistent intramural participators. Female (3.50 [CI: 1.34-9.16]) and male students (1.97 [1.28-3.02]) who did not participate in any intramurals were more likely to use cannabis than consistent participators. Male students who did not participate were also more likely to use cigarettes (1.81 [1.05-3.12]). No associations were found between intramural participation and e-cigarette use. Conclusion: Intramural participation may be associated with increased binge drinking among male high school students. More promisingly, consistent participation in intramurals may be protective against cannabis use among male and female students and cigarette use among male students.