The purpose of this study was to investigate the prevalence and configuration of Middle Mesial Canals (MMC) and isthmi in mandibular molars in Pakistani population using Cone Beam Computed Tomography (CBCT). METHODOLOGY: Sixty CBCTs of patients were selected randomly from database of University College of Dentistry, Lahore. First and second permanent mandibular molars without evidence of previous endodontic treatment, full coverage restoration or root resorption were evaluated for the number of roots, canals and the presence and configuration of MMC and isthmi. The age and gender of patients were also recorded. SPSS was used for statistical analysis using chi-square test. Significance level was set at 5% (P < 0.05). RESULTS: Out of 189 mandibular molars, MMCs were found in 9 and isthmi in 62 teeth. No statistical association was found between the presence of MMC and age, gender, side or tooth type. Significantly more isthmi were found in second molars, females and younger patients, P < 0.05. CONCLUSION: MMC and isthmi in mesial roots of mandibular molars are not unusual findings in our population and CBCT can be a valuable tool for their detection.