The aim of this study was to investigate central pain representations during loading of the periodontium induced by orthodontic and occlusal stress. Nineteen healthy male volunteers (25.7 ± 2.8 years) were tested on two consecutive days: after phenotyping (questionnaires) and determination of warmth (WPT) and heat (HPT) pain thresholds, functional magnetic resonance imaging was performed as event-related paradigm including 36 tooth clenchings of 3 s duration, alternating with rest periods varying between 20–30 s. The task was performed in absence (T1) and 24 h after placement of an elastic separator between the second bicuspid and the first molar on the right side of the lower jaw (T2). No significant changes in WPT and HPT were observed but pain ratings were significantly elevated at T2. Significantly elevated activation at T2, as compared to T1, was found in bilateral sensorimotor cortex, bilateral secondary sensory cortex, supplementary motor area, right rolandic operculum, and bilateral insula. Our data show for the first time in humans that periodontal stimulation, as tested by tooth clenching in the presence of an elastic separator, goes along with specific expressions of pain at behavioral and neuronal network levels. Findings supplement the existing neuroimaging literature on odontogenic pain.