OBJECTIVE The purpose of this study was to develop and test a clinically relevant method to mechanically stimulate lumbar functional spinal units while recording brain activity by means of functional magnetic resonance imaging (MRI). METHODS Subjects were investigated in the prone position with their face lying on a modified stabilization pillow. To minimize head motion, the pillow was fixed to the MRI headrest, and supporting straps were attached around the shoulders. An experienced manual therapist applied controlled, nonpainful pressure stimuli to 10 healthy subjects at 3 different lumbar vertebrae (L1, L3, and L5). Pressure applied to the thumb was used as a control. The stimulation consisted of posterior to anterior (PA) pressure movement. The therapist followed a randomized stimulation protocol projected onto a screen in the MRI room. Blood oxygenation level-dependent responses were analyzed in relation to the lumbar and the thumb stimulations. The study was conducted by the Chiropractic Department, Faculty of Medicine, University of Zürich, Switzerland. RESULTS No participant reported any discomfort due to the prone-lying position or use of the pillow. Importantly, PA-induced pressure produced only minimal head movements. Stimulation of the lumbar spinous processes revealed bilateral neural responses in medial parts of the postcentral gyrus (S1). Additional activity was observed in the secondary somatosensory cortex (S2), posterior parts of the insular cortex, different parts of the cingulate cortex, and the cerebellum. Thumb stimulations revealed activation only in lateral parts of the contralateral S1. CONCLUSION The current study demonstrates the feasibility of the application of PA pressure on lumbar spinous processes in an MRI environment. This approach may serve as a promising tool for further investigations regarding neuroplastic changes in chronic low back pain subjects.