A method for objectively determining supratentorial cerebrospinal fluid (CSF) production rate is described. The method employs cine phase-contrast magnetic resonance imaging with high temporal and spatial resolution at the level of the aqueduct of Sylvius. Automatic encircling of the aqueduct was accomplished by using a pulsatility-based segmentation approach. Results in 23 healthy adults (18 men, five women; age range, 21-39 years) yielded an average CSF production rate of 305 microL/min +/- 145 (standard deviation); this rate is in good agreement with literature-reported values obtained with invasive ventriculolumbar perfusion measurements. Average operator imprecision was 23.1% if automatic segmentation was not used. The proposed method is potentially an effective means for measuring supratentorial CSF production rate in humans.