This study was designed to test the usefulness of nitrogen-13 (N-13) glutamate imaging with positron emission tomography in defining myocardial ischemia in humans. Seventeen patients who had undergone coronary arteriography were studied with N-13 glutamate at peak supine exercise using a bicycle ergometer, as well as with the flow tracer N-13 ammonia at peak exercise during a second similar exercise test. Six of the patients also underwent imaging with N-13 glutamate at rest before exercise testing; in the remaining 11 patients imaging with fluorine-18 (F-18) fluorodeoxyglucose was performed to assess glucose metabolism after the second exercise test. Seven patients had classic metabolism-flow mismatches consistent with ischemia (that is, decreased N-13 ammonia uptake in a region with relatively increased F-18 fluorodeoxyglucose uptake). There was no evidence of increased N-13 glutamate uptake in the ischemic mismatched regions in any of these patients. In all 17 patients, the uptake of N-13 glutamate during exercise paralleled the uptake of N-13 ammonia during exercise, suggesting that N-13 glutamate behaves as a flow tracer rather than as a metabolic marker of ischemia in humans.