The present study aimed to investigate if (1) subcutaneous injection of glutamate induces pain, sensitization and vasomotor responses in humans and (2) if sex differences exist in these responses. Thirty healthy volunteers (men-15 and women-15) were included. Each subject received four subcutaneous injections (0.1ml; glutamate 100, 10, 1mM and isotonic saline 0.9%) into the forehead skin in two sessions separated by one week. Assessments of pain intensity (VAS), quality, distribution; area of pinprick hyperalgesia; pressure pain threshold (PPT) at the injection site; surface skin temperature and local blood flow were performed at predetermined time points. The highest concentration of glutamate evoked the highest pain intensity, the longest duration of pain and the largest pain area under the VAS-time curve (P<0.001) in both men and women, although responses in women were larger than in men (P<0.05). The face-chart pain area was the largest for the highest concentration of glutamate (P<0.001) and women drew a larger pain area than men (P=0.024). The area of pinprick hyperalgesia was the largest for glutamate 100mM (P<0.001) and women indicated a larger area than men (P<0.001). Concentration-dependent local vasomotor responses were found following the subcutaneous injection of glutamate but there was no sex difference in this effect. Glutamate 100mM significantly reduced the PPT values (P<0.001) without sex-related differences. The present study demonstrates for the first time that subcutaneous injection of glutamate evokes pain, vasomotor responses and pinprick hyperalgesia in human volunteers and that there are sex-related differences in some of these responses.