Nitric oxide (NO) has been implicated in mediating cerebral vasodilation during neuronal activation and specifically to pharmacologic activation of NMDA and kainate receptors. Possible mediators of cerebral vasodilation to AMPA have not been well studied in mature brain, although heme oxygenase activity has been implicated in newborn pigs. In anesthetized rats, 5 minutes of topical superfusion of 30 and 100 µM of AMPA on the cortical surface through a closed cranial window resulted in increases in pial arteriolar diameter. The dilatory response to AMPA was not inhibited by superfusion of a NO synthase inhibitor, a cyclooxygenase-2 inhibitor, or a cytochrome P450 epoxygenase inhibitor, all of which previously have been shown to inhibit the cortical blood flow response to sensory activation. However, the 48 ± 13% dilation to 100 µM AMPA was reduced 56-71% by superfusion of the adenosine A_2A antagonist ZM-241385, the A_2B antagonist alloxazine, and the heme oxygenase inhibitor, chromium mesoporphyrin. Combining the latter three inhibitors did not attenuate the dilator response more than the individual inhibitors, while an AMPA receptor antagonist fully blocked the vasodilation to AMPA. These results indicate that cortical pial arteriolar dilation to AMPA does not require activation of NO synthase, cyclooxygenase-2, or P450 epoxygenase, but does depend on activation of adenosine A_2A and A_2B receptors. In addition, carbon monoxide derived from heme oxygenase appears to play a role in the vascular response to AMPA receptor activation in mature brain by a mechanism that is not additive with that of adenosine receptor activation.