Cortical spreading depression (CSD) is a pathophysiological phenomenon. There are sufficient evidences to prove that CSD plays an important role in some neurological disorders. However, exact mechanisms of its initiation and propagation are still unclear. Previous studies showed that glutamate receptors could be concerned with CSD, but those studies were mostly performed oriented to ionotropic glutamate receptors (iGluRs). There is relatively little report about effects of metabotropic glutamate receptors (mGluRs) on CSD. Here, we applied optical intrinsic signal imaging (OISI) combined with direct current (DC) potential recording to examine influences of some mGluRs antagonist (or agonist) on CSD propagation in rat’s brain, to indirectly validate actions of some mGluRs on CSD. We found that N-acetyl-l-aspartyl-l-glutamate (NAAG, an agonist at mGluR3) inhibited the propagation of CSD, and the inhibition was gradually developed with time. However, 6-methyl-2-phenylethynyl-pyridine (MPEP, an antagonist of mGluR5) did not produce any significant alterations with the CSD propagation. Our findings suggest that mGluR3 could play an important role in the CSD propagation, but the activity of mGluR5 was comparatively weak. These findings can help to understand the propagation mechanism of CSD, and consider the therapy of some neurological diseases involved with CSD.