Similar expression of c-fos in the TNC was found in the optogenetic CSD group, NaCl group and EYFP group, however the pain-related regions were significantly activated in CSD group, indicating that activation of glutamatergic neurons to induce CSD may cause headache by directly activating cortical pain-related regions rather than activating TNC by meningeal afferents. This evidence concerns the gene FOS and Headache.