In this case–control study, we observed that (a) GFAP serum levels are significantly increased in migraine patients, particularly in those with medication overuse; (b) NfL levels are not increased in migraine patients; (c) attack at sampling/days from last attack and migraine frequency/attack severity do not influence NfL and GFAP levels; and (d) NfL but not GFAP levels are higher in patients with a long migraine course. This evidence concerns the gene GFAP and migraine disorder.