Cortical spreading depression (CSD) is the neurophysiological correlate of migraine aura and a trigger of migraine pain mechanisms. The mechanisms of initiation of CSD in migraineurs are unknown. Insights into these mechanisms can be obtained by studying the mechanisms of facilitation of CSD initiation in genetic mouse models of the disease. Here, we investigated these mechanisms in knock-in mice carrying a mutation in cacna1a causing familial hemiplegic migraine (FHM1 mice). High KCl puffs of increasing duration up to the threshold duration eliciting a CSD were applied on layer 2/3 of cerebral cortex slices whilst the membrane potential of a pyramidal neuron located near the site of CSD initiation and the intrinsic optic signal were simultaneously recorded before and after application of MK-801 to block NMDARs. We show that a threshold level of NMDAR activation is required to initiate CSD in FHM1 mice, similar to that required in WT mice. The delay of several seconds with which this occurs underlies the delay of CSD initiation relative to the rapid neuronal depolarization produced by KCl. In FHM1 mice, the threshold level of NMDAR activation is reached with a lower stimulation and in a shorter time than in WT mice and this explains the facilitation of CSD initiation in the mutants. While in WT mice block of NMDARs prevents CSD initiation by even largely suprathreshold stimuli, in FHM1 mice suprathreshold stimuli can initiate CSD, suggesting that, besides activation of NMDARs, other CaV2.1-dependent processes may contribute to CSD initiation by intense stimulation in FHM1.