NMDA receptors sustain but do not initiate neuronal depolarization in spreading depolarization.

Spreading depolarization (SD) represents a neurological process characterized by a massive, self-sustaining wave of brain cell depolarization. Understanding its mechanism is important for treating ischemic or hemorrhagic stroke and migraine with aura. Many believed that ion fluxes through NMDA receptors (NMDARs) are responsible for neuronal transmembrane currents of SD. However, the explicit role of NMDARs remains ambiguous. This is in part due to the limitation of traditional pharmacological approaches in resolving the contribution of NMDARs in different intercellular and intracellular processes of SD. Here, we applied single-cell blockade and genetic deletion methods to remove functional NMDARs from individual hippocampal CA1 neurons in order to examine the role of NMDARs in the depolarization mechanism without affecting the propagation of SD. We analyzed neuronal membrane potential changes to demonstrate that NMDARs are not required for initiating the depolarization. Consistently, neuronal input resistance (RN) revealed a sharp decline at the start of SD, which was unaffected by blocking NMDARs. Instead, the recovery of both membrane potential and RN during the late phase of SD was facilitated by inhibition of NMDARs, indicating that NMDARs are responsible for sustaining the depolarization. Our results strongly indicate that NMDAR activation is not a determinant of the initiation of depolarization but is important for sustaining transmembrane ion fluxes during SD.

Spreading Depression Promotes Astrocytic Calcium Oscillations and Enhances Gliotransmission to Hippocampal Neurons.

Spreading depression (SD) is a pathophysiological phenomenon characterized by propagating waves of profound neuronal and glial depolarization in central nervous system gray matter. Although SD is primarily mediated by neurons with a subsequent astrocytic response, it remains unclear how astrocytic activity is modulated after SD and how altered astrocyte signaling contribute to neuronal excitability. Here, we report that after the concurrent Ca2+ wave, SD enhanced astrocytic activity by promoting a secondary period of Ca2+ oscillations. SD-induced Ca2+ oscillations did not require the activation of metabotropic glutamate receptors or purinergic receptors; instead, they were mediated by the activation of GABAB receptors and 1,4,5-trisphosphate (IP3) receptors. Furthermore, SD increased the number of NMDA receptor-mediated slow inward currents (SICs) in CA1 pyramidal neurons. The frequency of SD-induced SICs was reduced by blockade of GABAB receptors or by limiting Ca2+ efflux from the ER. Selective inhibition of astrocytic Ca2+ signals by dialysis of BAPTA into astrocytes or by knocking out the astrocytic type of IP3 receptors suppressed SICs after SD. These results demonstrated a causative link between the SD-induced Ca2+ oscillations and the enhanced glutamatergic astrocyte-neuron signaling. Therefore, we conclude that SD enhances the astrocyte Ca2+ signals and further promotes gliotransmission and neuronal excitability.

Effects of anti-epileptic drugs on spreading depolarization-induced epileptiform activity in mouse hippocampal slices.

Epilepsy and spreading depolarization (SD) are both episodic brain disorders and often exist together in the same individual. In CA1 pyramidal neurons of mouse hippocampal slices, induction of SD evoked epileptiform activities, including the ictal-like bursts, which occurred during the repolarizing phase of SD, and the subsequent generation of paroxysmal depolarization shifts (PDSs), which are characterized by mild depolarization plateau with overriding spikes. The duration of the ictal-like activity was correlated with both the recovery time and the depolarization potential of SD, whereas the parameters of PDSs were not significantly correlated with the parameters of SD. Moreover, we systematically evaluated the effects of multiple anti-epileptic drugs (AEDs) on SD-induced epileptiform activity. Among the drugs that are known to inhibit voltage-gated sodium channels, carbamazepine, phenytoin, valproate, lamotrigine, and zonisamide reduced the frequency of PDSs and the overriding firing bursts in 20-25 min after the induction of SD. The GABA uptake inhibitor tiagabine exhibited moderate effects and partially limited the incidence of PDSs after SD. AEDs including gabapentin, levetiracetam, ethosuximide, felbamate, and vigabatrin, had no significant effect on SD-induced epileptic activity. Taken together, these results demonstrate the effects of AEDs on SD and the related epileptiform activity at the cellular level.