1 Changes in membrane potential and the intracellular calcium concentration 2 during CSD and OGD in layer V and layer II / III mouse cortical neurons

17 Cortical spreading depression (CSD) is an episode of electrical silence following intense 18 neuronal activity that propagates across the cortex at about 3-6 mm/min and is 19 associated with transient neuronal depolarization. CSD is benign in normally perfused 20 brain tissue but there is evidence suggesting that repetitive CSD contributes to infarct 21 growth following focal ischemia. Studies to date have assumed that the cellular 22 responses to CSD are uniform across neuronal types because there is no data to the 23 contrary. In this study we have investigated the effect of CSD on membrane potential 24 and the intracellular calcium concentration ([Ca]i) of mouse layer V and layer II/III 25 pyramidal neurons in brain slices. To place the data in context we have made similar 26 measurements during anoxic depolarization induced by oxygen and glucose deprivation 27 (OGD). The [Ca]i was quantified using the low-affinity ratiometric indicator Fura-4F. 28 During both CSDand OGD-induced depolarization the membrane potential 29 approached 0 mV in all neurons. In layer V pyramids OGD resulted in an increase in 30 [Ca]i to a maximum of 3.69 ± 0.73 μM (mean ± S.D; n = 12), significantly greater than 31 the increase to 1.81 ± 0.70 μM in CSD (n = 34; p < 0.0001). Membrane potential and 32 [Ca]i returned to nearly basal levels following CSD, but not OGD. Layer II/III neurons 33 responded to CSD with a greater peak increase in [Ca]i than layer V neurons (2.88 ± 34 0.6 μM; n = 9; p< 0.01). We conclude there is a laminar difference in the response of 35 pyramidal neurons to CSD; possible explanations are discussed. 36