CYTOTOXIC EDEMA INDUCES SIMULTANEOUS DEPOLARIZATION IN ANOXIC RAT BRAIN SLICES
Spreading depolarizations (SD) lead to cytotoxic edema and enhance lesion progression in cerebrovascular diseases. In our previous, in vivo experiments we observed a novel form of depolarization which affects the cortex simultaneously and more extensively (simultaneous depolarization, SiD). We have postulated and set out to prove that the underlying process of SiD must be neuronal hyperexcitability caused by astrocytic swelling. Live coronal brain slices (350 µm) prepared from Wistar rats (n=10) were perfused with artificial cerebrospinal fluid (aCSF). Hypo-osmotic medium (HM, for 20 min) was washed on the slices to induce edema, and O2 was withdrawn (2.5 min) to create anoxia. White light reflectance videos were captured with a CCD camera, local field potential (LFP) was recorded via glass capillary electrodes. Electron microscopy and Golgi-Cox staining was used to determine cell swelling. Bumetanide (1 mM) and TGN-020 (100 µM) were administered to block Na-K-Cl cotransporters and aquaporin-4 channels. HM caused permanent astrocyte swelling and triggered SDs (propagation velocity: 3.06 ± 0.61 mm/min, amplitude: 16.62±5.96 mV). Subsequent SiD elicited by anoxia in HM engaged the cortical area which had been involved in SD previously (HM: SiD; n=10, aCSF: SD; n=8). SiD enlarged the depolarized area (SiD vs. SD: 56.2±5.6 vs. 33.7±7.5 %). Pharmacological treatment reduced the duration of SDs and inhibited SiDs (bumetanide+TGN-020 vs. HM control 77.37±36.41 vs. 131.63±41.14 s). We conclude that astrocyte buffering capacity has pivotal role in SD propagation and lesion expansion. Moreover, pharmacological blockade of cytotoxic edema is essential to decrease secondary injury in cerebrovascular disease states.