Malignant astrocyte swelling and impaired glutamate clearance drive the expansion of injurious spreading depolarization foci

Abstract

Spreading depolarizations (SD) indicate infarct maturation and predict worse clinical outcome in ischemic stroke. We demonstrate here in rodents that brain edema formation upon ischemic stroke impairs astroglial glutamate clearance and increases the tissue area invaded by SD. The cytotoxic glutamate accumulation predisposes an extensive bulk of tissue for a yet undescribed simultaneous depolarization (SiD). We confirm in rat brain slices under hypo-osmotic stress that SiD is the pathological expansion of prior SD foci, is associated with astrocyte swelling and triggers oncotic neuron death. The blockade of astrocytic aquaporin-4 channels and Na+/K+/Cl- co-transporters, or volume-regulated anion channels mitigated slice edema, glutamate accumulation and SiD occurrence. Reversal of slice edema by hyperosmotic treatment counteracted glutamate accumulation and prevented SiD. In contrast, paralysis of astrocyte metabolism or inhibition of astrocyte glutamate uptake reproduced the SiD phenotype. We discuss our results in the light of evidence for SiD in the human cortex. Our results emphasize the need of preventive osmotherapy in ischemic stroke.

Comparative analysis of spreading depolarizations in brain slices exposed to osmotic or metabolic stress

“…The partial electrophysiological regeneration was supported by the propensity of the tissue to bear subsequent SDs either spontaneously occurring in HM (Fig. 1) or triggered with transient anoxia in HM [31]. In line with the functional data, the first SD in HM caused cellular injury in the cortex engaged in SD, but the damage was less severe than that seen after SD in OGD (Fig.…”

“…ischemia is the restricted availability of oxygen, as well as glucose) [40]. To study the impact of osmotic stress on SD, we incubated the brain slices in HM, following a previously established approach to model cerebral edema [26,31]. We experimented with HM solutions of decreasing Na + content in our pilot experiments (120-100-80-60-40 mM, HM 120 -HM 40 , respectively), and selected here HM 60 for systematic work, because incubation of the slices in HM 60 reliably produced SD within 10 min after HM application.…”

“…We accepted higher numbers of TTC-positive cellular compartments (i.e. particles) to indicate better tissue viability [31]. The SD-related TTC-positive particle loss was obvious after the single SD in HM or OGD (26.45 ± 10.64 and 11.44 ± 6.42 vs. 54.33 ± 21.21 per 1000 µm 2 , HM and OGD vs. KCl).…”

“…Also, the application of pharmacological agents may yield highly reproducible, consistent data due to the uniform conditions, to aid the mechanistic understanding of SD evolution. For example, specific ion channels, exchangers or transporters mediating ion currents or amino acid translocations with SD may be studied effectively [27][28][29][30][31]. Finally, brain slice preparations are an ideal, basic experimental model to test the efficacy of SD inhibiting agents (e.g.…”

“…The histological consequences of SD under these states are also evaluated. We pay particular attention here to spatial SD features, such as the volume of the area engaged in SD, which then corresponds to injury progression in the brain slice [31]. Rather than examining SD at the cellular level in a restricted tissue volume on the µm-scale [34,35], we provide large scale (mm) whole slice measurements.…”