Zn2+Influx Is Critical for Some Forms of Spreading Depression in Brain Slices

Abstract

Spreading depression (SD) is wave of profound depolarization that propagates throughout brain tissue and can contribute to the spread of injury after stroke or traumatic insults. The contribution of Ca²⁺influx to SD differs depending on the stimulus, and we show here that Zn²⁺can play a critical complementary role in murine hippocampal slices. In initial studies, we used the Na⁺/K⁺ATPase inhibitor ouabain and found conditions in which SD was always prevented by L-type Ca²⁺channel blockers; however, Ca²⁺influx was not responsible for L-type effects. Cytosolic Ca²⁺increases were not detectable in CA1 neurons before SD, and removal of extracellular Ca²⁺did not prevent ouabain-SD. In contrast, cytosolic Zn²⁺increases were observed in CA1 neurons before ouabain-SD, and L-type channel block prevented the intracellular Zn²⁺rises. A slow mitochondrial depolarization observed before ouabain-SD was abolished by L-type channel block, and Zn²⁺accumulation contributed substantially to initial mitochondrial depolarizations. Selective chelation of Zn²⁺withN,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) abolished SD, implying that Zn²⁺entry can play a critical role in the generation of ouabain-SD. TPEN was most effective when synaptic activity was reduced by adenosine A₁receptor activation, and a combination of Ca²⁺and Zn²⁺removal was required to prevent ouabain-SD when A₁receptors were blocked. Similarly, Zn²⁺chelation could prevent SD triggered by oxygen/glucose deprivation but Zn²⁺accumulation did not contribute to SD triggered by localized high K⁺exposures. These results identify Zn²⁺as a new target for the block of spreading depolarizations after brain injury.