Astrocyte Ca²⁺ signaling drives inversion of neurovascular coupling after subarachnoid hemorrhage

Physiologically, neurovascular coupling (NVC) matches focal increases in neuronal activity with local arteriolar dilation. Astrocytes participate in NVC by sensing increased neurotransmission and releasing vasoactive agents (e.g., K⁺) from perivascular endfeet surrounding parenchymal arterioles. Previously, we demonstrated an increase in the amplitude of spontaneous Ca²⁺ events in astrocyte endfeet and inversion of NVC from vasodilation to vasoconstriction in brain slices obtained from subarachnoid hemorrhage (SAH) model rats. However, the role of spontaneous astrocyte Ca²⁺signaling in determining the polarity of the NVC response remains unclear. Here, we used two-photon imaging of Fluo-4-loaded rat brain slices to determine whether altered endfoot Ca²⁺ signaling underlies SAH-induced inversion of NVC.Wereport a time-dependent emergence of endfoot high-amplitude Ca²⁺signals (eHACSs) afterSAHthat were not observed in endfeet from unoperated animals. Furthermore, the percentage of endfeet with eHACSs varied with time and paralleled the development of inversion of NVC. Endfeet with eHACSs were present only around arterioles exhibiting inversion of NVC. Importantly, depletion of intracellular Ca²⁺ stores using cyclopiazonic acid abolished SAH-induced eHACSs and restored arteriolar dilation in SAH brain slices to two mediators of NVC (a rise in endfoot Ca²⁺ and elevation of extracellular K⁺). These data indicate a causal link between SAH-induced eHACSs and inversion of NVC. Ultrastructural examination using transmission electron microscopy indicated that a similar proportion of endfeet exhibiting eHACSs also exhibited asymmetrical enlargement. Our results demonstrate that subarachnoid blood causes a delayed increase in the amplitude of spontaneous intracellular Ca²⁺ release events leading to inversion of NVC.