Perivascular Neutrophil Extracellular Traps Exacerbate Microvasospasm After Experimental Subarachnoid Hemorrhage

BACKGROUND: Subarachnoid hemorrhage (SAH) can lead to acute or delayed cerebral ischemia. Recent findings have revealed that spasm of microvessels, called microvasospasm, may contribute to SAH-related cerebral ischemia, and perivascular inflammation is considered important in the development of microvasospasms. However, owing to the difficulty in investigating the dynamics of vascular and perivascular events, little is known about the mechanisms underlying microvasospasms. METHODS: We established an experimental system aiming to investigate the vascular and perivascular pathology of SAH by combining a SAH mouse model with intravital 2-photon imaging. SAH was induced by intracisternal blood injection, and the distribution of erythrocytes, neutrophil behavior, and morphological changes in the pial arterioles were analyzed over time by 2-photon microscopy imaging. To further explore the role of neutrophils and neutrophil extracellular traps (NETs) in microvasospasm, we performed neutrophil depletion by intraperitoneal administration of neutrophil-specific antibody or NETs removal by intracisternal administration of DNase. RESULTS: Erythrocytes were immediately distributed in the perivascular space of the arterioles after SAH induction; neutrophils intensively infiltrated the perivascular space within 2 days and subsequently showed NETosis; and pial arterioles in the same region developed pearl-string-like microvasospasms in the subacute phase. Neutrophil depletion significantly reduced the number of microvasospasms. Furthermore, the removal of perivascular NETs drastically reduced microvasospasms. CONCLUSIONS: By establishing a unique experimental system, we demonstrated that perivascular NETs could be a new therapeutic target for microvasospasms.