Targeted single blood vessel occlusion in rodent pial arteries to study astrocyticvascular dysfunction in a mini-stroke model

Abstract

The ability to form targeted vascular occlusions in small vessels of the brain is an important technique to study the microscopic basis of cerebral ischemia. We utilize a method that enables the targeted occlusion of any single blood vessel within the upper 350 μm of mouse neocortex to generate highly localized regions of ischemia by blocking capillary or surface arteries. This method makes use of linear optical absorption by a photosensitizer, transiently circulated in the bloodstream, to induce a clot in a surface or near-surface segment of a vessel after activation with a green 532nm continuous laser. Using two-photon microscopy of green fluorescent protein-labeled astrocytes (GFEC) and 70-kDa Texas Red dextranlabeled blood flow, we explore the alteration of the spatial relationship between cortical microcirculation and astrocytic endfeet structure in vivo during ischemia. This two-photon imaging method allows extremely high spatial and temporal resolution for studying pathological mechanisms that underlie ischemic injury, including abnormal changes in cell signaling and structure, vascular dysfunction, and inflammation. We aim to identify the cellular basis of cerebrovascular dysregulation in transgenic animal models of neurological disease and identify molecular targets for the pharmacotherapy of these pathological pathways.