A Scalable Optogenetic System of Focal Stroke Induction in Zebrafish for Testing Stroke Disparities Genes

Stroke is one of the leading causes of death in the U.S. and also a leading cause of adult disability, with severe societal burdens. Ischemic strokes, caused by the thromboembolic occlusion of cerebral arteries, constitute the majority of stroke incidences, and the rest are hemorrhagic strokes with ruptured blood vessels.

In the U.S., stroke incidence shows a clear disparity between African Americans and Caucasians, with a much higher frequency (~240%) in blacks. Despite an intensive search for treatment, recombinant tissue plasminogen activator (rtPA) remains the only FDA-approved post-stroke medicine, with limited effectiveness. Behind this woefully inadequate dearth of stroke therapeutics lies the difficulties in generating a sufficiently large number of stroke-induced animals for effective drug screening, as a highly labor-intensive surgical procedure (middle cerebral artery occlusion) is still the method of choice to induce ischemic strokes in model animals.

Photothrombosis is a rapidly adopted new method to induce ischemic strokes in rodents, with a number of advantages such as highly reproducible infarct size and location with minimal mortality. In this procedure, focal illumination of defined wavelength light on the exposed skull activates an IV-injected photosensitive chemical (e.g., Rose-Bengal) in the bloodstream, causing injuries in endothelial cells and local platelet aggregation, leading to the clogging of the affected blood vessel.

However, even in this case, labor-intensive procedures that cannot be easily scaled up must be utilized to create the stroke model. Recently, photothrombic ischemic stroke has also been successfully induced in the adult zebrafish brain by focal illumination following a manual injection of Rose-Bengal, thus providing an additional vertebrate animal model system for stroke research. In this regard, an exciting novel genetic approach that eliminates the manual injection of photosensitive chemicals was recently created and tested successfully to selectively induce apoptotic cell death by light illumination in the adult zebrafish heart.

Proposed is the creation of a readily scalable, optogenetically induced stroke system using transgenic zebrafish, development of a behavioral test system to identify novel therapeutic chemicals, and testing of GWAS (genome-wide association study)-identified stroke risk variants of the human APOL1 gene, also known as prominent kidney disorder risk factors, in people of African ancestry.