Approximately 200,000 veterans receive cancer treatment at Veterans Health Administration facilities. With improving survival rates for many cancers, the deleterious long-term cardiovascular side effects of cancer therapies have become increasingly apparent. For many of the cancers that frequently occur among veterans, radiation therapy (RT) is an integral component of treatment. Despite improvements in the techniques used to target RT to the cancer tissue, some radiation always reaches surrounding normal tissue. Endothelial damage within the small blood vessels or “radiation endotheliopathy” has been postulated as a major cause of RT- induced injury of normal tissue. Many post-radiation syndromes have been attributed to radiation endotheliopathy. One of them is cognitive decline, which is estimated to affect as many as 90% of patients after brain RT. Mechanistically, radiation endotheliopathy is believed to be initiated by mitochondrial injury, leading to chronic oxidative stress and endothelial dysfunction over a period of years. Thus, intervention at the time of RT is likely critical for reducing radiation endotheliopathy and its sequelae, including cognitive decline. The discovery of effective mitigators will require a unified view of the pathways by which RT-related mitochondrial injury influences various steps in this progression; this has not been achieved to date. The objective of the proposed project is to identify the mechanisms by which mitochondrial injury promotes radiation endotheliopathy, and to test whether protection from mitochondrial injury prevents adverse short- and long-term effects of RT in small blood vessels. RT induces mitochondrial DNA damage, perturbs ATP production, enhances the mitochondrial membrane potential, Ca2+ uptake and reactive oxygen species (ROS) production. The mitochondrial Ca2+ uniporter (MCU) regulates Ca2+ uptake into the mitochondrial matrix and was recently reported as being redox-dependent. Mitochondrial Ca2+ augments ROS production that promotes further mitochondrial dysfunction. Thus, we posit that MCU in endothelial cells (ECs) drives a feed-forward circuit with mitochondrial ROS that leads to long-term adverse effects of RT, including cognitive decline after brain RT. Indeed, published data from our laboratory demonstrate that blocking mitochondrial Ca2+ uptake is sufficient to reduce mitochondrial ROS production and protect EC barrier function. Thus, our central hypothesis is that MCU-mediated Ca2+ uptake by mitochondria is required for excessive ROS production after RT, and thus for chronic mitochondrial DNA damage, blood brain barrier (BBB) breakdown, capillary loss, and cognitive decline. This hypothesis is further supported by our strong pilot data that inhibition of MCU in ECs during RT abolishes mitochondrial DNA damage in vitro and protects against BBB breakdown in vivo. Our novel tools and assays put us in the perfect position to perform the proposed study. These include genetic models...