Project Summary ! The neurovasculature supplies nutrients to the 100 billion neurons in the adult human brain via a 600 km network of capillaries and microvessels. As the interface between the brain and the vascular system, the blood-brain barrier, which includes the neurovasculature, is responsible for regulating the brain microenvironment by preventing fluctuations in chemistry, transport of immune cells, and the entry of toxins and pathogens. At the same time, almost all diseases of the brain are associated with disruption or dysfunction of the neurovasculature, which leads to entry of blood components, immune cells, and pathogens into the brain, and ultimately causes neuroinflammation, oxidative stress, and neurotoxicity. Functional human models have the potential to address many unresolved questions associated with the role of the neurovasculature in health and disease, and in developing more complex models of the human nervous system that will ultimately contribute towards the realization of integrated multicellular systems. There are two major challenges to developing physiologically-relevant, tissue-engineered models of the human neurovasculature: (1) a source of relevant cells, and (2) 3D cell culture methods to build the model. Stem cell technology provides a solution to providing a reliable source of human, brain-specific cells, a long-standing barrier to developing blood-brain barrier models. Similarly, advances in tissue engineering provide the tools for self-organization of perfusable vascular networks. Solving these problems will have significant impact on neuroscience research, elucidating mechanisms of central nervous system diseases, and in the development and translation of new therapies and technologies. In Aim 1 we will characterize the phenotype and barrier function of brain microvessels under quiescent conditions and in response to activation/stress. In Aim 2 we will develop and characterize brain-specific capillary networks. In Aim 3 we will integrate pericytes and astrocytes into our models. These models will enable a broad range of applications, including fundamental studies of neurogenesis, vascularization, and development, and mechanistic studies of disease progression, treatment, repair, drug and gene delivery, and toxicity.