The Role of Neurovascular Interactions in the Development and Regulation of the Blood-Brain Barrier

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Project Summary The blood-brain barrier (BBB) maintains homeostasis of the central nervous system (CNS) chemical environment, and is critical for proper neural function; BBB breakdown has been implicated in neurodegenerative diseases, including Alzheimer’s disease and Parkinson’s disease. The BBB is also a huge obstacle for drug delivery to the brain. For both of these reasons, we need a better understanding of the molecular regulators of BBB development and function. This K99/R00 proposal will support the PI, Dr. Natasha O’Brown, in developing innovative strategies and acquiring new skill sets to study the signaling between neurovascular cells and blood vessels involved in establishing BBB function. The experiments will be initiated during the mentored period (carried out in Dr. Sean Megason’s lab at Harvard Medical School) and continue in the PI’s own laboratory upon obtaining an independent position. Dr. O’Brown’s long-term career goal is to unravel the cellular and molecular interactions that direct BBB development and maintenance, with the hope of identifying new targets for therapeutic drug delivery into the brain. Dr. O’Brown’s previous work identified a novel zebrafish mutant with vastly increased BBB permeability. The leaky phenotype mapped to a missense mutation in the neuronally-produced secreted proteoglycan Spock1. Spock1 is the first identified neuronal factor that establishes BBB properties, and it therefore represents a huge opportunity to learn more about how endothelial cell permeability is regulated by the CNS microenvironment. Aim 1 will use zebrafish genetics, cell transplantation and tracer leakage assays to perturb and assess Spock1’s function in regulating BBB properties. Aim 2 will use a combination of lineage tracing, live imaging and RNA sequencing to characterize how loss of Spock1 alters CNS cellular interactions leading to increased BBB leakage. Finally, Aim 3 will establish a connection between BBB dysfunction and altered behavior, generating a new platform for a small molecule screen to identify novel molecules that alter BBB function. The discovery of novel molecular regulators of BBB function will form the foundation of Dr. O’Brown’s independent laboratory. Dr. O’Brown’s extensive training in developmental biology and genetics makes her an ideal candidate to further develop this important research area. Building on this training, she will develop expertise in zebrafish genetic and cellular manipulations, 4D high-resolution live imaging and analysis, and in performing and analyzing behavioral drug screens. She will be mentored throughout the K99 period by Drs. Sean Megason, Leonard Zon, Chenghua Gu and Randall Peterson, world-class experts in zebrafish development and quantitative imaging, zebrafish genetics and genomics, BBB physiology and small molecule screens. Together, the proposed experiments and training will position Dr. O’Brown to be an excellent candidate for independent faculty positions, and for a leadership role in research into the cellular and molecular control of BBB development and function.