Cellular Mechanisms Regulating Blood-Brain Barrier Permeability

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The brain microvessel endothelial cells that form the blood-brain barrier (BBB) have an important role in maintaining the proper extracellular environment within the brain required for central nervous system function. Morphologically, the brain microvessel endothelial cells form tight junctions that restrict the passage of solutes between the endothelial cells. In addition, the brain microvessel endothelial cells express a variety of transporters that move solutes and nutrients into and out of the brain. While often considered a static barrier, it is now understood that BBB permeability can be influenced by both neuronal and humoral signals that can alter tight junction and transporter function. However, the cellular mechanisms by which such alterations in BBB permeability can be produced remain to be fully characterized. Much emphasis has been placed on the tight junction complex that forms between adjacent brain microvessel endothelial cells and regulation of BBB permeability. However, it is increasingly recognized that there are plasma membrane proteins, in close proximity to the tight junction complex, that may also influence both paracellular and transcellular passage of solutes, macromolecules and even immune cells. The present Discovery grant application examines the role of two of these plasma membrane proteins in regulation of BBB permeability at the cellular and molecular level. The studies will proceed down two separate and independent research tracks. The first involves understanding the contribution of the adherens junction in regulating both paracellular diffusion and intracellular routes of BBB penetration involving solute carriers and vesicular transport. In particular, the role of the junction adhesion molecule, cadherin (CAD), will be systematically examined from the cellular to whole animal level. The hypothesis is that CAD not only regulates paracellular diffusion across brain endothelial cells but also influences intracellular transport through interactions with scaffold proteins involved in solute and macromolecules transport. The second research theme evaluates the role of connexins (Cxs) in solute and macromolecule permeability. For both CAD and Csx, approaches will first involve examination of these processes in various cell culture models of the BBB using a combination of specifically targeted peptide based inhibitors of these proteins along with siRNA knock-down and gene transfection methods. In vivo studies correlating BBB permeability changes with alterations in CAD and Csx function will also be performed providing research opportunities to trainees (2 grad and 1 undergrad) incorporating functional as well as cellular and molecular biology. Studies will provide key knowledge of cellular processes involved in regulating BBB permeability as well as information on cellular junction formation important in a variety of barrier cells.