The overall hypothesis of this proposal is that organophosphate compounds, such as chlorpyrifos, are metabolically activated by the blood brain barrier to a more potent compound (oxon) at a rate that is greater than the rate of inactivation. This active compound alters blood brain barrier structure, function and development by altering signaling pathways that are important in the maintenance of blood brain barrier integrity. We propose to test this hypothesis with the use of an in vitro model of the blood brain barrier, which consists of co-cultures of endothelial cells with astrocytes. We rationalize the use of an in vitro model because measurements of blood brain barrier function in live animals are difficult and costly to perform. The following specific aims will be used to test the above hypothesis: 1) structural and enzymatic characterization of an in vitro BBB model, which consists of primary bovine microvascular endothelial cells (BMEC) and neonatal rat astrocytes; 2) delineation of the effects of chlorpyrifos (CPF) and the activated CPF-oxon on structural and enzymatic characteristics of the blood brain barrier, including effects on BBB development, integrity of an established BBB, esterase activity and metabolite production; 3) determination of CPF and CPF-oxon effects on downstream signaling cascades that have been shown to be important in controlling BBB permeability and endothelial cell proliferation. ? ?
