Role of NSAID/RHO-GTPase mediated signalling in glia inflammation in primary and in organotypic cultures

Several epidemiological studies have established that treatment with non steroidal anti-inflammatory drugs (NSAIDs) reduces the risk for Alzheimer's disease (AD), but the mechanism of action remains controversial. The general hypothesis of this project is that the RhoGTPase family that controls cytoskeleton dynamics is a molecular target of NSAIDs in brain, and that NSAIDs provide protection in AD via Rho-GTPases by reducing chronic inflammation and improving synaptic plasticity. The project presents as novel ideas that alterations of the actin cytoskeleton paly a role in neurodegenerative diseases, and that NSAIDs can regulate synapticplasticity, a role independent of their anti-inflammatory effects. The study will be the coordinate effort of three groups to accomplish a comprehensive analysis of the role Rho-GTPases in NSAID-mediated protection. Subproject 1 will aim at characterizing the interplay between NSAIDs and Rho-GTPase (Rho, Rac, Cdc42) in the inflammatory reactivity of astrocytes and microglia in cultures and in organotypic cultures, looking at several parameters of inflammation. Rho-GTPase activities will be modulated either pharmacologically or molecularly, by infection with adenovirus containing "double negative" or "constitutively active" forms of Rho-GTPases. In Subproyect 2 we will test whether the NSAID ibuprofen modulates LTP (a functional expression of synaptic plasticity) in hippocampal slices. The molecular mechanism involved will be explored, in particular the role of cGMP-dependent signalling that is akey regulator of LTP. Subproject 3 will complete the analysis of synaptic plasticity by looking at neuronal remodelling in culture. Finally, in this suproject we will charcterize Rho-GTPase expression in Ad brains and in mice over-expressing the amyloid precursor protein (APP)