|Targeting Matrix Metalloproteinases to Treat Dendritic Spine Malformation and Behavioral Defects in Fragile X Mice
by Iryna Ethell, 5/1/2009
This project will determine if the beneficial effects of minocycline result from its ability to regulate MMP-9 expression and activity, with an emphasis on the role MMPs play in Fragile-X pathophysiology. There are three directions these studies will follow:
1. We will study the specific effects of inhibiting or depleting MMP-9 have on dendritic spine development and behavior in Fmr1 KO mice. This will be done with specific MMP-9 inhibitors and through genetic crosses with Fmr1 KO mice with mmp9 KO mice. We anticipate that reduced MMP-9 expression or activity in Fmr1 KO neurons will lead to similar behavioral and morphological effects as are seen with minocycline treatment. We will investigate the effects of MMP-9 depletion on dendritic spine/synapse development in Fmr1 KO hippocampal neurons by crossing mmp-9 KO mice (FVB.Cg-MMP9tm1Tvu/J; stock number 004104) with Fmr1 KO mice. Further, we will screen some new tetracycline derivatives for their ability to promote dendritic spine maturation in Fmr1 KO hippocampal neurons.
2. We will examine whether higher levels of MMP expression/activity can be detected in Fragile X patients. Since enhanced MMP-9 expression and activity have been found in the brains of Fmr1 KO mice, and may underlie the morphological and behavioral abnormalities in these mice, it is important to determine if MMP-2 and MMP-9 activity is higher in FXS patients. FXS may also affect MMP regulation outside the central nervous system (CNS) as patients display several characteristics of modified extracellular matrix (ECM) including large ears, long faces, hyperextensible joints and flat feet. We will use gelatinase assays, gelatin gel zymography and ELISAs to analyze the levels of MMP-9 expression and activity in blood and tissue samples from FXS patients and controls. We will also analyze the levels and activities of other MMPs as well as the activities of endogenous MMP inhibitors, the TIMPs, using specific ELISA and activity assays.
3. We will test the effects of minocycline and other tetracycline derivatives on the behavioral performance of adult Fmr1 KO mice. Our preliminary results have demonstrated that minocycline treatment of young Fmr1 KO mice reduces their anxiety level in the elevated plus maze test, and increases their visual-spatial recognition in Y-maze to behavior levels characteristic of the WT mice (Bilousova et al., 2009). Here we will compare the effects of minocycline and other tetracycline derivatives that are more potent MMP-9 inhibitors in adult Fmr1 KO mice. Behavioral testing will be performed in both WT and Fmr1 KO mice. The behavioral tests employed are designed to analyze levels of anxiety (elevated plus maze and open field; Yan et al., 2005), obsessive-compulsive behavior (marble burying), as well as learning and memory (RAWM and platform recognition, Ethell et al., 2006, Cao et al., 2009).