Ex Vivo and In Vivo Localized 1H NMR Spectroscopy of Brain Metabolites in the Study of MeCP2 Mutant Mice
B.Yerby; M.Sheffrin; N.Nag; J.E.Berger-Sweeney; N.H.Kolodny.
Abstract
Rett Syndrome (RTT) is caused by a mutation in the Mecp2 gene on the X-chromosome that alters the production of the transcriptional repressor Mecp2. RTT can be emulated by an Mecp2 mutant mouse model in which physical, behavioral and biochemical abnormalities are amenable to study in the first weeks of life. Nuclear magnetic resonance (NMR) spectroscopy has provided insights into pathogenic mechanisms in animal models of other neurological disorders via determination of regional metabolite levels. This study focuses on the aqueous metabolites N-acetylaspartate (NAA), creatine (Cre) and choline (Cho). We hypothesize that there will be abnormalities in the neurochemistry of Mecp2 mutant mouse brain during early development before physical symptoms become apparent. Mecp2 null males, heterozygous females and C57/BL6 wild-type (WT) controls of both sexes were used for the in vivo and ex vivo studies. In vivo localized NMR spectra were acquired on a 9.4 T NMR spectrometer equipped with a micro-imaging accessory, using the T2-weighted PRESS sequence. For ex vivo NMR studies the metabolites of interest were extracted from fresh frozen brain tissue using methanol-chloroform-water. Ex vivo NMR spectra were acquired at 7 T. We also examined the metabolite profiles of Mecp2 mutant male and WT mice after postnatal dietary Cho supplementation, a potential means of therapeutic intervention for RTT. We have been successful in identifying NAA, Cre, and Cho peaks in our in vivo and ex vivo spectra in mice between 3 and 5 weeks old. Currently, we are quantifying these peaks and analyzing metabolite levels longitudinally.
Lay Summary
Nuclear magnetic resonance (NMR) spectroscopy is one of the principal techniques used to obtain physical, chemical, electronic and structural information about a molecule, in real-time, in a living organism. NMR has provided insights into disease mechanisms in other animal models of neurological disorders. In this abstract, the authors describe their efforts to use NMR to study the neurochemistry of Mecp2 mutant mouse brain during early development before physical symptoms become apparent.