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. 2010 Dec 23;5(12):e15697. doi: 10.1371/journal.pone.0015697

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This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.

This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration, which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.

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Sucrose density fractionated was performed on brains from three different age groups of C57BL/6J mice: young (Yg, 3–6 months), middle aged (Md, 12 months), and aged (Ag, >18 months). Immunoblot analysis detected the majority of PSD-95 (post-synaptic density marker), NR2A, NR2B, AMPAR, TrkBR, and Cav-1 in buoyant fractions 4 and 5 (BFs) isolated from Yg brains (A). In contrast, the Md and Ag brains exhibited a drastic reduction in these synaptic signaling components, with the majority of these proteins detected in heavy fractions 11 and 12 (HFs) only. Densitometric analysis of the data is represented in B. (C) Cav-1 (C) and PSD-95 (P) immunoprecipitates pulled down NR2A, NR2B, AMPAR, and TrkB in the buoyant fractions of Yg mice, with decreased detection in Md and Ag. (D) Immunoblot analysis detected a significant decrease in PSD-95 (post-synaptic density marker), NR2A, NR2B, AMPAR, and Cav-1 in hippocampal synaptosomes from Md and Ag brains compared to Yg. PSD-95, NR2, NR2B, AMPAR, and Cav-1 decreased in PSD-95 immunoprecipitates of Md, and Ag synaptosomes compared to Yg. (E) Electron paramagnetic resonance (EPR) was performed on synaptosomal membranes from brains of C57BL/6J mice: young (Yg, 3–6 months) and aged (Old, >18 months). Membrane localized spin labels 5-doxylstearic acid (5-DSA) probes changes in the neuronal membrane fluidity closer to the membrane surface. Lineshape analysis of 5-DSA spin label using the indicated parameters revealed that neuronal membrane of aged mice exhibit significantly lower order parameter (i.e. increased fluidity) than young animals. Aged membranes were 8.5±1.2% more fluid than young membranes (F_(1,10) = 223.5, p = 0).