Macromolecular Architecture Of The Synapse

Neurological Disorders And Stroke

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Abstract

The post synaptic density (PSD) at excitatory glutamatergic synapses is a complex molecular machine which appears to be a key site of information storage. New methods to probe its structure show that a lattice-like backbone labeling for PSD-95 forms its core, while other structural components-synGAP, SHANK, and the kinase CaMKII-occupy various locations in the lattice. The PSDs in intact neurons, however, change size rapidly during activity and these changes are rapidly reversible. Immunolabeling shows that CaMKII is a major component of the added mass. Measurements of the mass of PSDs from ischemic brains shows an increase in mass sufficient to allow for the addition of up to 400 CaMKII holoenzymes. Furthermore, CaMKII at the PSD is in the phosphorylated form, and reversibility is inhibited by specific phosphatases, suggesting that addition of CaMKII depends on phosphorylation. A method has been developed to affinity purify PSDs from other components of the PSD fraction, which will allow independent measurement of CaMKII content. After high pressure freezing of cultures at rest and after NMDA stimulation they can be freeze-substituted and examined by post embedding immunogold and tomography of thin sections. Imunnogold labeling shows the distribution PSD molecules while tomography resolves PSD structure down to molecular dimensions.