title: Fine structure of synapses on dendritic spines.
abstract: Camillo Golgi's "Reazione Nera" led to the discovery of dendritic spines, small appendages originating from dendritic shafts. With the advent of electron microscopy (EM) they were identified as sites of synaptic contact. Later it was found that changes in synaptic strength were associated with changes in the shape of dendritic spines. While live-cell imaging was advantageous in monitoring the time course of such changes in spine structure, EM is still the best method for the simultaneous visualization of all cellular components, including actual synaptic contacts, at high resolution. Immunogold labeling for EM reveals the precise localization of molecules in relation to synaptic structures. Previous EM studies of spines and synapses were performed in tissue subjected to aldehyde fixation and dehydration in ethanol, which is associated with protein denaturation and tissue shrinkage. It has remained an issue to what extent fine structural details are preserved when subjecting the tissue to these procedures. In the present review, we report recent studies on the fine structure of spines and synapses using high-pressure freezing (HPF), which avoids protein denaturation by aldehydes and results in an excellent preservation of ultrastructural detail. In these studies, HPF was used to monitor subtle fine-structural changes in spine shape associated with chemically induced long-term potentiation (cLTP) at identified hippocampal mossy fiber synapses. Changes in spine shape result from reorganization of the actin cytoskeleton. We report that cLTP was associated with decreased immunogold labeling for phosphorylated cofilin (p-cofilin), an actin-depolymerizing protein. Phosphorylation of cofilin renders it unable to depolymerize F-actin, which stabilizes the actin cytoskeleton. Decreased levels of p-cofilin, in turn, suggest increased actin turnover, possibly underlying the changes in spine shape associated with cLTP. The findings reviewed here establish HPF as an appropriate method for studying the fine structure and molecular composition of synapses on dendritic spines.
authors.affiliation: Institute for Structural Neurobiology, Center for Molecular Neurobiology Hamburg, University Medical Center Hamburg-Eppendorf Hamburg Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Frotscher
authors.affiliation: Institute of Anatomy, University of Bern Bern Switzerland.
authors.forename: Daniel
authors.initials: D
authors.lastname: Studer
authors.affiliation: Institute of Anatomy, University of Bern Bern Switzerland.
authors.forename: Werner
authors.initials: W
authors.lastname: Graber
authors.affiliation: Institute for Structural Neurobiology, Center for Molecular Neurobiology Hamburg, University Medical Center Hamburg-Eppendorf Hamburg Germany.
authors.forename: Xuejun
authors.initials: X
authors.lastname: Chai
authors.affiliation: Institute of Anatomy and Cell Biology, University of Freiburg Freiburg Germany.
authors.forename: Sigrun
authors.initials: S
authors.lastname: Nestel
authors.affiliation: Institute for Structural Neurobiology, Center for Molecular Neurobiology Hamburg, University Medical Center Hamburg-Eppendorf Hamburg Germany.
authors.forename: Shanting
authors.initials: S
authors.lastname: Zhao
keywords: actin cytoskeleton; cofilin; dendritic spine; electron microscopy; high-pressure freezing; immunogold labeling