The structure of postsynaptic densities isolated from dog cerebral cortex: I. overall morphology and protein composition

Abstract

A postsynaptic density (PSD) fraction, including some adherent subsynaptic web material, has been isolated from dog cerebral cortex by a short-procedure modification of the methods of Davis and Bloom (21,22) and Cotman and Taylor (20), using Triton X-100. The fraction has been visualized by thin-section, replica, and negative (phosphotungstic acid) staining electron microscopy and its proteins separated by high-resolution SDS gel electrophoresis. Morphologically, the preparation seems to be quite pure, with very little membrane contamination. The density is composed of protein, no nucleic acids, and very little phospholipid being detectable. The fraction had no ATPase or GTPase activity, but it did have a very small amount of cytochrome c oxidase activity (of a specific activity less than 0.5 % that of a mitochondrial fraction) and a small amount of 5'-nucleotidase activity (of a specific activity between 6 and 7% that of a synaptic membrane fraction). Electron micrographs reveal cup-shaped structures -400 nm long and -40 nm wide, made up of apparent particles 13-28 nm in diameter. However, en face views, and particularly micrographs of replicas and PTA-stained preparations, reveal a disk-shaped structure, outside diameter -400 nm, in which filaments are seen to extend from the central part of the density. High resolution gel electrophoresis studies indicated some 15 major proteins and perhaps 10 more minor ones; the predominant protein had a mol wt of 51,000, followed by ones at 45,000, 40,000, 31,000, 26,000, and several at 100,000. A comparison by gel electrophoresis of density fraction proteins with those of a lysed synaptosomal membrane fraction containing some adherent densities indicated some comigrating proteins, but the major membrane fraction protein, mol wt 52,000, was not found in the density fraction. Antibodies raised against the density fraction reacted with a preparation of solubilized synaptic membrane proteins. By both these criteria, it was considered that the density and the synaptic membrane have some proteins in common. By separately mixing 125I-labeled myelin, synaptic vesicle, and mitochondrial fraction protel'~S with synaptosomes, and then isolating the density

Brevican isoforms associate with neural membranes

“…The presence of brevican in this fraction has already been reported (Seidenbecher et al 1998;Niederöst et al 1999) and it was suggested to contribute to the nonpermissive properties of CNS myelin (Niederöst et al 1999). The light membrane fraction is a less well defined membrane subpopulation, which contains a mixture of endoplasmic reticulum, Golgi and plasma membranes as estimated by electron microscopy (Cohen et al 1977;Hu et al 1998). This fraction contains extrasynaptic marker molecules of neuronal membranes, such as BDNF receptor gp145trkB and gp95trkB or the 5-hydroxytryptamine receptor 2A (Hu et al (b) (a) Fig.…”

Structural role of zinc ions bound to postsynaptic densities

“…Disc-shaped, segmented and perforated PSDs, consisting of granular and filamentous materials, have been observed by electron microscopy (Cohen and Siekevitz 1978;Spack 1985;Harris and Stevens 1989;Peters et al 1991;Geinisman et al 1992;Itarat and Jones 1993). Biochemical procedures have been developed to isolate PSDs from the brains of different animals (Cotman et al 1974;Cohen et al 1977;Gurd et al 1982). The isolated PSDs appeared to retain the morphological characteristics of their brain counterparts.…”

Tumour-associated antigens reacting with cytotoxic antibodies in serum of hepatoma-bearing rats

“…Column chromatography was performed in the cold on a Sepharose CL-4B column (1.6 x 100 cm) (Pharmacia, Uppsala, Sweden Neville (1971) as modified by Cohen et al (1977). The gels were stained with Coomassie Brilliant Blue R (Sigma) and destained as described by Chua & Bennoun (1975).…”

“…The gels were stained with Coomassie Brilliant Blue R (Sigma) and destained as described by Chua & Bennoun (1975). Human IgG and IgM, o-actinin, dog-brain neurofilament protein and post-synaptic density proteins (Cohen et al, 1977) were used as mol.-wt standards.…”