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ISOLATION AND PARTIAL CHARACTERIZATION OF RAT BRAIN SYNAPTIC PLASMA MEMBRANES 1
Author(s) -
Gurd J. W.,
Jones L. R.,
Mahler H. R.,
Moore W. J.
Publication year - 1974
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1974.tb11591.x
Subject(s) - synaptosome , membrane , microsome , chromatography , chemistry , biochemistry , gel electrophoresis , electrophoresis , ficoll , cell fractionation , mitochondrion , sodium , biophysics , enzyme , biology , in vitro , peripheral blood mononuclear cell , organic chemistry
— Synaptic plasma membranes from the cortices of adult rat brain were isolated from synaptosomes prepared by flotation of a washed mitochondrial pellet (P2) in a discontinuous Ficoll‐sucrose gradient. Contamination of the synaptosome fraction by microsomes was estimated by enzymic and chemical analysis to be less than 15 per cent. (2) The purified synaptosome fraction was subjected to osmotic shock, subfractionated on a discontinuous sucrose gradient and the distribution of enzymic and chemical markers for synaptic plasma membranes, microsomal membranes and mitochondria was determined. (3) Comparison of synaptosome subfractions prepared in the presence and absence of 1 mM NaH 2 PO 4 /0.1 mM EDTA buffer pH 7.5, indicated that the ionic composition of the isolation medium markedly affected the distribution and enzymic composition of the subfractions. (4) Synaptic plasma membranes prepared in the presence of PO 4 /EDTA exhibited a 10‐fold enrichment in [Na + + K + ] ATPase and were characterized by less than 15 and 10 per cent contamination by microsomes and mitochondria respectively. (5) The polypeptide composition of the purified synaptic plasma membranes was compared with the microsomes and mitochondria by polyacrylamide gel electrophoresis in sodium dodecyl sulphate. No differences between the protein and glycoprotein composition of the synaptic plasma membranes and microsomes were detected. The mitochondria, in contrast, possessed a unique protein composition.