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FRACTIONATION OF OLFACTORY TISSUE HOMOGENATES. ISOLATION OF A CONCENTRATED PLASMA MEMBRANE FRACTION
Author(s) -
Koch R. B.,
Norring N. L.
Publication year - 1969
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.1471-4159.1969.tb05933.x
Subject(s) - cytochrome c oxidase , biochemistry , differential centrifugation , atpase , chemistry , fractionation , membrane , microsome , cell fractionation , density gradient , centrifugation , nervous tissue , free nerve ending , enzyme , chromatography , biology , endocrinology , physics , quantum mechanics , neuroscience
— Differential and sucrose‐density‐gradient centrifugation techniques were used for studies on the separation of subcellular particles from rabbit brain and olfactory tissue. Comparisons were made among various fractions from the two types of tissue. These comparisons included protein concentration and enzyme activities of the individual fractions as well as their distribution in subfractions from density gradient separations. In tissue whole homogenates, the percentage of total ATPase activity as ouabain sensitive Na + ‐K + ATPase activity was about 4 times greater in brain cortex (63 per cent) than in olfactory tissue (17 per cent). Cytochrome oxidase and Na + ‐K + ATPase activities were used to indicate the presence and the concentration of mitochondria and of the plasma membranes. A fraction with properties similar to the mitochondria plus nerve ending fraction from brain homogenates (fraction B) was obtained from olfactory tissue. Nerve ending concentration subfractions (B 2 ) were prepared from the B primary fractions. Plasma membrane subfractions were obtained by osmotic shock treatment of B 2 , In the fraction of plasma membrane from olfactory tissue (E 2 ), 56 per cent of the total ATPase activity was Na + ‐K + ATPase activity. In E 2 from brain 71 per cent was Na + ‐K + ATPase activity. Deoxycholate (DOC)‐treated fractions containing nerve endings from brain preparations showed much greater increase in cytochrome oxidase activity than did similar fractions from olfactory tissue. DOC treatment increased the NADH cytochrome c reductase activity of all fractions and subfractions from brain, while it decreased activity in all but one fraction from olfactory tissue. DOC treatment decreased both the Mg 2+ and Na + ‐K + ATPase activities in both types of tissue. Electron photomicrographs of olfactory B 2 , B 3 , E 2 and E 3 show clear morphological differences among these subfractions. The presence of possible cilia and basal bodies on vesicles in B 2 gives morphological evidence for the presence of terminal swellings in this subtraction in agreement with enzyme marker activity results.