ISSN:
1471-4159
Source:
Blackwell Publishing Journal Backfiles 1879-2005
Topics:
Medicine
Notes:
Abstract: We report a method for the isolation of enriched fractions of intact Golgi apparatus from neurons of 10- to 12-day-old rat brains. Neurons were prepared according to a modified method of Farooq and Norton [J. Neurochem.31, 887–894 (1978)]. Golgi-enriched fractions were obtained after centrifugation of postmitochondrial supernatants in a discontinuous sucrose gradient. Golgi fractions 1 and 2, recovered at the interfaces of 28–34% and 34–36% sucrose densities, respectively, were examined with morphometric and enzymatic methods. Morphometric analyses showed that 21–34% of fraction 1 and 11–29% of fraction 2 consisted of intact Golgi apparatus. Lysosomes, mitochondria, ribosomes, and rough endoplasmic reticulum contaminated fraction 1 (6–10%) and fraction 2 (14–26%). Golgi fraction 1 showed a 25- to 65-fold enrichment over neurons of UDP Gal:GlcNAc galactosyltransferase, CMP-sialic acid:lactosylceramide sialyltransferase, and PAPS:cerebroside sulfotransferase activities. Golgi fraction 2 showed a 8-to 23-fold enrichment over neurons of the activities of the above glycolipid- and glycoprotein-synthesizing enzymes. The activities of the possible marker enzymes rotenone-insensitive NADH-cytochrome c reductase, succinate-cytochrome c reductase, and arylsulfatase were low or minimally elevated in the Golgi fractions. A sevenfold enrichment of Na+,K+-ATPase activities was found in the Golgi fractions. This is consistent either with significant plasma membrane contamination or with the presence of this enzyme in the neuronal Golgi apparatus.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1111/j.1471-4159.1985.tb04016.x
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