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PMID: 7159406 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

An efficient method for the isolation and separation of basolateral-membrane and luminal-membrane vesicles from rabbit kidney cortex.

The Biochemical journal ·Vol. 208 ·No. 2 ·1982-11-15 ·Pages 377-82

Sheikh MI, Kragh-Hansen U, Jørgensen KE, Røigaard-Petersen H

Abstract

A procedure for isolation and separation of purified luminal-membrane and basolateral-membrane vesicles from adult and newborn rabbit renal cortex by using Ca2+/Mg2+ precipitation, differential centrifugation and a self-orienting Percoll-gradient centrifugation is described. The purity of the membrane-vesicle suspensions was examined by electron microscopy and by measuring the activity of several marker enzymes. The activity of Na+ + K+-stimulated ATPase in the fraction mainly containing adult rabbit basolateral-membrane vesicles was enriched 16-fold, and the activity of alkaline phosphatase in the fraction mainly containing luminal-membrane vesicles was increased 13-fold, compared with the homogenate. Similar results were obtained with kidneys from newborn rabbits. Uptake studies, with a rapid filtration technique and the spectrophotometric method described in an accompanying paper [Kragh-Hansen, Jørgensen & Sheikh (1982) Biochem. J. 208, 359-368], showed that both adult and newborn rabbit luminal-membrane vesicles, in contrast with the basolateral-membrane preparations, possess an Na+-dependent electrogenic transport system for L-proline. Adult rabbit luminal-membrane vesicles take up citrate and L-malate by Na+-dependent electrogenic processes, whereas adult rabbit basolateral membrane vesicles do not exhibit electrogenic uptake of citrate. By contrast, these vesicles show Na+-dependent electrogenic uptake of L-malate.

MeSH Terms
Animals Cell Fractionation/methods Cell Membrane/enzymology,metabolism,ultrastructure Citrates/metabolism Citric Acid In Vitro Techniques Kidney Cortex/enzymology,metabolism,ultrastructure Malates/metabolism Microscopy, Electron Proline/metabolism Rabbits
Chemicals
Citrates Malates Citric Acid malic acid Proline
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Sheikh M I
Kragh-Hansen U
Jørgensen K E
Røigaard-Petersen H
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22 references, click to expand
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Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2080377
Published
1982-11-15
Pages
377-82
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1153973
Subset
IM
External Links
PubMed source DOI: 10.1042/bj2080377 PMC Full Text View journal details
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