Use of digitonin extraction to distinguish between autophagic-lysosomal sequestration and mitochondrial uptake of [14C]sucrose in hepatocytes
In isolated rat hepatocytes, electroinjected [14C]sucrose is sequestered both by mitochondria and by autophagosomes/lysosomes. Radioactivity can be selectively extracted from the latter organelles by low concentrations of digitonin, thereby providing a specific bioassay for autophagic sequestration....
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| Veröffentlicht in: | Biochemical journal 1985-12, Vol.232 (3), p.773-780 |
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| container_title | Biochemical journal |
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| creator | Gordon, P B Tolleshaug, H Seglen, P O |
| description | In isolated rat hepatocytes, electroinjected [14C]sucrose is sequestered both by mitochondria and by autophagosomes/lysosomes. Radioactivity can be selectively extracted from the latter organelles by low concentrations of digitonin, thereby providing a specific bioassay for autophagic sequestration. By including a digitonin extraction step in the assay procedure, autophagic [14C]sucrose sequestration could be shown to be virtually completely (greater than 90%) suppressed by the autophagy inhibitor 3-methyladenine (10 mM), whereas mitochondrial sugar uptake was unaffected. An amino acid mixture likewise suppressed autophagic sequestration very strongly, while having no detectable effect on the mitochondria. |
| doi_str_mv | 10.1042/bj2320773 |
| format | Article |
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Radioactivity can be selectively extracted from the latter organelles by low concentrations of digitonin, thereby providing a specific bioassay for autophagic sequestration. By including a digitonin extraction step in the assay procedure, autophagic [14C]sucrose sequestration could be shown to be virtually completely (greater than 90%) suppressed by the autophagy inhibitor 3-methyladenine (10 mM), whereas mitochondrial sugar uptake was unaffected. An amino acid mixture likewise suppressed autophagic sequestration very strongly, while having no detectable effect on the mitochondria.</description><identifier>ISSN: 0264-6021</identifier><identifier>EISSN: 1470-8728</identifier><identifier>DOI: 10.1042/bj2320773</identifier><identifier>PMID: 4091820</identifier><language>eng</language><publisher>England</publisher><subject>Adenine - analogs & derivatives ; Adenine - pharmacology ; Amino Acids - pharmacology ; Animals ; Autophagy - drug effects ; Digitonin - pharmacology ; In Vitro Techniques ; Liver - cytology ; Liver - drug effects ; Liver - metabolism ; Lysosomes - drug effects ; Lysosomes - metabolism ; Male ; Mitochondria, Liver - metabolism ; Phagocytosis - drug effects ; Rats ; Rats, Inbred Strains ; Sucrose - metabolism</subject><ispartof>Biochemical journal, 1985-12, Vol.232 (3), p.773-780</ispartof><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c370t-5f5bdc096258c5cdcef5f231b54ae373e22c04dcfa3a1e01a37bd452eb16cd923</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC1152950/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC1152950/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,727,780,784,885,27924,27925,53791,53793</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/4091820$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Gordon, P B</creatorcontrib><creatorcontrib>Tolleshaug, H</creatorcontrib><creatorcontrib>Seglen, P O</creatorcontrib><title>Use of digitonin extraction to distinguish between autophagic-lysosomal sequestration and mitochondrial uptake of [14C]sucrose in hepatocytes</title><title>Biochemical journal</title><addtitle>Biochem J</addtitle><description>In isolated rat hepatocytes, electroinjected [14C]sucrose is sequestered both by mitochondria and by autophagosomes/lysosomes. Radioactivity can be selectively extracted from the latter organelles by low concentrations of digitonin, thereby providing a specific bioassay for autophagic sequestration. By including a digitonin extraction step in the assay procedure, autophagic [14C]sucrose sequestration could be shown to be virtually completely (greater than 90%) suppressed by the autophagy inhibitor 3-methyladenine (10 mM), whereas mitochondrial sugar uptake was unaffected. An amino acid mixture likewise suppressed autophagic sequestration very strongly, while having no detectable effect on the mitochondria.</description><subject>Adenine - analogs & derivatives</subject><subject>Adenine - pharmacology</subject><subject>Amino Acids - pharmacology</subject><subject>Animals</subject><subject>Autophagy - drug effects</subject><subject>Digitonin - pharmacology</subject><subject>In Vitro Techniques</subject><subject>Liver - cytology</subject><subject>Liver - drug effects</subject><subject>Liver - metabolism</subject><subject>Lysosomes - drug effects</subject><subject>Lysosomes - metabolism</subject><subject>Male</subject><subject>Mitochondria, Liver - metabolism</subject><subject>Phagocytosis - drug effects</subject><subject>Rats</subject><subject>Rats, Inbred Strains</subject><subject>Sucrose - metabolism</subject><issn>0264-6021</issn><issn>1470-8728</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1985</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpVkc1q3DAUhUVJmE7TLvoABa8CXTi5-rNnNoUwpEkhkE2yCkXI8vVYqS25ltx0HiLvHM1kGJKVQPc759zLIeQrhTMKgp1Xj4wzKEv-gcypKCFflGxxRObACpEXwOhH8imERwAqQMCMzAQs6YLBnDzfB8x8k9V2baN31mX4P47aROtdFn36D9G69WRDm1UYnxBdpqfoh1avrcm7TfDB97rLAv6dMCTpTqldnfXJ0LTe1aNN82mI-s8u6oGK1e8wmdGn6BTY4qATuYkYPpPjRncBv-zfE3L_8_JudZ3f3F79Wl3c5IaXEHPZyKo2sCyYXBhpaoONbBinlRQaecmRMQOiNo3mmiJQzcuqFpJhRQtTLxk_IT9efYep6jHpXVq8U8Noez1ulNdWvZ8426q1_6colWwpIRmc7g1Gv7tb9TYY7Drt0E9BlYVMGOUJ_P4Kbs8NIzaHEApq2506dJfYb2-3OpD7svgLn-uaRA</recordid><startdate>19851215</startdate><enddate>19851215</enddate><creator>Gordon, P B</creator><creator>Tolleshaug, H</creator><creator>Seglen, P O</creator><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>19851215</creationdate><title>Use of digitonin extraction to distinguish between autophagic-lysosomal sequestration and mitochondrial uptake of [14C]sucrose in hepatocytes</title><author>Gordon, P B ; Tolleshaug, H ; Seglen, P O</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c370t-5f5bdc096258c5cdcef5f231b54ae373e22c04dcfa3a1e01a37bd452eb16cd923</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1985</creationdate><topic>Adenine - analogs & derivatives</topic><topic>Adenine - pharmacology</topic><topic>Amino Acids - pharmacology</topic><topic>Animals</topic><topic>Autophagy - drug effects</topic><topic>Digitonin - pharmacology</topic><topic>In Vitro Techniques</topic><topic>Liver - cytology</topic><topic>Liver - drug effects</topic><topic>Liver - metabolism</topic><topic>Lysosomes - drug effects</topic><topic>Lysosomes - metabolism</topic><topic>Male</topic><topic>Mitochondria, Liver - metabolism</topic><topic>Phagocytosis - drug effects</topic><topic>Rats</topic><topic>Rats, Inbred Strains</topic><topic>Sucrose - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Gordon, P B</creatorcontrib><creatorcontrib>Tolleshaug, H</creatorcontrib><creatorcontrib>Seglen, P O</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Biochemical journal</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Gordon, P B</au><au>Tolleshaug, H</au><au>Seglen, P O</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Use of digitonin extraction to distinguish between autophagic-lysosomal sequestration and mitochondrial uptake of [14C]sucrose in hepatocytes</atitle><jtitle>Biochemical journal</jtitle><addtitle>Biochem J</addtitle><date>1985-12-15</date><risdate>1985</risdate><volume>232</volume><issue>3</issue><spage>773</spage><epage>780</epage><pages>773-780</pages><issn>0264-6021</issn><eissn>1470-8728</eissn><abstract>In isolated rat hepatocytes, electroinjected [14C]sucrose is sequestered both by mitochondria and by autophagosomes/lysosomes. Radioactivity can be selectively extracted from the latter organelles by low concentrations of digitonin, thereby providing a specific bioassay for autophagic sequestration. By including a digitonin extraction step in the assay procedure, autophagic [14C]sucrose sequestration could be shown to be virtually completely (greater than 90%) suppressed by the autophagy inhibitor 3-methyladenine (10 mM), whereas mitochondrial sugar uptake was unaffected. An amino acid mixture likewise suppressed autophagic sequestration very strongly, while having no detectable effect on the mitochondria.</abstract><cop>England</cop><pmid>4091820</pmid><doi>10.1042/bj2320773</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | Biochemical journal, 1985-12, Vol.232 (3), p.773-780 |
| issn | 0264-6021 1470-8728 |
| language | eng |
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| source | MEDLINE; EZB-FREE-00999 freely available EZB journals; PubMed Central; Alma/SFX Local Collection |
| subjects | Adenine - analogs & derivatives Adenine - pharmacology Amino Acids - pharmacology Animals Autophagy - drug effects Digitonin - pharmacology In Vitro Techniques Liver - cytology Liver - drug effects Liver - metabolism Lysosomes - drug effects Lysosomes - metabolism Male Mitochondria, Liver - metabolism Phagocytosis - drug effects Rats Rats, Inbred Strains Sucrose - metabolism |
| title | Use of digitonin extraction to distinguish between autophagic-lysosomal sequestration and mitochondrial uptake of [14C]sucrose in hepatocytes |
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