Desensitization of .gamma.-aminobutyric acid receptor from rat brain: two distinguishable receptors on the same membrane
Transmembrane chloride flux mediated by gamma-aminobutyric acid (GABA) receptor can be measured with a mammalian brain homogenate preparation containing sealed membrane vesicles. The preparation can be mixed rapidly with solutions of defined composition. Influx of 36Cl- tracer initiated by mixing wi...
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| Veröffentlicht in: | Biochemistry (Easton) 1987-12, Vol.26 (24), p.7556-7562 |
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| creator | Cash, Derek J Subbarao, Katragadda |
| description | Transmembrane chloride flux mediated by gamma-aminobutyric acid (GABA) receptor can be measured with a mammalian brain homogenate preparation containing sealed membrane vesicles. The preparation can be mixed rapidly with solutions of defined composition. Influx of 36Cl- tracer initiated by mixing with GABA was rapidly terminated by mixing with bicuculline methiodide. The decrease in the isotope influx measurement due to prior incubation of the vesicle preparation with GABA, which increased with preincubation time and GABA concentration, was attributed to desensitization of the GABA receptor. By varying the time of preincubation with GABA between 10 ms and 50 s with quench-flow technique, the desensitization rates could be measured over their whole time course independently of the chloride ion flux rate. Most of the receptor activity decreased in a fast phase of desensitization complete in 200 ms (t 1/2 = 32 ms) at saturation with GABA. Remaining activity was desensitized in a few seconds (t 1/2 = 533 ms). These two phases of desensitization were each kinetically first order and were shown to correspond with two distinguishable GABA receptors on the same membrane. The receptor activities could be estimated, and the faster desensitizing receptor was the predominant one, giving on average ca. 80% of the total activity. The half-response concentrations were similar, 150 and 114 microM for the major and minor receptors, respectively. The dependence on GABA concentration indicated that desensitization is mediated by two GABA binding sites. The fast desensitization rate was approximately 20-fold faster than previously reported rates while the slower desensitization rate was slightly faster than previously reported rates. |
| doi_str_mv | 10.1021/bi00398a004 |
| format | Article |
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The preparation can be mixed rapidly with solutions of defined composition. Influx of 36Cl- tracer initiated by mixing with GABA was rapidly terminated by mixing with bicuculline methiodide. The decrease in the isotope influx measurement due to prior incubation of the vesicle preparation with GABA, which increased with preincubation time and GABA concentration, was attributed to desensitization of the GABA receptor. By varying the time of preincubation with GABA between 10 ms and 50 s with quench-flow technique, the desensitization rates could be measured over their whole time course independently of the chloride ion flux rate. Most of the receptor activity decreased in a fast phase of desensitization complete in 200 ms (t 1/2 = 32 ms) at saturation with GABA. Remaining activity was desensitized in a few seconds (t 1/2 = 533 ms). These two phases of desensitization were each kinetically first order and were shown to correspond with two distinguishable GABA receptors on the same membrane. The receptor activities could be estimated, and the faster desensitizing receptor was the predominant one, giving on average ca. 80% of the total activity. The half-response concentrations were similar, 150 and 114 microM for the major and minor receptors, respectively. The dependence on GABA concentration indicated that desensitization is mediated by two GABA binding sites. The fast desensitization rate was approximately 20-fold faster than previously reported rates while the slower desensitization rate was slightly faster than previously reported rates.</description><identifier>ISSN: 0006-2960</identifier><identifier>EISSN: 1520-4995</identifier><identifier>DOI: 10.1021/bi00398a004</identifier><identifier>PMID: 2447945</identifier><language>eng</language><publisher>Washington, DC: American Chemical Society</publisher><subject>550201 - Biochemistry- Tracer Techniques ; ALKALINE EARTH METAL COMPOUNDS ; AMINO ACIDS ; AMINOBUTYRIC ACID ; ANIMALS ; AUTONOMIC NERVOUS SYSTEM AGENTS ; BASIC BIOLOGICAL SCIENCES ; BETA DECAY RADIOISOTOPES ; BETA-MINUS DECAY RADIOISOTOPES ; Bicuculline - pharmacology ; BIOCHEMICAL REACTION KINETICS ; Biological and medical sciences ; BODY ; BRAIN ; CALCIUM CHLORIDES ; CALCIUM COMPOUNDS ; CALCIUM HALIDES ; CARBOXYLIC ACIDS ; CELL CONSTITUENTS ; Cell Membrane - metabolism ; CELL MEMBRANES ; Cell receptors ; Cell structures and functions ; CENTRAL NERVOUS SYSTEM ; Cerebral Cortex - metabolism ; CHLORIDES ; Chlorides - metabolism ; CHLORINE 36 ; CHLORINE COMPOUNDS ; CHLORINE ISOTOPES ; DRUGS ; ELECTRON CAPTURE RADIOISOTOPES ; Fundamental and applied biological sciences. Psychology ; gamma -aminobutyric acid ; gamma-Aminobutyric Acid - pharmacology ; HALIDES ; HALOGEN COMPOUNDS ; Ion Channels - drug effects ; Ion Channels - metabolism ; ISOTOPES ; KINETICS ; LIGHT NUCLEI ; MAGNESIUM CHLORIDES ; MAGNESIUM COMPOUNDS ; Male ; MAMMALS ; Mathematics ; MEMBRANE PROTEINS ; MEMBRANE TRANSPORT ; MEMBRANES ; Models, Theoretical ; Molecular and cellular biology ; NERVOUS SYSTEM ; NEUROREGULATORS ; NUCLEI ; ODD-ODD NUCLEI ; ORGANIC ACIDS ; ORGANIC COMPOUNDS ; ORGANS ; PROTEINS ; RADIOISOTOPES ; RATS ; Rats, Inbred Strains ; REACTION KINETICS ; RECEPTORS ; Receptors, GABA-A - drug effects ; Receptors, GABA-A - metabolism ; RODENTS ; SENSITIVITY ; VERTEBRATES ; YEARS LIVING RADIOISOTOPES</subject><ispartof>Biochemistry (Easton), 1987-12, Vol.26 (24), p.7556-7562</ispartof><rights>1988 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a422t-4c73bc692b364dd547b7f13a548d22fc1f8ca92103153af177c347c4fe46b37b3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/bi00398a004$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/bi00398a004$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,885,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=7743027$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/2447945$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://www.osti.gov/biblio/5438040$$D View this record in Osti.gov$$Hfree_for_read</backlink></links><search><creatorcontrib>Cash, Derek J</creatorcontrib><creatorcontrib>Subbarao, Katragadda</creatorcontrib><creatorcontrib>Univ. of Missouri, St. Louis</creatorcontrib><title>Desensitization of .gamma.-aminobutyric acid receptor from rat brain: two distinguishable receptors on the same membrane</title><title>Biochemistry (Easton)</title><addtitle>Biochemistry</addtitle><description>Transmembrane chloride flux mediated by gamma-aminobutyric acid (GABA) receptor can be measured with a mammalian brain homogenate preparation containing sealed membrane vesicles. The preparation can be mixed rapidly with solutions of defined composition. Influx of 36Cl- tracer initiated by mixing with GABA was rapidly terminated by mixing with bicuculline methiodide. The decrease in the isotope influx measurement due to prior incubation of the vesicle preparation with GABA, which increased with preincubation time and GABA concentration, was attributed to desensitization of the GABA receptor. By varying the time of preincubation with GABA between 10 ms and 50 s with quench-flow technique, the desensitization rates could be measured over their whole time course independently of the chloride ion flux rate. Most of the receptor activity decreased in a fast phase of desensitization complete in 200 ms (t 1/2 = 32 ms) at saturation with GABA. Remaining activity was desensitized in a few seconds (t 1/2 = 533 ms). These two phases of desensitization were each kinetically first order and were shown to correspond with two distinguishable GABA receptors on the same membrane. The receptor activities could be estimated, and the faster desensitizing receptor was the predominant one, giving on average ca. 80% of the total activity. The half-response concentrations were similar, 150 and 114 microM for the major and minor receptors, respectively. The dependence on GABA concentration indicated that desensitization is mediated by two GABA binding sites. The fast desensitization rate was approximately 20-fold faster than previously reported rates while the slower desensitization rate was slightly faster than previously reported rates.</description><subject>550201 - Biochemistry- Tracer Techniques</subject><subject>ALKALINE EARTH METAL COMPOUNDS</subject><subject>AMINO ACIDS</subject><subject>AMINOBUTYRIC ACID</subject><subject>ANIMALS</subject><subject>AUTONOMIC NERVOUS SYSTEM AGENTS</subject><subject>BASIC BIOLOGICAL SCIENCES</subject><subject>BETA DECAY RADIOISOTOPES</subject><subject>BETA-MINUS DECAY RADIOISOTOPES</subject><subject>Bicuculline - pharmacology</subject><subject>BIOCHEMICAL REACTION KINETICS</subject><subject>Biological and medical sciences</subject><subject>BODY</subject><subject>BRAIN</subject><subject>CALCIUM CHLORIDES</subject><subject>CALCIUM COMPOUNDS</subject><subject>CALCIUM HALIDES</subject><subject>CARBOXYLIC ACIDS</subject><subject>CELL CONSTITUENTS</subject><subject>Cell Membrane - metabolism</subject><subject>CELL MEMBRANES</subject><subject>Cell receptors</subject><subject>Cell structures and functions</subject><subject>CENTRAL NERVOUS SYSTEM</subject><subject>Cerebral Cortex - metabolism</subject><subject>CHLORIDES</subject><subject>Chlorides - metabolism</subject><subject>CHLORINE 36</subject><subject>CHLORINE COMPOUNDS</subject><subject>CHLORINE ISOTOPES</subject><subject>DRUGS</subject><subject>ELECTRON CAPTURE RADIOISOTOPES</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>gamma -aminobutyric acid</subject><subject>gamma-Aminobutyric Acid - pharmacology</subject><subject>HALIDES</subject><subject>HALOGEN COMPOUNDS</subject><subject>Ion Channels - drug effects</subject><subject>Ion Channels - metabolism</subject><subject>ISOTOPES</subject><subject>KINETICS</subject><subject>LIGHT NUCLEI</subject><subject>MAGNESIUM CHLORIDES</subject><subject>MAGNESIUM COMPOUNDS</subject><subject>Male</subject><subject>MAMMALS</subject><subject>Mathematics</subject><subject>MEMBRANE PROTEINS</subject><subject>MEMBRANE TRANSPORT</subject><subject>MEMBRANES</subject><subject>Models, Theoretical</subject><subject>Molecular and cellular biology</subject><subject>NERVOUS SYSTEM</subject><subject>NEUROREGULATORS</subject><subject>NUCLEI</subject><subject>ODD-ODD NUCLEI</subject><subject>ORGANIC ACIDS</subject><subject>ORGANIC COMPOUNDS</subject><subject>ORGANS</subject><subject>PROTEINS</subject><subject>RADIOISOTOPES</subject><subject>RATS</subject><subject>Rats, Inbred Strains</subject><subject>REACTION KINETICS</subject><subject>RECEPTORS</subject><subject>Receptors, GABA-A - drug effects</subject><subject>Receptors, GABA-A - metabolism</subject><subject>RODENTS</subject><subject>SENSITIVITY</subject><subject>VERTEBRATES</subject><subject>YEARS LIVING RADIOISOTOPES</subject><issn>0006-2960</issn><issn>1520-4995</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1987</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqF0c9rFDEUB_BBlLqtnjwLQaQeZNb8mslMb1p_VCgouIq38JJJuqmTyZpksPWvN2WWxYPgKYT3yePlfavqCcFrgil5pRzGrO8AY36vWpGG4pr3fXO_WmGM25r2LX5YHad0Xa4cC35UHVHORc-bVXXz1iQzJZfdb8guTChYtL4C72Fdg3dTUHO-jU4j0G5A0WizyyEiG4NHETJSEdx0hvKvgAaXspuuZpe2oEZzwAmVtnlrUAJvkDe-vJnMo-qBhTGZx_vzpPr6_t3m_KK-_PTh4_nryxo4pbnmWjCl254q1vJhaLhQwhIGDe8GSq0mttPQU4IZaRhYIoRmXGhuDW8VE4qdVM-WvqFMJ5N22eitDtNkdJYNZ11ZSUGnC9rF8HM2KUvvkjbjWAYNc5JC9ISwtv8vJLzjrO26Al8uUMeQUjRW7qLzEG8lwfIuNflXakU_3bedlTfDwe5jKvXn-zokDaMtC9QuHZgQnGEqCqsXVpIwN4cyxB-yFUw0cvP5i8Tdt82bC_Zd3n37xeJBJ3kd5jiVJP454B8ATrrD</recordid><startdate>19871201</startdate><enddate>19871201</enddate><creator>Cash, Derek J</creator><creator>Subbarao, Katragadda</creator><general>American Chemical Society</general><scope>BSCLL</scope><scope>IQODW</scope><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>7TK</scope><scope>8FD</scope><scope>FR3</scope><scope>M7Z</scope><scope>P64</scope><scope>7X8</scope><scope>OTOTI</scope></search><sort><creationdate>19871201</creationdate><title>Desensitization of .gamma.-aminobutyric acid receptor from rat brain: two distinguishable receptors on the same membrane</title><author>Cash, Derek J ; Subbarao, Katragadda</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a422t-4c73bc692b364dd547b7f13a548d22fc1f8ca92103153af177c347c4fe46b37b3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1987</creationdate><topic>550201 - Biochemistry- Tracer Techniques</topic><topic>ALKALINE EARTH METAL COMPOUNDS</topic><topic>AMINO ACIDS</topic><topic>AMINOBUTYRIC ACID</topic><topic>ANIMALS</topic><topic>AUTONOMIC NERVOUS SYSTEM AGENTS</topic><topic>BASIC BIOLOGICAL SCIENCES</topic><topic>BETA DECAY RADIOISOTOPES</topic><topic>BETA-MINUS DECAY RADIOISOTOPES</topic><topic>Bicuculline - pharmacology</topic><topic>BIOCHEMICAL REACTION KINETICS</topic><topic>Biological and medical sciences</topic><topic>BODY</topic><topic>BRAIN</topic><topic>CALCIUM CHLORIDES</topic><topic>CALCIUM COMPOUNDS</topic><topic>CALCIUM HALIDES</topic><topic>CARBOXYLIC ACIDS</topic><topic>CELL CONSTITUENTS</topic><topic>Cell Membrane - metabolism</topic><topic>CELL MEMBRANES</topic><topic>Cell receptors</topic><topic>Cell structures and functions</topic><topic>CENTRAL NERVOUS SYSTEM</topic><topic>Cerebral Cortex - metabolism</topic><topic>CHLORIDES</topic><topic>Chlorides - metabolism</topic><topic>CHLORINE 36</topic><topic>CHLORINE COMPOUNDS</topic><topic>CHLORINE ISOTOPES</topic><topic>DRUGS</topic><topic>ELECTRON CAPTURE RADIOISOTOPES</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>gamma -aminobutyric acid</topic><topic>gamma-Aminobutyric Acid - pharmacology</topic><topic>HALIDES</topic><topic>HALOGEN COMPOUNDS</topic><topic>Ion Channels - drug effects</topic><topic>Ion Channels - metabolism</topic><topic>ISOTOPES</topic><topic>KINETICS</topic><topic>LIGHT NUCLEI</topic><topic>MAGNESIUM CHLORIDES</topic><topic>MAGNESIUM COMPOUNDS</topic><topic>Male</topic><topic>MAMMALS</topic><topic>Mathematics</topic><topic>MEMBRANE PROTEINS</topic><topic>MEMBRANE TRANSPORT</topic><topic>MEMBRANES</topic><topic>Models, Theoretical</topic><topic>Molecular and cellular biology</topic><topic>NERVOUS SYSTEM</topic><topic>NEUROREGULATORS</topic><topic>NUCLEI</topic><topic>ODD-ODD NUCLEI</topic><topic>ORGANIC ACIDS</topic><topic>ORGANIC COMPOUNDS</topic><topic>ORGANS</topic><topic>PROTEINS</topic><topic>RADIOISOTOPES</topic><topic>RATS</topic><topic>Rats, Inbred Strains</topic><topic>REACTION KINETICS</topic><topic>RECEPTORS</topic><topic>Receptors, GABA-A - drug effects</topic><topic>Receptors, GABA-A - metabolism</topic><topic>RODENTS</topic><topic>SENSITIVITY</topic><topic>VERTEBRATES</topic><topic>YEARS LIVING RADIOISOTOPES</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Cash, Derek J</creatorcontrib><creatorcontrib>Subbarao, Katragadda</creatorcontrib><creatorcontrib>Univ. of Missouri, St. Louis</creatorcontrib><collection>Istex</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Neurosciences Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biochemistry Abstracts 1</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>MEDLINE - Academic</collection><collection>OSTI.GOV</collection><jtitle>Biochemistry (Easton)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Cash, Derek J</au><au>Subbarao, Katragadda</au><aucorp>Univ. of Missouri, St. Louis</aucorp><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Desensitization of .gamma.-aminobutyric acid receptor from rat brain: two distinguishable receptors on the same membrane</atitle><jtitle>Biochemistry (Easton)</jtitle><addtitle>Biochemistry</addtitle><date>1987-12-01</date><risdate>1987</risdate><volume>26</volume><issue>24</issue><spage>7556</spage><epage>7562</epage><pages>7556-7562</pages><issn>0006-2960</issn><eissn>1520-4995</eissn><abstract>Transmembrane chloride flux mediated by gamma-aminobutyric acid (GABA) receptor can be measured with a mammalian brain homogenate preparation containing sealed membrane vesicles. The preparation can be mixed rapidly with solutions of defined composition. Influx of 36Cl- tracer initiated by mixing with GABA was rapidly terminated by mixing with bicuculline methiodide. The decrease in the isotope influx measurement due to prior incubation of the vesicle preparation with GABA, which increased with preincubation time and GABA concentration, was attributed to desensitization of the GABA receptor. By varying the time of preincubation with GABA between 10 ms and 50 s with quench-flow technique, the desensitization rates could be measured over their whole time course independently of the chloride ion flux rate. Most of the receptor activity decreased in a fast phase of desensitization complete in 200 ms (t 1/2 = 32 ms) at saturation with GABA. Remaining activity was desensitized in a few seconds (t 1/2 = 533 ms). These two phases of desensitization were each kinetically first order and were shown to correspond with two distinguishable GABA receptors on the same membrane. The receptor activities could be estimated, and the faster desensitizing receptor was the predominant one, giving on average ca. 80% of the total activity. The half-response concentrations were similar, 150 and 114 microM for the major and minor receptors, respectively. The dependence on GABA concentration indicated that desensitization is mediated by two GABA binding sites. The fast desensitization rate was approximately 20-fold faster than previously reported rates while the slower desensitization rate was slightly faster than previously reported rates.</abstract><cop>Washington, DC</cop><pub>American Chemical Society</pub><pmid>2447945</pmid><doi>10.1021/bi00398a004</doi><tpages>7</tpages></addata></record> |
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| ispartof | Biochemistry (Easton), 1987-12, Vol.26 (24), p.7556-7562 |
| issn | 0006-2960 1520-4995 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_77911369 |
| source | MEDLINE; ACS Publications |
| subjects | 550201 - Biochemistry- Tracer Techniques ALKALINE EARTH METAL COMPOUNDS AMINO ACIDS AMINOBUTYRIC ACID ANIMALS AUTONOMIC NERVOUS SYSTEM AGENTS BASIC BIOLOGICAL SCIENCES BETA DECAY RADIOISOTOPES BETA-MINUS DECAY RADIOISOTOPES Bicuculline - pharmacology BIOCHEMICAL REACTION KINETICS Biological and medical sciences BODY BRAIN CALCIUM CHLORIDES CALCIUM COMPOUNDS CALCIUM HALIDES CARBOXYLIC ACIDS CELL CONSTITUENTS Cell Membrane - metabolism CELL MEMBRANES Cell receptors Cell structures and functions CENTRAL NERVOUS SYSTEM Cerebral Cortex - metabolism CHLORIDES Chlorides - metabolism CHLORINE 36 CHLORINE COMPOUNDS CHLORINE ISOTOPES DRUGS ELECTRON CAPTURE RADIOISOTOPES Fundamental and applied biological sciences. Psychology gamma -aminobutyric acid gamma-Aminobutyric Acid - pharmacology HALIDES HALOGEN COMPOUNDS Ion Channels - drug effects Ion Channels - metabolism ISOTOPES KINETICS LIGHT NUCLEI MAGNESIUM CHLORIDES MAGNESIUM COMPOUNDS Male MAMMALS Mathematics MEMBRANE PROTEINS MEMBRANE TRANSPORT MEMBRANES Models, Theoretical Molecular and cellular biology NERVOUS SYSTEM NEUROREGULATORS NUCLEI ODD-ODD NUCLEI ORGANIC ACIDS ORGANIC COMPOUNDS ORGANS PROTEINS RADIOISOTOPES RATS Rats, Inbred Strains REACTION KINETICS RECEPTORS Receptors, GABA-A - drug effects Receptors, GABA-A - metabolism RODENTS SENSITIVITY VERTEBRATES YEARS LIVING RADIOISOTOPES |
| title | Desensitization of .gamma.-aminobutyric acid receptor from rat brain: two distinguishable receptors on the same membrane |
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