Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors

Autores
Rayes, Diego Hernán; Spitzmaul, Guillermo Federico; Sine, Steven M.; Bouzat, Cecilia Beatriz
Año de publicación
2005
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
The receptor chimera 7–5HT3A has served as a prototype for understanding the pharmacology of 7 neuronal nicotinic receptors, yet its low single channel conductance has prevented studies of the activation kinetics of single receptor channels. In this study, we show that introducing mutations in the M3–M4 cytoplasmic linker of the chimera alters neither the apparent affinity for the agonist nor the EC50 but increases the amplitude of agonist-evoked single channel currents to enable kinetic analysis. Channel events appear as single brief openings flanked by long closings or as bursts of several openings in quick succession. Both the open and closed time distributions are described as the sum of multiple exponential components, but these do not change over a wide range of acetylcholine (ACh), nicotine, or choline concentrations. Bursts elicited by a saturating concentration of ACh contain brief and long openings and closings, and a cyclic scheme containing two open and two closed states is found to adequately describe the data. The analysis indicates that once fully occupied, the receptor opens rapidly and efficiently, and closes and reopens several times before it desensitizes. Channel closing and desensitization occur at similar rates and account for the invariant open and closed time distributions.
Fil: Rayes, Diego Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina
Fil: Spitzmaul, Guillermo Federico. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Sine, Steven M.. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina. Mayo Clinic College of Medicine; Estados Unidos
Fil: Bouzat, Cecilia Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina
Materia
Nicotinic
Receptor
Activation
Homomeric
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/48485
Acceder
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network_name_str CONICET Digital (CONICET)
spelling Single-Channel Kinetic Analysis of Chimeric 7-5HT3A ReceptorsRayes, Diego HernánSpitzmaul, Guillermo FedericoSine, Steven M.Bouzat, Cecilia BeatrizNicotinicReceptorActivationHomomerichttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1The receptor chimera 7–5HT3A has served as a prototype for understanding the pharmacology of 7 neuronal nicotinic receptors, yet its low single channel conductance has prevented studies of the activation kinetics of single receptor channels. In this study, we show that introducing mutations in the M3–M4 cytoplasmic linker of the chimera alters neither the apparent affinity for the agonist nor the EC50 but increases the amplitude of agonist-evoked single channel currents to enable kinetic analysis. Channel events appear as single brief openings flanked by long closings or as bursts of several openings in quick succession. Both the open and closed time distributions are described as the sum of multiple exponential components, but these do not change over a wide range of acetylcholine (ACh), nicotine, or choline concentrations. Bursts elicited by a saturating concentration of ACh contain brief and long openings and closings, and a cyclic scheme containing two open and two closed states is found to adequately describe the data. The analysis indicates that once fully occupied, the receptor opens rapidly and efficiently, and closes and reopens several times before it desensitizes. Channel closing and desensitization occur at similar rates and account for the invariant open and closed time distributions.Fil: Rayes, Diego Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; ArgentinaFil: Spitzmaul, Guillermo Federico. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Sine, Steven M.. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina. Mayo Clinic College of Medicine; Estados UnidosFil: Bouzat, Cecilia Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; ArgentinaAmerican Society for Pharmacology and Experimental Therapeutics2005-08info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/48485Rayes, Diego Hernán; Spitzmaul, Guillermo Federico; Sine, Steven M.; Bouzat, Cecilia Beatriz; Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors; American Society for Pharmacology and Experimental Therapeutics; Molecular Pharmacology; 68; 5; 8-2005; 1475-14830026-895CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1124/mol.105.015438info:eu-repo/semantics/altIdentifier/url/http://molpharm.aspetjournals.org/content/68/5/1475info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T10:03:17Zoai:ri.conicet.gov.ar:11336/48485instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-03 10:03:17.366CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
title Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
spellingShingle Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
Rayes, Diego Hernán
Nicotinic
Receptor
Activation
Homomeric
title_short Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
title_full Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
title_fullStr Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
title_full_unstemmed Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
title_sort Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors
dc.creator.none.fl_str_mv Rayes, Diego Hernán
Spitzmaul, Guillermo Federico
Sine, Steven M.
Bouzat, Cecilia Beatriz
author Rayes, Diego Hernán
author_facet Rayes, Diego Hernán
Spitzmaul, Guillermo Federico
Sine, Steven M.
Bouzat, Cecilia Beatriz
author_role author
author2 Spitzmaul, Guillermo Federico
Sine, Steven M.
Bouzat, Cecilia Beatriz
author2_role author
author
author
dc.subject.none.fl_str_mv Nicotinic
Receptor
Activation
Homomeric
topic Nicotinic
Receptor
Activation
Homomeric
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv The receptor chimera 7–5HT3A has served as a prototype for understanding the pharmacology of 7 neuronal nicotinic receptors, yet its low single channel conductance has prevented studies of the activation kinetics of single receptor channels. In this study, we show that introducing mutations in the M3–M4 cytoplasmic linker of the chimera alters neither the apparent affinity for the agonist nor the EC50 but increases the amplitude of agonist-evoked single channel currents to enable kinetic analysis. Channel events appear as single brief openings flanked by long closings or as bursts of several openings in quick succession. Both the open and closed time distributions are described as the sum of multiple exponential components, but these do not change over a wide range of acetylcholine (ACh), nicotine, or choline concentrations. Bursts elicited by a saturating concentration of ACh contain brief and long openings and closings, and a cyclic scheme containing two open and two closed states is found to adequately describe the data. The analysis indicates that once fully occupied, the receptor opens rapidly and efficiently, and closes and reopens several times before it desensitizes. Channel closing and desensitization occur at similar rates and account for the invariant open and closed time distributions.
Fil: Rayes, Diego Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina
Fil: Spitzmaul, Guillermo Federico. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Sine, Steven M.. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; Argentina. Mayo Clinic College of Medicine; Estados Unidos
Fil: Bouzat, Cecilia Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentina
description The receptor chimera 7–5HT3A has served as a prototype for understanding the pharmacology of 7 neuronal nicotinic receptors, yet its low single channel conductance has prevented studies of the activation kinetics of single receptor channels. In this study, we show that introducing mutations in the M3–M4 cytoplasmic linker of the chimera alters neither the apparent affinity for the agonist nor the EC50 but increases the amplitude of agonist-evoked single channel currents to enable kinetic analysis. Channel events appear as single brief openings flanked by long closings or as bursts of several openings in quick succession. Both the open and closed time distributions are described as the sum of multiple exponential components, but these do not change over a wide range of acetylcholine (ACh), nicotine, or choline concentrations. Bursts elicited by a saturating concentration of ACh contain brief and long openings and closings, and a cyclic scheme containing two open and two closed states is found to adequately describe the data. The analysis indicates that once fully occupied, the receptor opens rapidly and efficiently, and closes and reopens several times before it desensitizes. Channel closing and desensitization occur at similar rates and account for the invariant open and closed time distributions.
publishDate 2005
dc.date.none.fl_str_mv 2005-08
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/48485
Rayes, Diego Hernán; Spitzmaul, Guillermo Federico; Sine, Steven M.; Bouzat, Cecilia Beatriz; Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors; American Society for Pharmacology and Experimental Therapeutics; Molecular Pharmacology; 68; 5; 8-2005; 1475-1483
0026-895
CONICET Digital
CONICET
url http://hdl.handle.net/11336/48485
identifier_str_mv Rayes, Diego Hernán; Spitzmaul, Guillermo Federico; Sine, Steven M.; Bouzat, Cecilia Beatriz; Single-Channel Kinetic Analysis of Chimeric 7-5HT3A Receptors; American Society for Pharmacology and Experimental Therapeutics; Molecular Pharmacology; 68; 5; 8-2005; 1475-1483
0026-895
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1124/mol.105.015438
info:eu-repo/semantics/altIdentifier/url/http://molpharm.aspetjournals.org/content/68/5/1475
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv American Society for Pharmacology and Experimental Therapeutics
publisher.none.fl_str_mv American Society for Pharmacology and Experimental Therapeutics
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.13397

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