Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies

Substance P and other polycationic peptides are thought to stimulate mast cell degranulation via direct activation of G proteins. We investigated the ability of extracellularly applied substance P to translocate into mast cells and the ability of intracellularly applied substance P to stimulate degr...

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Автори: Lorenz, Dorothea, Wiesner, Burkhard, Zipper, Josef, Winkler, Anett, Krause, Eberhard, Beyermann, Michael, Lindau, Manfred, Bienert, Michael
Формат: Стаття
Мова:English
Опубліковано: The Rockefeller University Press 1998
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Онлайн доступ:https://ncbi.nlm.nih.gov/pmc/articles/PMC2229441/
https://ncbi.nlm.nih.gov/pubmed/9806967
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spelling pubmed-22294412008-04-22 Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies Lorenz, Dorothea Wiesner, Burkhard Zipper, Josef Winkler, Anett Krause, Eberhard Beyermann, Michael Lindau, Manfred Bienert, Michael J Gen Physiol Article Substance P and other polycationic peptides are thought to stimulate mast cell degranulation via direct activation of G proteins. We investigated the ability of extracellularly applied substance P to translocate into mast cells and the ability of intracellularly applied substance P to stimulate degranulation. In addition, we studied by reverse transcription–-PCR whether substance P-specific receptors are present in the mast cell membrane. To study translocation, a biologically active and enzymatically stable fluorescent analogue of substance P was synthesized. A rapid, substance P receptor- and energy-independent uptake of this peptide into pertussis toxin-treated and -untreated mast cells was demonstrated using confocal laser scanning microscopy. The peptide was shown to localize preferentially on or inside the mast cell granules using electron microscopic autoradiography with (125)I-labeled all-D substance P and (3)H-labeled substance P. Cell membrane capacitance measurements using the patch-clamp technique demonstrated that intracellularly applied substance P induced calcium transients and activated mast cell exocytosis with a time delay that depended on peptide concentration (delay of 100–500 s at concentrations of substance P from 50 to 5 μM). Degranulation in response to intracellularly applied substance P was inhibited by GDPβS and pertussis toxin, suggesting that substance P acts via G protein activation. These results support the recently proposed model of a receptor-independent mechanism of peptide-induced mast cell degranulation, which assumes a direct interaction of peptides with G protein α subunits subsequent to their translocation across the plasma membrane. The Rockefeller University Press 1998-11-01 /pmc/articles/PMC2229441/ /pubmed/9806967 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
institution US National Library of Medicine
collection PubMed Central
language English
format Article
topic Article
spellingShingle Article
Lorenz, Dorothea
Wiesner, Burkhard
Zipper, Josef
Winkler, Anett
Krause, Eberhard
Beyermann, Michael
Lindau, Manfred
Bienert, Michael
Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies
description Substance P and other polycationic peptides are thought to stimulate mast cell degranulation via direct activation of G proteins. We investigated the ability of extracellularly applied substance P to translocate into mast cells and the ability of intracellularly applied substance P to stimulate degranulation. In addition, we studied by reverse transcription–-PCR whether substance P-specific receptors are present in the mast cell membrane. To study translocation, a biologically active and enzymatically stable fluorescent analogue of substance P was synthesized. A rapid, substance P receptor- and energy-independent uptake of this peptide into pertussis toxin-treated and -untreated mast cells was demonstrated using confocal laser scanning microscopy. The peptide was shown to localize preferentially on or inside the mast cell granules using electron microscopic autoradiography with (125)I-labeled all-D substance P and (3)H-labeled substance P. Cell membrane capacitance measurements using the patch-clamp technique demonstrated that intracellularly applied substance P induced calcium transients and activated mast cell exocytosis with a time delay that depended on peptide concentration (delay of 100–500 s at concentrations of substance P from 50 to 5 μM). Degranulation in response to intracellularly applied substance P was inhibited by GDPβS and pertussis toxin, suggesting that substance P acts via G protein activation. These results support the recently proposed model of a receptor-independent mechanism of peptide-induced mast cell degranulation, which assumes a direct interaction of peptides with G protein α subunits subsequent to their translocation across the plasma membrane.
author Lorenz, Dorothea
Wiesner, Burkhard
Zipper, Josef
Winkler, Anett
Krause, Eberhard
Beyermann, Michael
Lindau, Manfred
Bienert, Michael
author_facet Lorenz, Dorothea
Wiesner, Burkhard
Zipper, Josef
Winkler, Anett
Krause, Eberhard
Beyermann, Michael
Lindau, Manfred
Bienert, Michael
author_sort Lorenz, Dorothea
title Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies
title_short Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies
title_full Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies
title_fullStr Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies
title_full_unstemmed Mechanism of Peptide-induced Mast Cell Degranulation : Translocation and Patch-Clamp Studies
title_sort mechanism of peptide-induced mast cell degranulation : translocation and patch-clamp studies
publisher The Rockefeller University Press
publisher_facet The Rockefeller University Press
publishDate 1998
url https://ncbi.nlm.nih.gov/pmc/articles/PMC2229441/
https://ncbi.nlm.nih.gov/pubmed/9806967
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