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Spatial and Temporal Aspects of Exocytosis Studied on the Isolated Plasma Membranes

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Exocytosis and Endocytosis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2233))

Abstract

Exocytosis of large-dense core vesicles in neuroendocrine cells is a highly regulated, calcium-dependent process, mediated by networks of interrelated proteins and lipids. Here, I describe experimental procedures for studies of selective spatial and temporal aspects of exocytosis at the plasma membrane, or in its proximity, using adrenal chromaffin cells. The assay utilizes primary cells subjected to a brief ultrasonic pulse, resulting in the formation of thin, flat inside-out plasma membranes with attached secretory vesicles and elements of cell cytoskeleton. In this model, secretion of plasma membrane–attached secretory vesicles was found to be dependent on calcium and sensitive to clostridial neurotoxins. Depending on the probe selected for secretory vesicle cargo, protein, and/or lipid detection, this simple assay is versatile, fast and inexpensive, and offers excellent spatial resolution.

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Acknowledgments

I thank Dr. N. Raimundo for a discussion, and Dr. A. Milosevic for help with the figures. This work is supported by the Emmy Noether Young Investigator Award (1702/1) of the German Research Foundation (DFG) and the Schram Stiftung (T287/25457). No competing financial interests are declared.

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Authors and Affiliations

  1. European Neuroscience Institute (ENI), A Joint Initiative of the University Medical Center Göttingen and the Max Planck Society, Göttingen, Germany

    Ira Milosevic

  2. Wellcome Centre for Human Genetics, Nuffield Department of Medicine, NIHR Oxford Biomedical Research Centre, University of Oxford, Oxford, UK

    Ira Milosevic

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  1. Ira Milosevic
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Correspondence to Ira Milosevic .

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Editors and Affiliations

  1. Department of Infection, Immunity & Inflammation, Institut Cochin (Institut National de la santé et de la recherche médicale, Centre National de la Recherche Scientifique and Université de Paris), Paris, France

    Florence Niedergang

  2. Centre National de la Recherche Scientifique, Institut des Neurosciences Cellulaires et Intégratives, Université de Strasbourg, Strasbourg, France

    Nicolas Vitale

  3. Centre National de la Recherche Scientifique, Institut des Neurosciences Cellulaires et Intégratives, Université de Strasbourg, Strasbourg, France

    Stéphane Gasman

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Milosevic, I. (2021). Spatial and Temporal Aspects of Exocytosis Studied on the Isolated Plasma Membranes. In: Niedergang, F., Vitale, N., Gasman, S. (eds) Exocytosis and Endocytosis. Methods in Molecular Biology, vol 2233. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1044-2_21

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  • DOI: https://doi.org/10.1007/978-1-0716-1044-2_21

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1043-5

  • Online ISBN: 978-1-0716-1044-2

  • eBook Packages: Springer Protocols

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