Neurotransmitters and hormones are released by Ca2+-triggered exocytosis. This tightly regulated process controls the delivery of chemical signals throughout the nervous and endocrine systems. At present, we have a very poor understanding of the molecular mechanism of Ca2+-triggered exocytosis, and this limits our ability to investigate fundamental processes such as synaptic transmission and endocrine regulation. Exocytosis entails the fusion of a vesicle with a cell's plasma membrane. A pivotal step in exocytosis is the formation of a fusion pore, which initiates membrane fusion. The fusion pore can be detected in biophysical measurements, but its molecular properties are poorly understood and its chemical composition is unknown. The experiments proposed here will investigate 1) the mechanism by which released substances permeate the fusion pore, 2) the mechanism by which fusion pores open and close, and 3) the mechanism by which the protein synaptotagmin regulates the dynamics of the fusion pore. 4) Additional experiments will test the hypothesis that membrane spanning segments of the proteins synaptotagmin, syntaxin, and synaptobrevin form the fusion pore. These experiments employ electrical measurements of flux and current through single fusion pores. The experiments will be performed in PC12 cells, a clonal cell line that exhibits Ca2+-triggered secretion, and in which molecular manipulations are easily carried out. The three specific aims of this proposal approach the question of the molecular properties of the fusion pore from different directions. The improved understanding of fusion pore permeation, dynamics, regulation, and structure emerging from this work will lead to a better understanding of how nerve and endocrine cells release substances. This in turn will improve our understanding of the many physiological functions and systems under neural and endocrine control.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS044057-03
Application #
6758025
Study Section
Molecular, Cellular and Developmental Neurosciences 2 (MDCN)
Program Officer
Stewart, Randall R
Project Start
2002-07-01
Project End
2007-06-30
Budget Start
2004-07-01
Budget End
2005-06-30
Support Year
3
Fiscal Year
2004
Total Cost
$205,159
Indirect Cost
Name
University of Wisconsin Madison
Department
Physiology
Type
Schools of Medicine
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Chiang, Chung-Wei; Chang, Che-Wei; Jackson, Meyer B (2018) The Transmembrane Domain of Synaptobrevin Influences Neurotransmitter Flux through Synaptic Fusion Pores. J Neurosci 38:7179-7191
Jackson, Meyer B (2017) Chemistry in a vesicle. J Gen Physiol 149:893-896
Chang, Che-Wei; Chiang, Chung-Wei; Jackson, Meyer B (2017) Fusion pores and their control of neurotransmitter and hormone release. J Gen Physiol 149:301-322
Chang, Che-Wei; Chiang, Chung-Wei; Gaffaney, Jon D et al. (2016) Lipid-anchored Synaptobrevin Provides Little or No Support for Exocytosis or Liposome Fusion. J Biol Chem 291:2848-57
McMahon, Shane M; Chang, Che-Wei; Jackson, Meyer B (2016) Multiple cytosolic calcium buffers in posterior pituitary nerve terminals. J Gen Physiol 147:243-54
Jackson, Meyer B (2016) The Hydrophobic Effect in Solute Partitioning and Interfacial Tension. Sci Rep 6:19265
Chang, Che-Wei; Jackson, Meyer B (2015) Synaptobrevin transmembrane domain influences exocytosis by perturbing vesicle membrane curvature. Biophys J 109:76-84
Chang, Che-Wei; Hui, Enfu; Bai, Jihong et al. (2015) A structural role for the synaptobrevin 2 transmembrane domain in dense-core vesicle fusion pores. J Neurosci 35:5772-80
McMahon, Shane M; Jackson, Meyer B (2014) In situ Ca2+ titration in the fluorometric study of intracellular Ca2+ binding. Cell Calcium 56:504-12
Yoo, Jejoong; Jackson, Meyer B; Cui, Qiang (2013) A comparison of coarse-grained and continuum models for membrane bending in lipid bilayer fusion pores. Biophys J 104:841-52

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