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Fusion Pore Size Limits 5‐HT Release From Single Enterochromaffin Cell Vesicles
Author(s) -
Raghupathi Ravinarayan,
Jessup Claire F.,
Lumsden Amanda L.,
Keating Damien J.
Publication year - 2016
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.25256
Subject(s) - enterochromaffin cell , exocytosis , vesicle , chemistry , biophysics , enteroendocrine cell , serotonin , neurotransmitter , microbiology and biotechnology , secretion , biology , biochemistry , membrane , receptor , endocrine system , hormone
Enterochromaffin cells are the major site of serotonin (5‐HT) synthesis and secretion providing ∼95% of the body's total 5‐HT. 5‐HT can act as a neurotransmitter or hormone and has several important endocrine and paracrine roles. We have previously demonstrated that EC cells release small amounts of 5‐HT per exocytosis event compared to other endocrine cells. We utilized a recently developed method to purify EC cells to demonstrate the mechanisms underlying 5‐HT packaging and release. Using the fluorescent probe FFN511, we demonstrate that EC cells express VMAT and that VMAT plays a functional role in 5‐HT loading into vesicles. Carbon fiber amperometry studies illustrate that the amount of 5‐HT released per exocytosis event from EC cells is dependent on both VMAT and the H + ‐ATPase pump, as demonstrated with reserpine or bafilomycin, respectively. We also demonstrate that increasing the amount of 5‐HT loaded into EC cell vesicles does not result in an increase in quantal release. As this indicates that fusion pore size may be a limiting factor involved, we compared pore diameter in EC and chromaffin cells by assessing the vesicle capture of different‐sized fluorescent probes to measure the extent of fusion pore dilation. This identified that EC cells have a reduced fusion pore expansion that does not exceed 9 nm in diameter. These results demonstrate that the small amounts of 5‐HT released per fusion event in EC cells can be explained by a smaller fusion pore that limits 5‐HT release capacity from individual vesicles. J. Cell. Physiol. 231: 1593–1600, 2016. © 2015 Wiley Periodicals, Inc.