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J. Biol. Chem., Vol. 277, Issue 32, 29101-29107, August 9, 2002
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From the Department of Chemistry, University of North Carolina at
Chapel Hill, Chapel Hill, North Carolina 27599
During exocytosis, vesicles in secretory cells
fuse with the cellular membrane and release their contents in a
Ca2+-dependent process. Release occurs
initially through a fusion pore, and its rate is limited by the
dissociation of the matrix-associated contents. To determine whether
this dissociation is promoted by osmotic forces, we have examined the
effects of elevated osmotic pressure on release and extrusion from
vesicles at mast and chromaffin cells. The identity of the molecules
released and the time course of extrusion were measured with fast scan
cyclic voltammetry at carbon fiber microelectrodes. In external
solutions of high osmolarity, release events following entry of
divalent ions (Ba2+ or Ca2+) were less
frequent. However, the vesicles appeared to be fused to the membrane
without extruding their contents, since the maximal observed
concentrations of events were less than 7% of those evoked in isotonic
media. Such an isolated, intermediate fusion state, which we term
"kiss-and-hold," was confirmed by immunohistochemistry at
chromaffin cells. Transient exposure of cells in the kiss and hold
state to isotonic solutions evoked massive release. These results
demonstrate that an osmotic gradient across the fusion pore is an
important driving force for exocytotic extrusion of granule contents
from secretory cells following fusion pore formation.
Temporal Separation of Vesicle Release from
Vesicle Fusion during Exocytosis*
*
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed. Tel.: 919-962-1472;
Fax: 919-962-2388; E-mail: rmw@unc.edu.
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