Push-and-pull regulation of the fusion pore by synaptotagmin-7.

In chromaffin cells, Ca(2+) binding to synaptotagmin-1 and -7 triggers exocytosis by promoting fusion pore opening and fusion pore expansion. Synaptotagmins contain two C2 domains that both bind Ca(2+) and contribute to exocytosis; however, it remains unknown whether the C2 domains act similarly or differentially to promote opening and expansion of fusion pores. Here, we use patch amperometry measurements in WT and synaptotagmin-7-mutant chromaffin cells to analyze the role of Ca(2+) binding to the two synaptotagmin-7 C2 domains in exocytosis.

Quantitative investigations of amperometric spike feet suggest different controlling factors of the fusion pore in exocytosis at chromaffin cells.

Around 30% of exocytosis events recorded by amperometry at carbon fiber microelectrodes exhibit a pre-spike feature (PSF) termed a "foot". This wave is associated with the release of the neurotransmitters via a transitory fusion pore, whilst the large, main exocytotic spike is due to complete release. The amperometric data reported herein were obtained using bovine chromaffin cells stimulated with either potassium or barium ions, two commonly-employed elicitors of exocytosis.

Relationship between amperometric pre-spike feet and secretion granule composition in chromaffin cells: an overview.

Amperometry is a simple and powerful technique to study exocytosis at the single cell level. By positioning and polarizing (at an appropriate potential at which the molecules released by the cell can be oxidized) a carbon fiber microelectrode at the top of the cell, each exocytotic event is detected as an amperometric spike. More particularly, a portion of these spikes has previously been shown to present a foot, i.

Vesicular exocytosis under hypotonic conditions shows two distinct populations of dense core vesicles in bovine chromaffin cells.

Several previous reports have discussed the effects of external osmolarity on vesicular exocytotic processes. However, few of these studies considered hypotonic conditions on chromaffin cells. Herein, the exocytosis of catecholamines by chromaffin cells was investigated in a medium of low osmolarity (200 mOsm) by amperometry at carbon fiber microelectrodes.

Correlation between vesicle quantal size and fusion pore release in chromaffin cell exocytosis.

A significant number of exocytosis events recorded with amperometry demonstrate a prespike feature termed a "foot" and this foot has been correlated with messengers released via a transitory fusion pore before full exocytosis. We have compared amperometric spikes with a foot with spikes without a foot at chromaffin cells and found that the probability of detecting a distinct foot event is correlated to the amount of catecholamine released. The mean charge of the spikes with a foot was found to be twice that of the spikes without a foot, and the frequency of spikes displaying a foot was zero for small spikes increasing to approximately 50% for large spikes.

Dynamics of full fusion during vesicular exocytotic events: release of adrenaline by chromaffin cells.

Vesicular exocytosis is important in the communication between cells in complex organisms. It controls the release of specific chemical or biochemical messengers stored in the emitting cell, which elicit a response upon detection by the target cells. Secretion of a messenger molecule (a neurotransmitter) was measured electrochemically, which allowed the quantification of cellular events and the validation of current physicochemical models.