scholarly journals Annexin II in exocytosis: catecholamine secretion requires the translocation of p36 to the subplasmalemmal region in chromaffin cells.

1996 ◽  
Vol 133 (6) ◽  
pp. 1217-1236 ◽  
Author(s):  
S Chasserot-Golaz ◽  
N Vitale ◽  
I Sagot ◽  
B Delouche ◽  
S Dirrig ◽  
...  
Keyword(s):  
Chromaffin Cells ◽  
Insoluble Fraction ◽  
Catecholamine Secretion ◽  
Annexin Ii ◽  
Cell Periphery ◽  
High K ◽  
Confocal Immunofluorescence ◽  
S 100 ◽  
Triton X ◽  
Insoluble Protein Fraction

Annexin II is a Ca(2+)-dependent membrane-binding protein present in a wide variety of cells and tissues. Within cells, annexin II is found either as a 36-kD monomer (p36) or as a heterotetrameric complex (p90) coupled with the S-100-related protein, p11. Annexin II has been suggested to be involved in exocytosis as it can restore the secretory responsiveness of permeabilized chromaffin cells. By quantitative confocal immunofluorescence, immunoreplica analysis and immunoprecipitation, we show here the translocation of p36 from the cytosol to a subplasmalemmal Triton X-100 insoluble fraction in chromaffin cells following nicotinic stimulation. A synthetic peptide corresponding to the NH2-terminal domain of p36 which contains the phosphorylation sites was microinjected into individual chromaffin cells and catecholamine secretion was monitored by amperometry. This peptide blocked completely the nicotine-induced recruitment of p36 to the cell periphery and strongly inhibited exocytosis evoked by either nicotine or high K+. The light chain of annexin II, p11, was selectively expressed by adrenergic chromaffin cells, and was only present in the subplasmalemmal Triton X-100 insoluble protein fraction of both resting and stimulated cells. p11 can modify the Ca(2+)- and/or the phospholipid-binding properties of p36. We found that loss Ca2+ was required to stimulate the translocation of p36 and to trigger exocytosis in adrenergic chromaffin cells. Our findings suggest that the translocation of p36 to the subplasmalemmal region is an essential event in regulated exocytosis and support the idea that the presence of p11 in adrenergic cells may confer a higher Ca2+ affinity to the exocytotic pathway in these cells.

scholarly journals Effect of inositol 1,4,5-trisphosphate receptor stimulation on mitochondrial [Ca2+] and secretion in chromaffin cells

2002 ◽  
Vol 365 (2) ◽  
pp. 451-459 ◽  
Author(s):  
Mayte MONTERO ◽  
Maria Teresa ALONSO ◽  
Almudena ALBILLOS ◽  
Inmaculada CUCHILLO-IBÁÑEZ ◽  
Román OLIVARES ◽  
...  
Keyword(s):  
Chromaffin Cells ◽  
Ryanodine Receptors ◽  
Catecholamine Secretion ◽  
Mitochondrial Inhibitors ◽  
High K ◽  
Inositol 1,4,5 Trisphosphate ◽  
Insp3 Receptors ◽  
Voltage Dependent ◽  
Trisphosphate Receptor ◽  
Stimulation Of

Ca2+ uptake by mitochondria is a potentially important buffering system able to control cytosolic [Ca2+]. In chromaffin cells, we have shown previously that stimulation of either Ca2+ entry or Ca2+ release via ryanodine receptors triggers large increases in mitochondrial [Ca2+] ([Ca2+]M) approaching the millimolar range, whose blockade dramatically enhances catecholamine secretion [Montero, Alonso, Carnicero, Cuchillo-Ibañez, Albillos, Garcia, Carcia-Sancho and Alvarez (2000) Nat. Cell Biol. 2, 57–61]. In the present study, we have studied the effect of stimulation of inositol 1,4,5-trisphosphate (InsP3) receptors using histamine. We find that histamine produces a heterogeneous increase in [Ca2+]M, reaching peak levels at approx. 1μM in 70% of the mitochondrial space to several hundred micromolar in 2–3% of mitochondria. Intermediate levels were found in the rest of the mitochondrial space. Single-cell imaging experiments with aequorin showed that the heterogeneity had both an intercellular and a subcellular origin. Those mitochondria responding to histamine with increases in [Ca2+]M much greater than 1μM (30%) were the same as those that also responded with large increases in [Ca2+]M following stimulation with either high-K+ medium or caffeine. Blocking mitochondrial Ca2+ uptake with protonophores or mitochondrial inhibitors also enhanced catecholamine secretion induced by histamine. These results suggest that some InsP3 receptors tightly co-localize with ryanodine receptors and voltage-dependent Ca2+ channels in defined subplasmalemmal functional units designed to control secretion induced by different stimuli.

scholarly journals Cortical filamentous actin disassembly and scinderin redistribution during chromaffin cell stimulation precede exocytosis, a phenomenon not exhibited by gelsolin.

1991 ◽  
Vol 113 (5) ◽  
pp. 1057-1067 ◽  
Author(s):  
M L Vitale ◽  
A Rodríguez Del Castillo ◽  
L Tchakarov ◽  
J M Trifaró
Keyword(s):  
Actin Filament ◽  
Chromaffin Cells ◽  
Chromaffin Cell ◽  
Catecholamine Secretion ◽  
Cortical Region ◽  
Cortical Surface ◽  
Filamentous Actin ◽  
Cell Stimulation ◽  
Rhodamine Phalloidin ◽  
Filament Networks

Immunofluorescence and cytochemical studies have demonstrated that filamentous actin is mainly localized in the cortical surface of the chromaffin cell. It has been suggested that these actin filament networks act as a barrier to the secretory granules, impeding their contact with the plasma membrane. Stimulation of chromaffin cells produces a disassembly of actin filament networks, implying the removal of the barrier. The presence of gelsolin and scinderin, two Ca(2+)-dependent actin filament severing proteins, in the cortical surface of the chromaffin cells, suggests the possibility that cell stimulation brings about activation of one or more actin filament severing proteins with the consequent disruption of actin networks. Therefore, biochemical studies and fluorescence microscopy experiments with scinderin and gelsolin antibodies and rhodamine-phalloidin, a probe for filamentous actin, were performed in cultured chromaffin cells to study the distribution of scinderin, gelsolin, and filamentous actin during cell stimulation and to correlate the possible changes with catecholamine secretion. Here we report that during nicotinic stimulation or K(+)-evoked depolarization, subcortical scinderin but not gelsolin is redistributed and that this redistribution precedes catecholamine secretion. The rearrangement of scinderin in patches is mediated by nicotinic receptors. Cell stimulation produces similar patterns of distribution of scinderin and filamentous actin. However, after the removal of the stimulus, the recovery of scinderin cortical pattern of distribution is faster than F-actin reassembly, suggesting that scinderin is bound in the cortical region of the cell to a component other than F-actin. We also demonstrate that peripheral actin filament disassembly and subplasmalemmal scinderin redistribution are calcium-dependent events. Moreover, experiments with an antibody against dopamine-beta-hydroxylase suggest that exocytosis sites are preferentially localized to areas of F-actin disassembly.

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