On the role of intravesicular calcium in the motion and exocytosis of secretory organelles

Abstract

Secretory vesicles of sympathetic neurons and chromaffin granules maintain a pH gradient towards the cytosol (5.5 vs. 7.2) promoted by the V-ATPase activity. This gradient of pH is mainly responsible for the accumulation of amines. The secretory vesicles contain large amounts of total Ca(2+), but the free intragranular [Ca(2+)], the mechanisms for Ca(2+) uptake and release from the granules and their physiological relevance regarding exocytosis are still matters of debate.We have recently shown that disruption of the pH gradient of secretory vesicles slowed down exocytosis. Fluorimetric measurements, using the dye Oregon green BAPTA-2, showed that the V-ATPase inhibitor bafilomycin A1 directly released Ca(2+) from freshly isolated vesicles. Accordingly, vesicle alkalinization released Ca(2+) from the granules to the cytosol, measured with fura-2 in intact chromaffin cells. Using TIRFM in cells overexpressing the EGFP-labeled synaptobrevin (VAMP2-EGFP) protein, we have then shown that the Ca(2+) released from the vesicles to the cytosol in the presence of bafilomycin, dramatically increased the granule motion of chromaffin- or PC12-derived granules, and triggered exocytosis (measured by amperometry).We conclude that the gradient of pH of secretory vesicles might be involved in the homeostatic regulation of the local cytosolic Ca(2+) around the vesicles and in two of the major functions of secretory cells, vesicle motion and exocytosis.1.

Theoretical Model of Neurotransmitter Release during In Vivo Vesicular Exocytosis Based on a Grainy Biphasic Nano-Structuration of Chromogranins within Dense Core Matrixes

“…61 necessarily leads to an increase of the volume of the lessbound areas through swelling within the present view of dense core vesicle structure. Based on Tanaka and Fillmore's swelling mechanism in Equations 1 and 3, 35 the external areas of the initial matrixes must swell faster than the zones closer to their centers where swelling is necessarily delayed kinetically by the increasingly stronger shear forces.…”

“…cium channel pumps. [61][62][63][64] Though the relative amount of Ca 2+ vs Cat + is small, it is noted that calcium ions have a high affinity for hydroxyl groups. Besides the effect of Ca 2+ dications per se (see above) they may chelate up to 3-6 Cat + per Ca 2+ .…”

“…60 In this respect, it is worth recalling that in addition to a high concentration of catecholamine cations, vesicle matrixes contain also non-negligible concentrations of strongly chelated calcium ions (ca. 40 mM) 61 whose long term permanence inside vesicles is maintained by specific cal- When the cation charge 'm+' increases the radius R blob of the compacted "blobs" increases while the length L strand of the near unfolded domains between blobs decreases (see text). For simplicity only one free chain is represented.…”

Calcium dynamics in the secretory granules of neuroendocrine cells

“…Given that Ca 2+ is the key signal that triggers secretion of granule contents, the possible functional role of the calcium stored in the granules has attracted considerable attention for more than thirty years. The available evidence suggests moreover that intragranular Ca 2+ is important for secretion [37,[73][74][75]. However, in spite of the efforts of many research groups, most of the key questions related with granule Ca 2+ homeostasis, e.g., the free granular [Ca 2+ ], the physical state of the stored Ca 2+ or the mechanisms for Ca 2+ accumulation and release from the granules, remain obscure.…”

Cytosolic organelles shape calcium signals and exo–endocytotic responses of chromaffin cells

“…Experimental protocols and techniques capable of distinguishing the various Ca 2+ sources contributing to such HCMDs may enlighten the contribution of vesicular Ca 2+ release in the various steps of exocytosis. For further analysis of this topic, see two recent reviews [74,75].…”