257Large secretory vesicles are still accumulating CA even in the absence of Cgs.
Although amine concentration resulted halved (400–500 mM), it is still hyper-
tonic to cytosol. As the only intravesicular species present inside LDCV is ATP
and considering that ATP is present almost in all secretory vesicles from all ani-
mal cells (Borges 2013 ), we decided to test the role of ATP as a colligative agent.
This ability of ATP to bind CA to reduce the osmotic pressure was demonstrated
in vitro (CITA). We were successful showing that reduction of vesicular ATP,
assessed by siVNUT treatment, was accompanied with a reduction in vesicular
ATP. In addition, the overexpression of VNUT resulted in an increase of vesicular
CA (Estevez-Herrera et al. 2016a). This later results offers a new view of the
physic-chemical interaction between the solutes present in the vesicular cocktail
(Estevez-Herrera et al. 2016b).
3 Concluding Remarks
Since their discovery, Cgs have captivated the attention of scientists and they have
been implicated in several processes, including granule biogenesis and sorting, cat-
echolamine storage and release, as prohormones for the production of bioactive
peptides, tumour marking, and the pathophysiology of neurodegenerative diseases.
Data from Cg-KO mice are providing direct evidence implicating Cgs in vesicular
storage and in the exocytotic release of catecholamines. While the exocytosis pro-
cess is maintained, even in the complete absence of Cgs, the absence of Cgs impairs
vesicular catecholamine accumulation. Hence, although LDCV biogenesis does not
appear to be affected, the saturation of vesicular storage capacity might well be.
Protein analysis of the secretory vesicle fraction revealed the compensatory overex-
pression of CgA in the absence of CgB, and vice versa. Unexpectedly, other pro-
teins that are apparently unrelated with secretion were only present in the
adrenomedullary tissue of CgB-KO animals. In conclusion, in accordance with ear-
lier hypotheses and findings, Cgs are highly efficient direct mediators of mono-
amine accumulation, influencing the kinetics of exocytosis in LDCVs.
Acknowledgments LC is the recipient of a fellowship from the Fundación CajaCanarias. This
work is partially funded by the grant BFU2013-45253-P from the MINECO (Spain) to RB and
JDM.
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Chromogranins and Exocytosis