Calcium dynamics in bovine adrenal medulla chromaffin cell secretory granules

Abstract

The secretory granules constitute one of the less well-known compartments in terms of Ca 2+ dynamics. ] SG ), the mechanisms for Ca 2+ uptake and release from the granules and their physiological significance regarding exocytosis are still matters of debate. We used in the present work an aequorin chimera targeted to the granules to investigate [Ca 2+ ] SG homeostasis in bovine adrenal chromaffin cells. We found that most of the intracellular aequorin chimera is present in a compartment with 50-100 lm Ca 2+ . Ca 2+ accumulation into this compartment takes place mainly through an ATP-dependent mechanism, namely, a thapsigargin-sensitive Ca 2+ -ATPase. In addition, fast Ca during stimulation. In conclusion, our data show that chromaffin cell secretory granules have the machinery required for rapid uptake and release of Ca 2+ , and this strongly supports the hypothesis that granular Ca 2+ may contribute to its own secretion.

Calcium dynamics in the secretory granules of neuroendocrine cells

“…Similarly, loading with Ca 2+ of dense-core insulin storage granules of mouse pancreatic ␤-cells was sensitive to both SERCA inhibitors BHQ and thapsigargin [42]. More recently, using aequorin to monitor intragranular [Ca 2+ ] we have also described both in chromaffin cells and in the cell lines PC12 and INS1 [23,24], that Ca 2+ uptake by the secretory granules was ATP-dependent and fully sensitive to thapsigargin and BHQ (see Fig. 1).…”

“…Because aequorin is consumed faster in high-Ca 2+ environments, high-Ca 2+ -affinity aequorin is rapidly consumed in those regions and finally tends to reflect only the behavior of minor compartments with low [Ca 2+ ] [22]. Using an aequorin with lower Ca 2+ -affinity and correcting for the effect of acidic pH on aequorin luminescence, we have reported free [Ca 2+ ] values of 50-100 M in chromaffin granules [23] and 20-40 M in PC12 and INS1 cells [24]. We should note also that the small size of some of the vesi- In conclusion, the free [Ca 2+ ] in several types of secretory granules appears to range between 20 and 100 M. These values are significantly lower, nearly by one order of magnitude, than the free [Ca 2+ ] in the endoplasmic reticulum, which is about 500 M [3,25].…”

“…Immunocytochemical localization of all three types of InsP 3 R in serially sectioned bovine chromaffin cells showed that the secretory granules contained about 60% of the InsP 3 R in the cell [50], and additional evidence for the presence of InsP 3 R in the granules was the finding that both chromogranin A and B interact with InsP 3 R and modulate its activity [26,[51][52][53][54]. More recently, using targeted aequorin to measure [Ca 2+ ] in the secretory granules of chromaffin cells and the neurosecretory cell lines PC12 and INS1 [23,24], we have shown that InsP 3 and InsP 3 -releasing agonists can release Ca 2+ from the granular compartment (see Fig. 2).…”

“…The ryanodine receptor activator cyclic ADP ribose (cADPR) has been reported to release Ca 2+ from zymogen granules [55,69]. Both cADPR and caffeine released also Ca 2+ from the secretory granules of insulin-secreting MIN6 cells [20], and caffeine, but not cADPR, released Ca 2+ from secretory granules in chromaffin cells [23]. The lack of effect of cADPR may reflect an insensitivity to this agonist of the ryanodine receptors in these cells, as cADPR was also unable to release Ca 2+ from the ER in chromaffin cells [7].…”

Expression and reconstitution of the bioluminescent Ca2+ reporter aequorin in human embryonic stem cells, and exploration of the presence of functional IP3 and ryanodine receptors during the early stages of their differentiation into cardiomyocytes

“…Viral transduction methods have been previously used to genetically express apo-aequorin in the cytosol, or targeted to distinct intracellular organelles, in a variety of cell types, including HeLa cells, adrenal medulla chromaffin cells, HEK293 cells, GH 3 pituitary cells and PC12 cells (Chamero et al, 2002;Missiaen et al, 2004;SantoDomingo et al, 2008). In addition, apo-aequorin has previously been expressed genetically in cardiomyocytes isolated from adult rats (Bell et al, 2006), but this is the first time that the protein has been expressed in hESCs and in hESC-CMs.…”