Characterization of inositol 1,4,5-trisphosphate-sensitive (IsCaP) and-insensitive (IisCaP) nonmitochondrial Ca2+ pools in rat pancreatic acinar cells

Abstract

We have measured Ca2+ uptake and Ca2+ release in isolated permeabilized pancreatic acinar cells and in isolated membrane vesicles of endoplasmic reticulum prepared from these cells. Ca2+ uptake into cells was monitored with a Ca2+ electrode, whereas Ca2+ uptake into membrane vesicles was measured with 45Ca2+. Using inhibitors of known action, such as the H+ ATPase inhibitors NBD-Cl and NEM, the Ca2+ ATPase inhibitor vanadate as well as the second messenger inositol 1,4,5-trisphosphate (IP3) and its analog inositol 1,4,5-trisphosphorothioate (IPS3), we could functionally differentiate two nonmitochondrial Ca2+ pools. Ca2+ uptake into the IP3-sensitive Ca2+ pool (IsCaP) occurs by a MgATP-dependent Ca2+ uptake mechanism that exchanges Ca2+ for H+ ions. In the absence of ATP Ca2+ uptake can occur to some extent at the expense of an H+ gradient that is established by a vacuolar-type MgATP-dependent H+ pump present in the same organelle. The other Ca2+ pool takes up Ca2+ by a vanadate-sensitive Ca2+ ATPase and is insensitive to IP3 (IisCaP). The IsCaP is filled at "higher" Ca2+ concentrations (approximately 10(-6) mol/liter) which may occur during stimulation. The low steady-state [Ca2+] of approximately 10(-7) mol/liter is adjusted by the IisCaP. It is speculated that both Ca2+ pools can communicate with each other, the possible mechanism of which, however, is at present unknown.

ATP-driven Ca2+/H+ antiport in acid vesicles from Dictyostelium.

“…It has been proposed that rat parotid (17) and pancreatic (18) (13) have presented evidence that the plasma membrane of Neurospora possesses an ATP-driven Ca2+/H+ exchanger that is stimulated by the inwardly directed proton gradient due to the plasma membrane proton pump and advance a convincing thermodynamic argument that this interaction is important for the maintenance of low cytosolic [Ca2W] in that organism. In Dictyostelium and possibly in other cell types where there is evidence for protonophore-sensitive calcium sequestration (17)(18)(19)(20)(21)(22), if such an enzyme pumps calcium ions into an acidic compartment, then the cell could use the energy stored in the proton gradient to augment that available from ATP (13). The efficiency of calcium sequestration into acidic intracellular stores would then depend on the magnitude of the transmembrane pH gradient and should be reduced by exposure to weak bases such as ammonia.…”

Oscillating intracellular Ca2+ signals evoked by activation of receptors linked to inositol lipid hydrolysis: Mechanism of generation

“…4 requires two separate Ca 2+ pools with Ca 2+ channels having different characteristics. The existence of at least two nonmitochondrial intracellular Ca 2+ pools has been proposed on the basis of studies where it can be shown that Ins (1,4,5) P3 is only able to release Ca 2+ from a minor compartment of the endoplasmic reticulum Ca 2+ pool (Biden, Wollheim & Schlegel, 1986;Thevenod et al, 1989). In the pancreatic acinar cells the two Ca 2 + pools seem to have different types of Ca 2 + uptake mechanisms.…”

“…This idea is based on the data from Osipchuk et al (1990). The proposal that the Ca 2" uptake mechanisms in the Ins (1,4,5) P3-sensitive and insensitive pools are different is based on the data from Thevenod et al (1989) polypeptides with M r -400,000) with square shape (Maeda et al, 1990).…”

Ca2+ dependence of inositol 1,4,5‐trisphosphate‐induced Ca2+ release in renal epithelial LLC‐PK1 cells

“…When cells are loaded a t high free [Ca ' 3, this component accounts for up to 35% of the sequestered Ca2 ' . It is not very likely that the thapsigargin-insensitive Ca2+ pool in LLC-PK, cells is related to the vanadateinsensitive Ca"+ pool described in pancreatic acinar cells (Thevenod et al, 1989). The vanadate-insensitive Ca" pool in the latter study was found to be InsP,-sensitive and was filled by a NEM-sensitive mechanism, which was not the case for the thapsigargin-insensitive Ca2+ pool in our study.…”