ausblenden:
Schlagwörter:
H+ pump; H+ ATPase inhibitors; vanadate; inositol 1,4,5-trisphosphorothioate; Ca2+ electrode; acridine orange
Zusammenfassung:
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 with45Ca2+. 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 non-mitochondrial 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 (∼10−6 mol/liter) which may occur during stimulation. The low steady-state [Ca2+] of ∼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.
