Imaging of Intracellular Calcium Stores in Individual Permeabilized Pancreatic Acinar Cells

Abstract

2؉ release in a dose-dependent, "quantal" fashion. The kinetics of this release were similar to those reported for suspensions of permeabilized pancreatic acinar cells. Interestingly, the permeabilized acinar cells showed no intercellular variation in Ins(1,4,5)P 3 sensitivity. Although SLO treatment is known to result in a considerable loss of cytosolic factors, permeabilization did not result in a redistribution of zymogen granules, as judged by electron microscope analysis. These results suggest that Ins(1,4,5)P 3 -sensitive Ca 2؉ stores are unlikely to be redistributed as a result of SLO treatment. The effects of Ins(1,4,5)P 3 were therefore subsequently studied at the subcellular level. Detailed analysis demonstrated that no regional differences in Ins(1,4,5)P 3 sensitivity exist in this permeabilized cell system. Therefore, we propose that additional cytosolic factors and/or the involvement of ryanodine receptors underlie the polarized pattern of agonist-induced Ca 2؉ signaling in intact pancreatic acinar cells.

Slow kinetics of inositol 1,4,5-trisphosphate-induced Ca2+ release: is the release ‘quantal’ or ‘non-quantal’?

“…The slow phase of the InsP $ -induced Ca# + release in pancreatic acinar cells only occurred in the absence of Ca# + pumping and was prevented when the Ca# + pumps were functional [18,33]. A similar phenomenon was observed for the Ca# + release via the ryanodine receptor in smooth muscle [34].…”

“…Effects of InsP $ are often presented as linear plots of the total store Ca# + content, the luminal [Ca# + ] or the medium free [Ca# + ] against time [2,17,19,31,[37][38][39]. In addition the signals coming from luminally trapped Ca# + indicators are usually presented as linear plots of fluorescence intensities [16,17] or ratios [18,21] against time. We have therefore replotted the content of the Ca# + stores on a linear scale (Figure 1B).…”

“…The concept of quantal or partial Ca# + release was recently challenged when imaging techniques made it possible to measure directly the effects of InsP $ on the luminal [Ca# + ]. An almost total (nonquantal) release of the stores by a submaximal [InsP $ ] was reported when the luminal [Ca# + ] was monitored with chlortetracycline [16], Calcium Green-5N [17] or with Magfura 2 in the presence of thapsigargin [18]. However, other experiments with fura 2 [17,19], furaptra [20] or Magfura 2 [21] resulted in a quantal or partial release pattern.…”

Newly Developed Mg2+–Selective Fluorescent Probe Enables Visualization of Mg2+ Dynamics in Mitochondria

“…Because of the absence of Mg 2+ selective indicators, however, the changes in [Mg 2+ ] i in cells and organelles are still not well understood. Commercially available fluorescent Mg 2+ probes are also sensitive to changes in the intracellular Ca 2+ concentration, and therefore, those probes are often used as Ca 2+ indicators [10], [11], [12]. In our group, novel Mg 2+ selective fluorescent probes, the KMG series, have been developed [13], [14].…”

Phasic characteristic of elementary Ca2+release sites underlies quantal responses to IP3

“…In particular, the graded nature of Ca 2+ release is difficult to reconcile with the well‐established stimulatory effect of cytosolic Ca 2+ on IP 3 receptor (IP 3 R) activity (Iino, 1990; Bezprozvanny et al ., 1991; Finch et al ., 1991), which would be expected to lead to regenerative, all‐or‐none responses through Ca 2+ ‐induced Ca 2+ release (CICR). Two major models have been proposed to resolve this discrepancy: (i) all‐or‐none release from stores exhibiting heterogeneous sensitivities to IP 3 , as discussed above (Muallem et al ., 1989; Bootman et al ., 1992; Cheek et al ., 1994; Beecroft and Taylor, 1997); and (ii) a ‘steady‐state’ model, involving intracellular stores with similar sensitivities to IP 3 that each release Ca 2+ in a phasic or adaptive manner in response to stepwise increments of [IP 3 ] (Irvine, 1990; Missiaen et al ., 1991, 1992, 1999; Nunn and Taylor, 1992; Tanimura and Turner, 1996; van de Put and Elliott, 1996; Koizumi et al ., 1999).…”