Imaging of Intracellular Calcium Stores in Individual Permeabilized Pancreatic Acinar Cells

• Published in: The Journal of biological chemistry 1996-03, Vol.271 (9), p.4999-5006
• Main Authors: van de Put, Frans H.M.M., Elliott, Austin C.
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.9.4999

Several lines of evidence suggest that the existence of a heterogeneous population of inositol 1,4,5-trisphosphate (Ins(1,4,5)P )-sensitive Ca stores underlies the polarized agonist-induced rise in cytosolic Ca concentration ([Ca ] ) in pancreatic acinar cells (Kasai, H., Li, Y. X., and Miyashita, Y.(1993) Cell 74, 669-677; Thorn, P., Lawrie, A. M., Smith, P. M., Gallacher, D. V., and Petersen, O. H.(1993) Cell 74, 661-668). To investigate whether the apical pole of acinar cells contains Ca stores which are relatively more sensitive to Ins(1,4,5)P than those in basolateral areas, we studied Ca handling by Ca stores in individual streptolysin O (SLO) permeabilized cells using the low affinity Ca indicator Magfura-2 and an in situ imaging technique. The uptake of Ca by intracellular Ca stores was ATP-dependent. A steady-state level was reached within 10 min, and the free Ca concentration inside loaded Ca stores was estimated to be 70 μM. Ins(1,4,5)P induced Ca 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 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 -sensitive Ca stores are unlikely to be redistributed as a result of SLO treatment. The effects of Ins(1,4,5)P were therefore subsequently studied at the subcellular level. Detailed analysis demonstrated that no regional differences in Ins(1,4,5)P 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 signaling in intact pancreatic acinar cells.