title: Imaging of intracellular calcium stores in individual permeabilized pancreatic acinar cells. Apparent homogeneous cellular distribution of inositol 1,4,5-trisphosphate-sensitive stores in permeabilized pancreatic acinar cells.
abstract: Several lines of evidence suggest that the existence of a heterogeneous population of inositol 1,4,5-trisphosphate (Ins(1,4,5)P3)-sensitive Ca2+ stores underlies the polarized agonist-induced rise in cytosolic Ca2+ concentration ([Ca2+]i) 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 Ca2+ stores which are relatively more sensitive to Ins(1,4,5)P3 than those in basolateral areas, we studied Ca2+ handling by Ca2+ stores in individual streptolysin O (SLO) permeabilized cells using the low affinity Ca2+ indicator Magfura-2 and an in situ imaging technique. The uptake of Ca2+ by intracellular Ca2+ stores was ATP-dependent. A steady-state level was reached within 10 min, and the free Ca2+ concentration inside loaded Ca2+ stores was estimated to be 70 microM. Ins(1,4,5)P3 induced Ca2+ 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)P3 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)P3-sensitive Ca2+ stores are unlikely to be redistributed as a result of SLO treatment. The effects of Ins(1,4,5)P3 were therefore subsequently studied at the subcellular level. Detailed analysis demonstrated that no regional differences in Ins(1,4,5)P3 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 Ca2+ signaling in intact pancreatic acinar cells.
authors.affiliation: School of Biological Sciences, G. 38 Stopford Building, Oxford Road, Manchester M13 9PT, United Kingdom.
authors.forename: F H
authors.initials: FH
authors.lastname: van de Put
authors.forename: A C
authors.initials: AC
authors.lastname: Elliott
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001426:Bacterial Proteins; D002118:Calcium / Q000032:analysis / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002463:Cell Membrane Permeability; D003594:Cytoplasmic Granules / Q000378:metabolism / Q000648:ultrastructure; D003600:Cytosol / Q000378:metabolism; D005456:Fluorescent Dyes; D016257:Fura-2 / Q000031:analogs & derivatives; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D008854:Microscopy, Electron; D010179:Pancreas / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013301:Streptolysins; D013997:Time Factors
chemical_list: D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D013301:Streptolysins; C025964:streptolysin O; C072323:Mag-Fura-2-acetoxymethyl ester; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D002118:Calcium; D016257:Fura-2