Dynamic properties of an inositol 1,4,5-trisphosphate- and thapsigargin-insensitive calcium pool in mammalian cell lines.
The functional characteristics of a nonacidic, inositol 1,4,5-trisphosphate- and thapsigargin-insensitive Ca2+ pool have been characterized in mammalian cells derived from the rat pituitary gland (GH3, GC, and GH3B6), the adrenal tissue (PC12), and mast cells (RBL-1). This Ca2+ pool is released into the cytoplasm by the Ca2+ ionophores ionomycin or A23187 after the discharge of the inositol 1,4,5-trisphosphate-sensitive store with an agonist coupled to phospholipase C activation and/or thapsigargin. The amount of Ca2+ trapped within this pool increased significantly after a prolonged elevation of intracellular Ca2+ concentration elicited by activation of Ca2+ influx. This pool was affected neither by caffeine-ryanodine nor by mitochondrial uncouplers. Probing mitochondrial Ca2+ with recombinant aequorin confirmed that this pool did not coincide with mitochondria, whereas its homogeneous distribution across the cytosol, as revealed by confocal microscopy, and its insensitivity to brefeldin A make localization within the Golgi complex unlikely. A proton gradient as the driving mechanism for Ca2+ uptake was excluded since ionomycin is inefficient in releasing Ca2+ from acidic pools and Ca2+ accumulation/release in/from this store was unaffected by monensin or NH4Cl, drugs known to collapse organelle acidic pH gradients. Ca2+ sequestration inside this pool, thus, may occur through a low-affinity, high-capacity Ca2+-ATPase system, which is, however, distinct from classical endosarcoplasmic reticulum Ca2+-ATPases. The cytological nature and functional role of this Ca2+ storage compartment are discussed.
Pizzo
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P
Department of Biomedical Sciences, University of Padova, Italy.
Fasolato
C
C
Pozzan
T
T
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002451:Cell Compartmentation; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology*; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008928:Mitochondria / Q000378:metabolism; D016716:PC12 Cells; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D012433:Ryanodine / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D014475:Uncoupling Agents / Q000494:pharmacology
D015220:Calcium Channels; D007476:Ionophores; D014475:Uncoupling Agents; D012433:Ryanodine; D000001:Calcimycin; D002110:Caffeine; D015759:Ionomycin; D011189:Potassium Chloride; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000252:Calcium-Transporting ATPases; D002118:Calcium