The lysosomal compartment as intracellular calcium store in MDCK cells: a possible involvement in InsP3-mediated Ca2+ release.
To test for a possible role of lysosomes in intracellular Ca2+ homeostasis, the effects of glycyl-L-phenylalanine-beta-naphthylamide (GPN), known to permeabilize these organelles by osmotic swelling, were studied in single MDCK cells. Fluorescence of acridine orange, rhodol green dextran, lysotracker green and FITC-dextran indicated that GPN (0.2 mmol/l) elicited a reversible permeabilization of lysosomes. Cytosolic Ca2+ ([Ca2+]i) as determined by Fura-2 fluorescence increased from 60 +/- 11 to 534 +/- 66 nmol/l (n = 41) in the presence of GPN. Whereas only a single intracellular Ca2+ release could be induced by GPN in a Ca(2+)-free perfusate, repetitive release could be evoked in Ca2+ containing solutions suggesting reuptake of Ca2+ into lysosomal stores. GPN-induced Ca2+ release was blunted after pretreatment with thapsigargin (TG), an inhibitor of Ca(2+)-ATPase, or repeated applications of ATP inducing Ca2+ release from inositol trisphosphate (InsP3) sensitive Ca2+ stores. The effect of ATP on Ca2+ release was, however, not abolished by preceding GPN treatment. GPN-induced Ca2+ release from lysosomes was independent of InsP3 formation or Ca(2+)-induced Ca2+ release, since it was unaffected by the phospholipase C inhibitor U-73, 122 or by caffeine and ruthenium red. These results suggest that Ca2+ largely accumulates in lysosomal vesicles. Moreover, these organelles seem to be part or functionally coupled with InsP3-sensitive Ca2+ stores.
Haller
T
T
Department of Physiology, University of Innsbruck, Austria.
Dietl
P
P
Deetjen
P
P
Völkl
H
H
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000641:Ammonia / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002451:Cell Compartmentation / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000187:drug effects; D004151:Dipeptides / Q000494:pharmacology; D004285:Dogs; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D007668:Kidney / Q000166:cytology; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin
D004151:Dipeptides; D004791:Enzyme Inhibitors; D013729:Terpenes; C041865:glycylphenylalanine 2-naphthylamide; D019284:Thapsigargin; D000641:Ammonia; D015544:Inositol 1,4,5-Trisphosphate; D000255:Adenosine Triphosphate; D002118:Calcium