Polarized Ca2+ release in saponin-permeabilized parotid acinar cells evoked by flash photolysis of 'caged' inositol 1,4,5-trisphosphate.
In exocrine acinar cells, agonist stimulation results in a polarized Ca2+ signal, termed the 'Ca2+ wave', that propagates from the apical pole towards the basolateral region. We attempted to detect the inositol 1,4,5-trisphosphate (InsP3)-induced Ca2+ wave in saponin-permeabilized rat parotid acinar cells using a digital imaging system. The permeabilized acinar cells were labelled with the membrane-bound Ca2+ indicator Calcium Green C18 to detect changes in Ca2+ concentration adjacent to the membrane of intracellular organelles. Application of InsP3 was made by the photolysis of InsP3 P4(5)-1-(2-nitrophenyl)ethyl ester (caged InsP3) to expose simultaneously all regions of the permeabilized acinar cells to InsP3. The increase in fluorescence ratio following the photolysis of 0.5 microM caged InsP3 started at the apical region of the acinar cells within 0.1 s and spread towards the basolateral region, indicating that Ca2+ release from intracellular Ca2+ stores was initially evoked at the apical region. Pretreatment with thapsigargin, an inhibitor of endoplasmic reticulum Ca2+ pumps, failed to prevent the InsP3-induced Ca2+ wave, suggesting that the generation of the Ca2+ wave is not attributed to the polarized distribution of the Ca2+ pumps. The photolysis of a high concentration (10 microM) of caged InsP3 caused a homogeneous increase in the fluorescence ratio throughout the cells, indicating that all regions of intracellular Ca2+ stores similarly responded to the high concentration of InsP3. The present study is the first demonstration of the InsP3-induced Ca2+ wave in permeabilized exocrine acinar cells. The result provides fresh evidence that the apical region contains elements of intracellular Ca2+ stores particularly sensitive to InsP3 and that the Ca2+ wave results from a polarized distribution of InsP3-sensitive Ca2+ stores.
Tanimura
A
A
Department of Dental Pharmacology, School of Dentistry, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Hokkaido 061-02, Japan.
Matsumoto
Y
Y
Tojyo
Y
Y
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000187:drug effects; D005456:Fluorescent Dyes; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D015544:Inositol 1,4,5-Trisphosphate / Q000031:analogs & derivatives / Q000494:pharmacology* / Q000528:radiation effects; D007424:Intracellular Fluid / Q000378:metabolism / Q000528:radiation effects; D007425:Intracellular Membranes / Q000378:metabolism; D008297:Male; D009930:Organic Chemicals; D010306:Parotid Gland / Q000166:cytology / Q000378:metabolism* / Q000528:radiation effects; D010782:Photolysis; D051381:Rats; D017208:Rats, Wistar; D012503:Saponins / Q000494:pharmacology*
D005456:Fluorescent Dyes; D009930:Organic Chemicals; D012503:Saponins; C095636:calcium green C18; C061868:inositol 1,4,5-trisphosphate 1-(2-nitrophenyl)ethyl ester; D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium