ISSN:
0021-9541
Keywords:
Life and Medical Sciences
;
Cell & Developmental Biology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Medicine
Notes:
We have studied arginine vasopressin (AVP)-, thapsigargin- and inositol 1,4,5-trisphosphate (InsP3)-mediated Ca2+ release in renal epithelial LLC-PK1 cells. AVP-induced changes in the intracellular free calcium concentration ([Ca2+]i) were studied in indo-1 loaded single cells by confocal laser cytometry. AVP-mediated Ca2+ mobilization was also observed in the absence of extracellular Ca2+, but was completely abolished after depletion of the intracellular Ca2+ stores by 2 μM thapsigargin. Using 45Ca2+ fluxes in saponin-permeabilized cell monolayers, we have analysed how InsP3 affected the Ca2+ content of nonmitochondrial Ca2+ pools in different loading and release conditions. Less than 10% of the Ca2+ was taken up in a thapsigargin-insensitive pool when loading was performed in a medium containing 0.1 μM Ca2+. The thapsigargin-insensitive compartment amounted to 35% in the presence of 110 μM Ca2+, but Ca2+ sequestered in this pool could not be released by InsP3. The thapsigargin-sensitive Ca2+ pool, in contrast, was nearly completely InsP3 sensitive. A submaximal [InsP3], however, released only a fraction of the sequestered Ca2+. This fraction was dependent on the cytosolic as well as on the luminal [Ca2+]. The cytosolic free [Ca2+] affected the InsP3-induced Ca2+ release in a biphasic way. Maximal sensitivity toward InsP3 was found at a free cytosolic [Ca2+] between 0.1 and 0.5 μM, whereas higher cytosolic [Ca2+] decreased the InsP3 sensitivity. Other divalent cations or La3+ did not provoke similar inhibitory effects on InsP3-induced Ca2+ release. The luminal free [Ca2+] was manipulated by varying the time of incubation of Ca2+ -loaded cells in an EGTA-containing medium. Reduction of the Ca2+ content to one-third of its initial value resulted in a fivefold decrease in the InsP3 sensitivity of the Ca2+ release. © 1993 Wiley-Liss, Inc.
Additional Material:
6 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/jcp.1041550113
