Ca2+ dependence of inositol 1,4,5-trisphosphate-induced Ca2+ release in renal epithelial LLC-PK1 cells

✍ Scribed by M. Tshipamba; H. De Smedt; L. Missiaen; B. Himpens; L. Van Den Bosch; R. Borghgraef

• Publisher: John Wiley and Sons
• Year: 1993
• Tongue: English
• Weight: 901 KB
• Volume: 155
• Category: Article
• ISSN: 0021-9541
• DOI: 10.1002/jcp.1041550113

✦ Synopsis

We have studied arginine vasopressin (AVP)-, thapsigarginand inositol 1,4,5trisphosphate (InsPJ-mediated CaL+ release in renal epithelial LLC-PK, cells. AVP-induced changes in the intracellular free calcium concentration ([Ca"],) were studied in indo-1 loaded single cells by confocal laser cytometry. AVPmediated Ca2+ mobilization was also observed in the absence of extracellular Ca2+, but was completely abolished after depletion of the intracellular Ca'+ stores by 2 p M thapsigargin. Using 4sCa'+ fluxes in saponin-permeabilized cell monolayers, we have analysed how InsP, affected the Ca'+ content of non-mitochondria1 Ca2+ pools in different loading and release conditions. Less than 10% of the Ca'+ was taken up in a thapsigargin-insensitive pool when loading was performed in a medium containing 0.1 IJ.M Caz+. The thapsigargin-insensitive compartment amounted to 35% in the presence of 110 pM Ca2+, but Ca2+ sequestered in this pool could not be released by InsP,. The thapsigargin-sensitive Ca'+ pool, in contrast, was nearly completely InsP, sensitive. A submaximal [InsP,], however, released only a fraction of the sequestered CaL+. This fraction was dependent on the cytosolic as well as on the luminal [CaH]. The cytosolic free [Ca2+] affected the InsP,-induced Ca2+ release in a biphasic way. Maximal sensitivity toward InsP, was found at a free cytosolic [Ca"] between 0.1 and 0.5 pM, whereas higher cytosolic [Ca2+l decreased the InsP, sensitivity. Other divalent cations or La'+ did not provoke similar inhibitory effects on InsP,-induced Ca2+ release. The luminal free [Ca2+] was manipulated by varying the time of incubation of Ca2+-loaded cells in an ECTA-containing medium. Reduction of the Ca2+ content to one-third of its initial value resulted in a fivefold decrease in the InsP, sensitivity of the Ca'+ release. o 1993 WiIcy-Liss, Inc.