IP3-mediated cytosolic and nuclear calcium elevation in NRK-52E cells using 'caged' GPIP2.
Alterations in calcium homeostasis play a pivotal role in the cellular response to injury. Increases in the concentration of cytosolic free calcium ([Ca2+]i) result in a variety of calcium mediated toxic responses such as cytoskeletal alterations, mitochondrial damage, and over-expression of gene products. Inositol trisphosphate is a second messenger that links external cell surface signals to [Ca2+]i elevation. The present study explored the use of caged glycerophosphoryl-myo-inositol-1,4,5-bisphosphate (GPIP2) to mediate a rapid and prolonged increase in [Ca2+]i in a normal rat kidney epithelial cell line (NRK-52E). In intact NRK-52E cells, UV photolysis of microinjected GPIP2 resulted in a 3-4-fold sustained increase in [Ca2+]i. Graded photolytic release of GPIP2 also resulted in calcium-mediated morphologic alterations, as shown by confocal microscopy, with cellular blebs apparent within 30 min. There was no apparent increase in [Ca2+]i or morphologic alterations in control cells microinjected with calcium indicator and equally exposed to UV light. Subsequent application of thapsigargin or ionomycin (1.0 microM) produced a rapid and transient increase in [Ca2+]i. In addition, we show that activation of IP3 stores results in increased concentration of ionized nuclear calcium, ([Ca2+]n) which persists longer than the increase in [Ca2+]i. These findings indicate that GPIP2 mediates a rapid and sustained elevation in [Ca2+]n and [Ca2+]i and this IP3-mediated calcium elevation is translated to the nucleus in rat kidney epithelial cells.
Department of Pathology, School of Medicine, University of Maryland, Baltimore, USA.
M A
MA
Davis
S H
SH
Chang
B F
BF
Trump
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002467:Cell Nucleus / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D003600:Cytosol / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D004847:Epithelial Cells; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism* / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism* / Q000494:pharmacology; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D010782:Photolysis; D051381:Rats
C087840:1-(glycerophosphoryl)-myo-inositol 4,5-diphosphate P(4(5))-1-(2-nitrophenyl)ethyl ester; D007295:Inositol Phosphates; D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium