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Probing differential subcellular calcium dynamics using targeted fluorescence calcium biosensors in rat pulmonary artery smooth muscle cells
Author(s) -
Subedi Krishna Prasad,
Paudel Omkar,
Sham James S.K.
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.923.18
Subject(s) - nucleoplasm , cytoplasm , subcellular localization , nucleus , microbiology and biotechnology , calcium , biology , chemistry , biophysics , nucleolus , organic chemistry
Intracellular calcium (Ca 2+ ) is involved in distinct cellular functions through global and local signaling in various subcellular compartments. To get better insight if the Ca 2+ signals are independently regulated in subcellular compartments, we targeted genetically encoded Ca 2+ indicator, D3cpv, into three different subcellular regions and used laser scanning confocal microscopy to study how Ca 2+ signals are triggered in the plasma membrane (PM), cytoplasm and nucleus in cultured rat pulmonary artery smooth muscle cells (PASMCs). The in situ K d for Ca 2+ of D3cpv was comparable in the cytoplasm and nucleoplasm, but it was slightly higher in the PM. Application of IP 3 or Endothelin‐1 to PASMCs caused rapid increase of fluorescence emission ratio in all three regions. The increase of cytoplasmic/PM Ca 2+ preceded the nucleoplasmic Ca 2+ and the response was more sustained in the nucleus. Platelet derived growth factor (PDGF) generated variable Ca 2+ signals with Ca 2+ increase occurred first in the cytoplasm followed by the nucleoplasm in some cells, while an early Ca 2+ response in the nucleus was observed in the others. Similarly, the increase in PM Ca 2+ was slower than that in the nucleoplasm in some PDGF treated cells. These data suggest a differential regulation of Ca 2+ in the cellular compartments that may be essential for the independent regulation of contraction, cell signaling and gene expression in PASMCs.