Calcium regulation in aortic smooth muscle cells during the initial phase of tunicamycin-induced endo/sarcoplasmic reticulum stress.
Endo/sarcoplasmic reticulum stress and the unfolded protein response have been implicated as underlying mechanisms of cell death in many pathological conditions. We have confirmed that long-term exposure to 10µM tunicamycin induced the endo/sarcoplasmic reticulum stress in cultured vascular smooth muscle cells. Since tunicamycin is reported to induce the stress response by inhibiting protein glycosylation, we attempted to investigate a causal link between accumulation of unfolded proteins and dysregulation of cellular calcium transport. However, we found that tunicamycin caused an immediate release of calcium from the endo/sarcoplasmic reticulum, which was sensitive to thapsigargin, and an influx of calcium through the plasma membrane, resulting in a significant increase in cytoplasmic calcium and depletion of endo/sarcoplasmic reticulum calcium. Furthermore, we observed that tunicamycin also induced contraction in intact vascular smooth muscle. By applying established procedures and antagonists, we established that tunicamycin did not directly activate physiological calcium channels, such as store-operated channels, voltage gated calcium channels, ryanodine receptors or inositol trisphosphate receptors. Instead, we found that its effects on cellular calcium fluxes closely resembled those of the known calcium ionophore, ionomycin. We have concluded that tunicamycin directly permeabilizes the plasma membrane and endo/sarcoplasmic reticulum to calcium, and is, therefore, inappropriate for studying the relationship between accumulation of unfolded proteins and endo/sarcoplasmic reticulum calcium dysregulation during the endo/sarcoplasmic reticulum stress response. In contrast, we also report that two other well-known endo/sarcoplasmic reticulum stress inducers, brefeldin A and dithiothreitol, did not exhibit similar increases in calcium permeability.
Child & Family Research Institute, Department of Anesthesiology, Pharmacology, and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada.
Gabriela
G
Ziomek
Child & Family Research Institute, Department of Anesthesiology, Pharmacology, and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada.
Parisa
P
Cheraghi Zanjani
Child & Family Research Institute, Department of Anesthesiology, Pharmacology, and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada.
Darian
D
Arman
Child & Family Research Institute, Department of Anesthesiology, Pharmacology, and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada. Electronic address: casey@ti.ubc.ca.
Cornelis
C
van Breemen
Child & Family Research Institute, Department of Anesthesiology, Pharmacology, and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada; Department of Biomedical Sciences, College of Health Sciences, Midwestern University, Glendale, AZ, USA. Electronic address: mesfan@midwestern.edu.
Mitra
M
Esfandiarei
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001013:Aorta, Thoracic / Q000166:cytology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D059865:Endoplasmic Reticulum Stress / Q000502:physiology*; D008297:Male; D008810:Mice, Inbred C57BL; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism; D011921:Rats, Inbred WKY; D014415:Tunicamycin / Q000494:pharmacology*; D056811:Unfolded Protein Response
D000900:Anti-Bacterial Agents; D014415:Tunicamycin; D002118:Calcium
Calcium; Endoplasmic reticulum stress; Tunicamycin; Unfolded protein response; Vascular smooth muscle cell