Thapsigargin stimulates Ca2+ entry in vascular smooth muscle cells: nicardipine-sensitive and -insensitive pathways.
We have investigated the role of the sarcoplasmic reticulum Ca2+ pool in regulating Ca2+ entry in vascular smooth muscle cells using a receptor-independent means of mobilizing the intracellular Ca2+ pool. Thapsigargin (TG) has been shown to inhibit the endoplasmic reticulum Ca(2+)-ATPase, mobilize intracellular Ca2+, and activate Ca2+ entry in nonmuscle tissues. When smooth muscle cells were treated with 0.2 microM TG, cytosolic Ca2+ concentrations rose gradually over 8 min to a peak value of 365 +/- 18 nM. Cytosolic Ca2+ remained elevated for at least 20 min and was supported by continued entry of extracellular Ca2+. TG also stimulated entry of Mn2+ and 45Ca2+ from outside the cell. Importantly, TG-induced Ca2+ entry and Mn2+ entry were found to occur through mechanisms that were independent of L-type Ca2+ channel activation because influx was not inhibited by concentrations of nicardipine that were found to block either endothelin- or 100 mM extracellular K(+)-induced cation influx. The mechanism through which TG activates cation entry appears to involve mobilization of the inositol 1,4,5-trisphosphate-responsive intracellular Ca2+ pool. In permeabilized cells, TG prevented ATP-stimulated Ca2+ uptake into the sarcoplasmic reticulum and slowly released sequestered Ca2+. The Ca2+ pool involved was responsive to inositol 1,4,5-trisphosphate. However, TG did not initiate the formation of inositol polyphosphates. Thus TG mobilizes the sarcoplasmic reticulum Ca2+ pool and activates Ca2+ entry through a nicardipine-insensitive Ca2+ channel in vascular smooth muscle. The mechanism is independent of inositol polyphosphate formation.
Xuan
Y T
YT
Department of Pharmacology, Duke University Medical Center, Durham, North Carolina 27710.
Wang
O L
OL
Whorton
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AR
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002478:Cells, Cultured; D007295:Inositol Phosphates / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D008345:Manganese / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D009529:Nicardipine / Q000494:pharmacology*; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D013729:Terpenes / Q000494:pharmacology*; D019284:Thapsigargin
D007295:Inositol Phosphates; D013729:Terpenes; D008345:Manganese; D019284:Thapsigargin; D009529:Nicardipine; D000252:Calcium-Transporting ATPases; D002118:Calcium