Calcium entry-dependent oscillations of cytoplasmic calcium concentration in cultured endothelial cell monolayers.
Bovine endothelial cell monolayers grown to confluence and stimulated with bradykinin responded with periodic fluctuations in intracellular Ca2+ concentration ([Ca2+]i) when exposed to K(+)-free Hepes-buffered saline. The fluctuations in [Ca2+]i measured with fura-2 were synchronized among the population of cells observed and were sensitive to extracellular Ca2+ concentration ([Ca2+]o). Thapsigargin, which inhibits the endoplasmic reticular Ca2(+)-ATPase, did not inhibit the [Ca2+]i oscillations. Removal of extracellular Ca2+ or inhibition of Ca2+ entry by using La3+ or 1-(beta- [3-(4-methoxyphenyl)proproxy]-4-methoxyphenethyl)-1H-imidazole hydrochloride (SKF 96365) abolished the [Ca2+]i oscillations in endothelial cell monolayers. The fluctuations in [Ca2+]i were therefore dependent on Ca2+ influx rather than Ca2+ mobilization from intracellular stores. Simultaneous measurements of membrane potential (Em) using the potential-sensitive bisoxonol dye bis(1,3-dibutylbarbituric acid)trimethine oxonol [Di-BAC4(3)] and [Ca2+]i using fura-2 showed that Em oscillated at the same frequency as the fluctuations in [Ca2+]i. The peak depolarization signal coincided with the maximum rate of increase in the [Ca2+]i signal. Oscillations in the Em signal were inhibited by removal of Ca2+ or by addition of 1 mM Ni2+ to the external solution. Taken together, these observations suggest that the change in Em is the consequence of oscillatory changes in a membrane conductance that also allows Ca2+ to enter the cell. Oscillations in the DiBAC4(3) signal may reflect a rhythmic entry of Ca2+ through nonselective cation channels.
Laskey
R E
RE
Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine, FL 33101.
Adams
D J
DJ
Cannell
M
M
van Breemen
C
C
D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D066298:In Vitro Techniques; D008564:Membrane Potentials; D010507:Periodicity; D011188:Potassium / Q000502:physiology
D011188:Potassium; D001920:Bradykinin; D002118:Calcium