Sodium-calcium exchange in cultured bovine pulmonary artery endothelial cells.
1. Intracellular free calcium ([Ca2+]i) was measured in cultured bovine pulmonary artery endothelial cell monolayers loaded with the fluorescent calcium indicator Fura-2. 2. Resting [Ca2+]i was 112 +/- 10 nM. Application of ouabain (20 microM) was without effect on [Ca2+]i for periods of up to 1 h. Monensin (10 microM) resting [Ca2+]i to 145 +/- 32 nM over approximately 2 min. In the presence of ouabain (20 microM), 10 microM-monensin increased [Ca2+]i to 146 +/- 15 nM. 3. Removal of extracellular sodium was without effect in resting cells or cells exposed to ouabain alone. However, in the presence of monensin, replacement of extracellular Na+ with Li+ resulted in a prompt increase in [Ca2+]i to a peak of 280 +/- 37 nM, which then returned towards resting levels. When Na+ was removed in the presence of both ouabain and monensin, [Ca2+]i reached a peak of 585 +/- 53 nM. 4. When extracellular Na+ was replaced with K+, to achieve simultaneous Na+ removal and depolarization, [Ca2+]i reached a peak of 568 +/- 63 nM, compared with a peak of 462 +/- 38 nM when Li+ was used as a Na+ substitute in paired experiments. The transient increase in [Ca2+]i evoked by sodium removal peaked earlier when K+ was used as the sodium substitute, showing that depolarization increased the rate of calcium influx into the cell when sodium was removed from the bathing medium. 5. Removal of extracellular K+ had no effect on the low-Na(+)-evoked increase in [Ca2+]i. 6. Returning extracellular Na+ during the increase in [Ca2+]i resulting from Na+ removal increased the rate of return of [Ca2+]i towards basal levels. In the absence of Na+, [Ca2+]i took 41 +/- 5 s to decline from 400 to 200 nM, and this was reduced to 26 +/- 6 s (n = 4, S.E.M.) when Na+ was returned to the bathing solution. 7. These results indicate endothelial cells possess a voltage-dependent Na(+) -Ca2+ exchange mechanism in the surface membrane. However, this mechanism does not appear to be of primary importance in the maintenance of resting [Ca2+]i since cells were able to restore a low [Ca2+]i in the absence of extracellular Na+. The evidence for the existence of a Na(+) -Ca2+ exchanger in the surface membrane of endothelial cells and the possibility that this mechanism may contribute to calcium entry and/or extrusion during agonist-evoked responses is discussed.
Sage
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SO
Department of Pharmacology, University of Miami School of Medicine, FL 33136.
van Breemen
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Cannell
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D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D006706:Homeostasis / Q000502:physiology; D066298:In Vitro Techniques; D007474:Ion Exchange; D008985:Monensin / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D011651:Pulmonary Artery / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*
D010042:Ouabain; D008985:Monensin; D012964:Sodium; D002118:Calcium