Na (+) -Ca2+ exchange in intact endothelium of rabbit cardiac valve
Li, L.; van Breemen, C.
Circulation Research 76(3): 396-404
1995
ISSN/ISBN: 0009-7330 PMID: 7859386 Document Number: 445707
A new method of measuring cytoplasmic free Ca-2+ ((Ca-2+)-i) of individual intact cardiovascular endothelial cells by using imaging fluorescence microscopy was designed. Application of agonist to the aortic or pulmonary valve of the rabbit triggered an increase in (Ca-2+)-i, which depended on the existence of endothelium on the surface of the valve. Under resting conditions, sudden reversal of the Na+ gradient by substituting external Na+ with N-methyl D-glucamine (NMDG) resulted in a (Ca-2+)-i spike, which then returned toward the resting level. Increasing intracellular Na+ concentration ((Na+)-i) by application of ouabain or monensin induced a sustained (Ca-2+)-i increase. Na+ substitution by NMDG during the agonist- or monensin-induced (Ca-2+)-i increase gave rise to a further (Ca-2+)-i spike, which subsequently declined to a level higher than that before removal of external Na+. A selective inhibitor of Na+-Ca-2+ exchange, 3',4'-dichlorobenzamyl (DCB), abolished the transient (Ca-2+)-i increase induced by Na+ substitution, and Mg-2+, an inorganic inhibitor of Na+-Ca-2+ exchanger, markedly reduced this transient (Ca-2+)-i increase. On the other hand, the selective Na+-H+ exchanger blocker 5-(N,N-hexamethylene)amiloride (HMA) did not abolish the transient (Ca-2+)-i increase caused by Na+ substitution. In summary, decreasing the Na+ gradient of the endothelial cells through either receptor stimulation (agonist), Na+-K+ pump inhibition (ouabain), pretreatment with Na+ ionophore (monensin), or reversing the Na+ gradient through Na+ substitution (NMDG) all increased (Ca-2+)-i. This raised (Ca-2+)-i was antagonized by agents such as DCB or Mg-2+, which are thought to inhibit Na+-Ca-2+ exchange, but not by HMA, an inhibitor of Na+-H+ exchange. Taken together, these results strongly imply the presence of Na+-Ca-2+ exchange as a viable mechanism for Ca-2+ transport in intact cardiovascular endothelium and that the Ca-2+ entry component is enhanced when (Na+)-i is elevated.