Abstract
Although inhibition of the Na+/Ca2+ exchanger normally increases [Ca2+](i) in neonatal cardiac myocytes, application of the inhibitor Ni2+ appears to reduce [Ca2+](i) measured by fluo-3. To investigate how the apparent reduction in [Ca2+](i) occurs we examined Ca2+ transport by the human Na+/Ca2+ exchanger expressed in Sf9 cells. Transport of Ca2+ by the Na+/Ca2+ exchanger was examined using a laser-scanning confocal microscope and the fluorescent Ca2+ indicator fluo-3, and the electrogenic function was determined by measuring the Na+/Ca2+ exchange current (I(NaCa)) using patch clamp methods. I(NaCa) was elicited with voltage-clamp steps or flash photolysis of caged Ca2+. We show significant expression of Na+/Ca2+ exchanger function in Sf9 cells infected with a recombinant Baculovirus carrying the Na+/Ca2+ exchanger. In addition to measurements of I(NaCa), characterization includes Ca2+ transport via the Na+/Ca2+ exchanger and the voltage dependence of Ca2+ transport. Application of Ni2+ blocked I(NaCa) but, contrary to expectation, decreased fluo-3 fluorescence. Experiments with infected Sf9 cells suggested that Ni2+ was transported via the Na+/Ca2+ exchanger at a rate comparable to the Ca2+ transport. Once inside the cells, Ni2+ reduced fluorescence, presumably by quenching fluo-3. We conclude that Ni2+ does indeed block I(NaCa), but is also rapidly translocated across the cell membrane by the Na+/Ca2+ exchanger itself, most likely via an electroneutral partial reaction of the exchange cycle.
Original language | English (US) |
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Pages (from-to) | 9-17 |
Number of pages | 9 |
Journal | Cell Calcium |
Volume | 25 |
Issue number | 1 |
DOIs | |
State | Published - Jan 1999 |
Bibliographical note
Funding Information:The work was supported by grants from the Swiss National Science Foundation to E.N. (31.–50564.97), the NIH (AG15555) and by a grant from the University of Maryland SRIF to D.H.S. We would like to thank Mrs M. Herrenschand and Mrs J. Gygax for skillful technical help.