Role of the cardiac Na(+)/H(+)exchanger in [Ca(2+)](i)and [Na(+)](i)handling during intracellular acidosis. Effect of cariporide (Hoe 642).
Intracellular acidosis is one of the alterations occurring in cardiac ischemia and has been discussed to be important in altering excitation--contraction coupling. The aim of this study was to determine how intracellular acidosis may affect intracellular sodium and calcium handling. Cardiomyocytes were isolated from the hearts of adult male guinea-pigs by standard techniques and superfused with modified Tyrode's solution at room temperature, either HEPES buffered containing 10 mM NaHCO(3)or HEPES buffered without NaHCO(3), in order to examine a possible interaction with the sodium bicarbonate symport. The whole cell voltage clamp technique was used utilizing 3 M Omega pipettes filled with (mM): Cs aspartate 120, CsCl 20, MgCl(2)1, NaCl 5, Mg-ATP 2, HEPES 10 and either 100 microM Fura-2 or 100 microM SBFI. The pH of the pipette solution was either 7.2 or 6.5. Cells were kept at a holding potential of -80 mV and after a pre-pulse to -40 mV the membrane was continuously clamped to potentials from -30 to +80 mV in 10 mV steps. Intracellular Ca(2+)or Na(+)were estimated using the Fura-2 or SBFI technique (impermeable salt), respectively. The cardiac Na(+)/H(+)exchanger was inhibited using the Na(+)/H(+)- exchange inhibitor cariporide (Hoe 642) (1 microM), when indicated. In NaHCO(3)-free experiments we found an increase in intracellular sodium reflected by a rise in the SBFI ratio of 0.326 +/- 0.01 upon intracellular acidification, in contrast to cells perfused at pH = 7.2 (no significant increase in intracellular Na(+)) (P< 0.05). There was no difference in intracellular calcium handling between cells perfused with solutions of pH = 7.2 or 6.5 (Fura-2 Delta ratio: 0.79 +/- 0.10 vs 0.82 +/- 0.07, n.s.). The l -type calcium current also remained unchanged. Blockade of the Na(+)/H(+)exchanger by Hoe 642 had no influence on cells perfused at pH = 7.2 but inhibited the increase in intracellular Na(+)at pH = 6.5 (0.023 +/- 0.026 in the presence of Hoe 642 vs 0.326 +/- 0.01 without Hoe 642, P< 0.05) without affecting [Ca(2+)](i)or the L-type calcium current. In cells superfused with a Tyrode solution containing NaHCO(3), the increase in intracellular sodium concentration was even more pronounced. Under these conditions Hoe 642 also antagonized this increase in intracellular sodium but without reaching the control level. We conclude that under these experimental conditions intracellular acidification causes an increase in [Na(+)](i)without changing intracellular Ca(2+)or the L-type calcium current. In addition in bicarbonate-buffered systems the acidosis-induced increase in sodium is enhanced which may involve the Na(+)/HCO(3)(minus sign)symport. The effect of cariporide (Hoe 642) in intracellular acidosis seems to be based on antagonization of the rise in intracellular sodium rather than calcium in this model.
Salameh
Aida
A
Medizinische Klinik I, Abt. Kardiologie, Universität Leipzig, Johannisallee 32, 04103 Leipzig, Germany. sala@medizin.uni-leipzig.de
Dhein
Stefan
S
Beuckelmann
Dirk J
DJ
D000818:Animals; D000889:Anti-Arrhythmia Agents / Q000494:pharmacology*; D001572:Benzofurans / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000187:drug effects; D004988:Ethers, Cyclic / Q000494:pharmacology; D016257:Fura-2 / Q000494:pharmacology; D006146:Guanidines / Q000494:pharmacology*; D006168:Guinea Pigs; D006321:Heart / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D012964:Sodium / Q000378:metabolism*; D017693:Sodium Bicarbonate / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000502:physiology*; D013450:Sulfones / Q000494:pharmacology*
D000889:Anti-Arrhythmia Agents; D001572:Benzofurans; D020746:Calcium Channels, L-Type; D004988:Ethers, Cyclic; D006146:Guanidines; D017923:Sodium-Hydrogen Exchangers; D013450:Sulfones; C061665:sodium-binding benzofuran isophthalate; C093373:cariporide; D017693:Sodium Bicarbonate; D012964:Sodium; D002118:Calcium; D016257:Fura-2