Physiological effects of Na+/Ca2+ exchanger knockdown by antisense oligodeoxynucleotides in arterial myocytes.
Antisense oligodeoxynucleotides (AS-oligos) targeted to the Na+/Ca2+ exchanger (NCX) inhibit NCX-mediated Ca2+ influx in mesenteric artery (MA) myocytes [Am. J. Physiol. 269 (Cell Physiol. 38): C1340-C1345, 1995]. Here, we show AS-oligo knockdown of NCX-mediated Ca2+ efflux. In initial experiments, the cytosolic free Ca2+ concentration ([Ca2+]cyt) was raised, and sarcoplasmic reticulum (SR) Ca2+ sequestration was blocked with caffeine and cyclopiazonic acid; the extracellular Na+-dependent (NCX) component of Ca2+ efflux was then selectively inhibited in AS-oligo-treated cells but not in controls (no oligos or nonsense oligos). In contrast, the La3+-sensitive (plasmalemma Ca2+ pump) component of Ca2+ efflux was unaffected in AS-oligo-treated cells. Knockdown of NCX activity was reversed by incubating AS-oligo-treated cells in normal media for 5 days. Transient [Ca2+]cyt elevations evoked by serotonin (5-HT) at 15-min intervals in AS-oligo-treated cells were indistinguishable from those in controls. When cells were stimulated every 3 min, however, the peak amplitudes of the second and third responses were larger, and [Ca2+]cyt returned to baseline more slowly, in AS-oligo-treated cells than in controls. Peak 5-HT-evoked responses in the controls, but not AS-oligo-treated cells, were augmented more than twofold in Na+-free media. This implies that NCX is involved in Na+ gradient modulation of SR Ca2+ stores and cell responsiveness. The repetitive stimulation data suggest that the NCX may be important during tonic activation of arterial myocytes.
Department of Physiology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
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D000818:Animals; D001483:Base Sequence; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002462:Cell Membrane / Q000201:enzymology; D002478:Cells, Cultured; D003470:Culture Media; D003600:Cytosol / Q000378:metabolism; D007211:Indoles / Q000494:pharmacology; D007700:Kinetics; D007811:Lanthanum / Q000494:pharmacology; D008638:Mesenteric Arteries; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D051381:Rats; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D012701:Serotonin / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D019831:Sodium-Calcium Exchanger / Q000096:biosynthesis* / Q000235:genetics
D003470:Culture Media; D007211:Indoles; D016376:Oligonucleotides, Antisense; D019831:Sodium-Calcium Exchanger; D012701:Serotonin; D002110:Caffeine; D007811:Lanthanum; D012964:Sodium; D000252:Calcium-Transporting ATPases; D002118:Calcium; C000543:cyclopiazonic acid