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.
Slodzinski
M K
MK
Department of Physiology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
Blaustein
M P
MP
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