Altered mRNA abundance of calcium transport genes in cardiac myocytes induced by angiotensin II.
Recent studies have implicated angiotensin II (angiotensin) in the pathogenesis of cardiac hypertrophy and heart failure. Heart failure is associated with alterations in intracellular Ca2+ movements mediated by sarcolemmal (SL) and sarcoplasmic reticular (SR) membranes in cardiac myocytes. As it was suspected that alteration gene expression of proteins responsible for controlling transmembrane Ca2+ fluxes may contribute to loss of Ca2+ homeostasis in failing hearts, we undertook a study of the effect of angiotensin on the expression of some target genes in the myocardium. Specifically, we tested the effect of angiotensin on mRNA abundance of cardiac Ca(2+)-transport genes including SL Na+/Ca2+ exchange (EX), SR ryanodine receptor (RYR), and sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA). The mRNA abundance of target gene was assessed by Northern blot assay in (i) direct hormonal stimulation of cultured isolated neonatal and adult rat myocytes and (ii) adult rat hearts after implantation of osmotic mini-pumps for delivery of hormone. In all experiments, Northern blot data were normalized using cDNA (Glyceraldehyde 3-phosphate dehydrogenase signal, GAPDH) hybridization to RNA samples. The results indicate that the ratios of EX/GAPDH, RYR/GAPDH, and SERCA2/GAPDH signals were decreased by 51.6%, 55.0%, and 49.4% respectively after neonatal cardiac myocytes were treated (24 h) with 10(-7) M angiotensin. These decreases were blocked completely by treatment with angiotensin subtype 1 (AT1) receptor antagonist (losartan), whereas angiotensin subtype 2 (AT2) receptor antagonist (PD123319) treatment had no effect on the angiotensin-mediated decrease in target gene mRNA abundance. In contrast, angiotensin had no effect on EX, RYR nor SERCA2 gene mRNA abundance in cultured adult myocytes. In a separate series of experiments wherein adult male Sprague-Dawley rats were infused with different dose of angiotensin for 3 days via osmotic mini-pump, we did not detect any alterations in mRNA abundance of cardiac EX/GAPDH, RYR/GAPDH or SERCA/GAPDH genes in either left or right ventricular samples. Thus our results indicate that, in neonatal rat myocytes, angiotensin affects SL and SR calcium transport gene expression by direct agonism of AT1-receptors. As the infusion of low and high dose angiotensin did not affect the expression of target genes in adult hearts, we suggest that the mechanisms for transduction of the angiotensin signaling in neonatal and adult myocytes may be different and may depend on the stage of development. We conclude that regulation of myocardial Ca(2+)-transport gene mRNA abundance by angiotensin may differ among neonatal and adult animals. Nonetheless, our finding with respect to neonatal preparation led us to believe that in neonatal myocytes, the mRNA abundance of SL Na+/Ca2+ exchange, SR ryanodine receptor, and SR Ca(2+)-ATPase are all decreased in response to stimulation by angiotensin.
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Division of Cardiovascular Sciences, St Boniface General Hospital Research Centre, University of Manitoba, Winnipeg, Canada.
Scammel-La Fleur
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Dixon
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D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000096:biosynthesis* / Q000235:genetics; D000252:Calcium-Transporting ATPases / Q000096:biosynthesis* / Q000235:genetics; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000187:drug effects; D017136:Ion Transport / Q000187:drug effects; D008297:Male; D009124:Muscle Proteins / Q000096:biosynthesis* / Q000235:genetics; D009206:Myocardium / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D019837:Ryanodine Receptor Calcium Release Channel; D014662:Vasoconstrictor Agents / Q000494:pharmacology*
D015220:Calcium Channels; D009124:Muscle Proteins; D012333:RNA, Messenger; D019837:Ryanodine Receptor Calcium Release Channel; D014662:Vasoconstrictor Agents; D000804:Angiotensin II; D000252:Calcium-Transporting ATPases; D002118:Calcium