Constitutively active mutants of the alpha(1a)- and the alpha(1b)-adrenergic receptor subtypes reveal coupling to different signaling pathways and physiological responses in rat cardiac myocytes.
Activation of alpha(1)-adrenergic receptors influences both the contractile activity and the growth potential of cardiac myocytes. However, the signaling pathways linking activation of specific alpha(1)-adrenergic receptor (AR) subtypes to these physiological responses remain controversial. In the present study, a molecular approach was used to identify conclusively the signaling pathways activated in response to the individual alpha(1A)- and alpha(1B)-AR subtypes in cardiac myocytes. For this purpose, a mutant alpha(1a)-AR subtype (alpha(1a)-S(290/293)-AR) was constructed based on analogy to the previously described constitutively active mutant alpha(1b)-AR subtype (alpha(1b)-S(288-294)-AR). The mutant alpha(1a)-S(290/293)-AR subtype displayed constitutive activity based on four criteria. To introduce the constitutively active alpha(1)-AR subtypes into cardiac myocytes, recombinant Sindbis viruses encoding either the alpha(1a)-S(290/293)-AR or alpha(1b)-S(288-294)-AR subtype were used to infect the whole cell population with >90% efficiency, thereby allowing the biochemical activities of the various signaling pathways to be measured. When expressed at comparable levels, the alpha(1a)-S(290/293)-AR subtype exhibited a significantly elevated basal level as well as agonist-stimulated level of inositol phosphate accumulation, coincident with activation of atrial natriuretic factor-luciferase gene expression. By contrast, the alpha(1b)-S(288-294)-AR subtype displayed a markedly increased serum response element-luciferase gene expression but no activation of atrial natriuretic factor-luciferase gene expression. Taken together, this study provides the first molecular evidence for coupling of the alpha(1a)-AR and the alpha(1b)-AR subtypes to different signaling pathways in cardiac myocytes.
McWhinney
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Henry Hood Research Program, Pennsylvania State College of Medicine, Danville, Pennsylvania 17822-2614, USA.
Wenham
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Kanwal
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Kalman
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Hansen
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Robishaw
J D
JD
D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D019556:COS Cells; D002417:Cattle; D002478:Cells, Cultured; D004789:Enzyme Activation; D017930:Genes, Reporter; D006868:Hydrolysis; D008156:Luciferases / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009206:Myocardium / Q000378:metabolism*; D010656:Phenylephrine / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D018340:Receptors, Adrenergic, alpha-1 / Q000235:genetics* / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection
C547715:Adra1a protein, rat; C547718:Adra1b protein, rat; D000316:Adrenergic alpha-Agonists; D000317:Adrenergic alpha-Antagonists; D010716:Phosphatidylinositols; D018340:Receptors, Adrenergic, alpha-1; D010656:Phenylephrine; D008156:Luciferases; D020929:Mitogen-Activated Protein Kinase Kinases