Constitutively Active Mutants of the alpha sub(1a)- and the alpha sub(1b)-Adrenergic Receptor Subtypes Reveal Coupling to Different Signaling Pathways and Physiological Responses in Rat Cardiac Myocytes

Activation of alpha sub(1)-adrenergic receptors influences both the contractile activity and the growth potential of cardiac myocytes. However, the signaling pathways linking activation of specific alpha sub(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 sub(1A)- and alpha sub(1B)-AR subtypes in cardiac myocytes. For this purpose, a mutant alpha sub(1a)-AR subtype ( alpha sub(1a)-S super(290/293)-AR) was constructed based on analogy to the previously described constitutively active mutant alpha sub(1b)-AR subtype ( alpha sub(1b)-S super(288-294)-AR). The mutant alpha sub(1a)-S super(290/293)-AR subtype displayed constitutive activity based on four criteria. To introduce the constitutively active alpha sub(1)-AR subtypes into cardiac myocytes, recombinant Sindbis viruses encoding either the alpha sub(1a)-S super(290/293)-AR or alpha sub(1b)-S super(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 sub(1a)-S super(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 sub(1b)-S super(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 sub(1a)-AR and the alpha sub(1b)-AR subtypes to different signaling pathways in cardiac myocytes.