β 2 -Adrenergic cAMP Signaling Is Uncoupled From Phosphorylation of Cytoplasmic Proteins in Canine Heart

Abstract

Background —Recent studies of β-adrenergic receptor (β-AR) subtype signaling in in vitro preparations have raised doubts as to whether the cAMP/protein kinase A (PKA) signaling is activated in the same manner in response to β 2 -AR versus β 1 -AR stimulation. Methods and Results —The present study compared, in the intact dog, the magnitude and characteristics of chronotropic, inotropic, and lusitropic effects of cAMP accumulation, PKA activation, and PKA-dependent phosphorylation of key effector proteins in response to β-AR subtype stimulation. In addition, many of these parameters and L-type Ca 2+ current ( I Ca ) were also measured in single canine ventricular myocytes. The results indicate that although the cAMP/PKA-dependent phosphorylation cascade activated by β 1 -AR stimulation could explain the resultant modulation of cardiac function, substantial β 2 -AR–mediated chronotropic, inotropic, and lusitropic responses occurred in the absence of PKA activation and phosphorylation of nonsarcolemmal proteins, including phospholamban, troponin I, C protein, and glycogen phosphorylase kinase. However, in single canine myocytes, we found that β 2 -AR–stimulated increases in both I Ca and contraction were abolished by PKA inhibition. Thus, the β 2 -AR–directed cAMP/PKA signaling modulates sarcolemmal L-type Ca 2+ channels but does not regulate PKA-dependent phosphorylation of cytoplasmic proteins. Conclusions —These results indicate that the dissociation of β 2 -AR signaling from cAMP regulatory systems is only apparent and that β 2 -AR–stimulated cAMP/PKA signaling is uncoupled from phosphorylation of nonsarcolemmal regulatory proteins involved in excitation-contraction coupling.