Differential Downregulation of β1-Adrenergic Receptor Signaling in the Heart

Background: Chronic sympathetic stimulation drives desensitization and downregulation of β1 Adrenergic Receptor (β₁AR) in heart failure. We aim to explore the differential downregulation subcellular pools of β₁AR signaling in the heart.

Methods and results: We applied chronic infusion of isoproterenol to induced cardiomyopathy in male C57BL/6J mice. We applied confocal and proximity ligation assay to examine β₁AR association with L-type calcium channel, ryanodine receptor 2, and SERCA2a ((Sarco)endoplasmic reticulum calcium ATPase 2a) and Förster resonance energy transfer-based biosensors to probe subcellular β₁AR-PKA (protein kinase A) signaling in ventricular myocytes. Chronic infusion of isoproterenol led to reduced β₁AR protein levels, receptor association with L-type calcium channel and ryanodine receptor 2 measured by proximity ligation (puncta/cell, 29.65 saline versus 14.17 isoproterenol, P<0.05), and receptor-induced PKA signaling at the plasma membrane (Förster resonance energy transfer, 28.9% saline versus 1.9% isoproterenol, P<0.05) and ryanodine receptor 2 complex (Förster resonance energy transfer, 30.2% saline versus 10.6% isoproterenol, P<0.05). However, the β₁AR association with SERCA2a was enhanced (puncta/cell, 51.4 saline versus 87.5 isoproterenol, P<0.05), and the receptor signal was minimally affected. The isoproterenol-infused hearts displayed decreased PDE4D (phosphodiesterase 4D) and PDE3A and increased PDE2A, PDE4A, and PDE4B protein levels. We observed a reduced role of PDE4 and enhanced roles of PDE2 and PDE3 on the β₁AR-PKA activity at the ryanodine receptor 2 complexes and myocyte shortening. Despite the enhanced β₁AR association with SERCA2a, the endogenous norepinephrine-induced signaling was reduced at the SERCA2a complexes. Inhibiting Monoamine Oxidase A rescued the norepinephrine-induced PKA signaling at the SERCA2a and myocyte shortening.

Conclusions: This study reveals distinct mechanisms for the downregulation of subcellular β₁AR signaling in the heart under chronic adrenergic stimulation.

(Sarco)endoplasmic reticulum calcium ATPase 2a; Förster resonance energy transfer; cardiac contractility; phosphodiesterase; protein kinase a; ryanodine receptor; β1 adrenergic receptor.