Phosphodiesterase 2 and 3 Regulate Compartmentalized Beta2-Adrenergic Receptor Camp Signaling

The two main β-adrenergic receptor (β-AR) subtypes present in cardiac myocytes, β1-AR and β2-AR, both stimulate production of the diffusible second messenger 3',5'-cyclic adenosine monophosphate (cAMP). However, the cAMP signals from the different receptor subtypes result in distinct protein kinase A (PKA) dependent functional responses. While activation of both receptors produces an increase in the force and rate of cardiac contraction, only β1-AR stimulation causes an increase in the rate of relaxation. It is hypothesized that β1-AR, but not β2-AR, produce cAMP that accesses the subcellular location where PKA-dependent phosphorylation of phospholamban occurs. Once phosphorylated, phospholamban reduces its inhibition of sarco/endoplasmic reticulum Ca²⁺-ATPase (SERCA) causing an increase in the rate of cardiac relaxation. Using adult rat ventricular myocytes, this hypothesis was tested by comparing cAMP responses detected by a fluorescence resonance energy transfer (FRET) based biosensor (Epac2-camps) expressed uniformly throughout the cytoplasm of the entire cell and a FRET-based biosensor (Epac2-αKAP) targeted to the SERCA/phospholamban complex. Selective activation of β1-AR or β2-AR produced cAMP responses that could be detected by the Epac2-camps probe. However, only β1-AR selective activation produced a change in cAMP that the Epac2-αKAP probe could detect. Furthermore, selective inhibition of individual phosphodiesterases (PDEs) involved in cAMP metabolism (PDE2, PDE3, or PDE4) increased the basal cAMP detected by both probes. Yet only inhibition of PDE2 or PDE3 allowed a β2-AR cAMP signal to be detected by the Epac2-αKAP probe. These results support the conclusions that cAMP produced by β2-AR stimulation is compartmentalized and that this involves the activity of PDE2 and PDE3.