G i-Dependent Localization of β2-Adrenergic Receptor Signaling to L-Type Ca 2+ Channels

Abstract

A plausible determinant of the specificity of receptor signaling is the cellular compartment over which the signal is broadcast. In rat heart, stimulation of β 1-adrenergic receptor ( β 1-AR), coupled to G s-protein, or β 2-AR, coupled to G s- and G i-proteins, both increase L-type Ca 2+ current, causing enhanced contractile strength. But only β 1-AR stimulation increases the phosphorylation of phospholamban, troponin-I, and C-protein, causing accelerated muscle relaxation and reduced myofilament sensitivity to Ca 2+. β 2-AR stimulation does not affect any of these intracellular proteins. We hypothesized that β 2-AR signaling might be localized to the cell membrane. Thus we examined the spatial range and characteristics of β 1-AR and β 2-AR signaling on their common effector, L-type Ca 2+ channels. Using the cell-attached patch-clamp technique, we show that stimulation of β 1-AR or β 2-AR in the patch membrane, by adding agonist into patch pipette, both activated the channels in the patch. But when the agonist was applied to the membrane outside the patch pipette, only β 1-AR stimulation activated the channels. Thus, β 1-AR signaling to the channels is diffusive through cytosol, whereas β 2-AR signaling is localized to the cell membrane. Furthermore, activation of G i is essential to the localization of β 2-AR signaling because in pertussis toxin-treated cells, β 2-AR signaling becomes diffusive. Our results suggest that the dual coupling of β 2-AR to both G s- and G i-proteins leads to a highly localized β 2-AR signaling pathway to modulate sarcolemmal L-type Ca 2+ channels in rat ventricular myocytes.