Abstract
Inhibition of agonist-induced M(2) muscarinic receptor (M(2)R) activation by functional anti-M(2)R autoantibodies has been associated with cardiac parasympathetic dysfunction in patients with chronic Chagas disease (CD). This study explored the allosteric nature of that inhibitory effect by assessing the ability of serum IgG from patients with CD and dysautonomia (DCD IgG) to modulate the interaction between M(2)R and β-arrestins in HEK 293T cells using bioluminescence resonance energy transfer. DCD IgG alone did not stimulate arrestin-2 or arrestin-3 recruitment. When cells were preincubated with DCD IgG and then treated with carbachol, arrestin-2 translocation decreased in a concentration-dependent manner, while arrestin-3 recruitment remained unaffected. Inhibition curve analysis showed a submaximal inhibitory effect (68.1 ± 2.4%) and a Hill slope less than -1 (-4.03 ± 0.39). Carbachol concentration-response assays after preincubation with DCD IgG revealed a noncompetitive inhibition of arrestin-2 recruitment, with no change in arrestin-3 translocation. Unlikely, simultaneous exposure to DCD IgG and carbachol potentiated agonist-induced Arr-2 recruitment. We conclude that anti-M(2)R autoantibodies selectively inhibit agonist-induced arrestin-2 recruitment, acting as negative allosteric modulators of agonist efficacy. The direction of autoantibody-induced allosteric modulation depends on the timing of IgG application relative to the agonist and the duration of receptor exposure to autoantibodies.