Figure 1. Effects of YIL781 on in cellulo signaling of the GHSR1a determined by G-protein mediated Ca²⁺ responsiveness and βarrestin-2 translocation.

(A–B) Ca²⁺ response of the GHSR1a in HEK-293 cells was assessed by bioluminescence in the presence of an aequorin reporter. (A) YIL781 treatment alone over the range of concentrations was modeled by a line with slope = 0 ± 0.0078 and intercept = 0.47 ± 0.0077 (B) The ability of the GHSR1a agonist L-692,585 to activate calcium was inhibited by increasing doses of YIL781, IC₅₀= (1.85 ± 0.093) × 10⁻⁷ M). (C–D) βarrestin-2 translocation to the activated GHSR1a in U2OS cells was measured by assessing the formation of receptor/ βarrestin-2 aggregates using a robotic microscope plate reader. (C) Linear regression of YIL781 treatment over the dose range, Intercept = 18 ± 43, slope = 0.35 ± 6.2. (D) The ability of L-692,585 to induce βarrestin-2 translocation was inhibited by YIL781 with an IC₅₀= (1 ± 0.26) × 10⁻⁶ M. Data were analyzed by GraphPad Prism, version 7.02 and are presented as mean ± sem, N = 3 independent experiments.