FIGURE 5.

Co-expression of D₁ or MC₃ attenuates GHS-R1a mediated signaling in Hek293A cells. Ghrelin-mediated intracellular calcium increase (A) is significantly reduced when the D₁ receptor (B) or the MC₃ receptor (C) is co-expressed with the GHS-R1a receptor in Hek293A cells. The cells were incubated for 30 min with 100 nm ghrelin, 10 μm 6,7-ADTN hydrobromide, [Nle⁴,d-Phe⁷]-α-MSH, or a combination of both, and intracellular calcium influx was measured. Intracellular calcium increase was depicted as a percentage of maximal calcium increase as elicited by control (3% FBS). The data are representative figures from three independent experiments depicting the means ± S.E. with each concentration point performed in triplicate. Statistical significance was analyzed using a two-way ANOVA followed by Bonferroni multiple comparison test; statistical significance is notated as follows: ***, p < 0.001; and **, p < 0.01.