Fig. 1.
Distinct signaling properties of lysergic acid diethylamide (LSD) and ergotamine (ERG) at 5-HT1B, 5-HT7A and 5-HT2B receptors. We used luminescence-based assays to measure 5-HT1B receptor mediated Gi activation and cAMP production; fluorescence-based calcium mobilization assays to measure 5-HT2B receptor mediated Gq activation and β-arrestin translocation-dependent luciferase reporter assays to measure 5-HT1B and 5-HT2B receptors mediated β-arrestin recruitment, all in HEK293 derived cells. (A) Normalized concentration-response studies for LSD and ERG at human cloned 5-HT1B receptor-mediated activation of Gi and non-canonical (Arrestin) signaling. (B) Normalized concentration-response studies for LSD and ERG at human cloned 5-HT7A receptor-mediated activation of Gs signaling in the presence and absence of 5-Hydroxytryptamine (5-HT). (C) Normalized concentration-response studies for LSD and ERG at human cloned 5-HT2B receptor-mediated activation of Gq and non-canonical (Arrestin) signaling. The solid circles for LSD-Gs signals are superimposed by the solid squares for ERG-Gs signals and thus, not visible. (D) Mean β-arrestin bias factors were calculated for serotonergic agonists (dihydroergotamine (DHE), methylergonovine (MTE), pergolide (PER) and cabergoline (CAB)), at 5-HT2B and 5-HT1B receptors. Concentration-responses curves were fit to the Black and Leff operational model to obtain transduction coefficients [Log(τ/KA)] for each ligand at each corresponding pathway. The ΔLog(τ/KA) was then calculated with 5-HT as a reference agonist for each pathway, and the ΔΔLog(τ/KA) was calculated between two pathways for each ligand. The bias factor is unit-less and defined as 10ΔΔLog(τ/KA) (28). Compounds with values close to one represent unbiased agonists while compounds with large numerical values, typically >100, represent extremely biased agonists. *p<0.0001 via two-way ANOVA comparing 5-HT2B vs 5-HT1B bias factors; N=3–6 separate experiments. ERG, DHE and, to a lesser extent, LSD, MTE and PER show strong β-arrestin bias at 5-HT2B receptor, but not 5-HT1B receptor.