Pathway-selective signaling of serotonergic psychedelics at 5-HT receptors: Implications for psychoactivity, safety, and therapeutic potential

Serotonergic psychedelics primarily exert their effects through activation of the serotonin (5-HT) 2A receptor (5-HT 2A R). However, the full pharmacological profiles of these substances involve multiple serotonin receptor subtypes and intracellular signaling pathways. This study characterized the signaling behavior of a panel of serotonergic psychedelics using stable cell lines and a series of bioassays targeting phospholipase C (PLC) activation via inositol monophosphate (IP1) formation, phospholipase A 2 (PLA 2 ) activation, β-arrestin2 recruitment, and Gα i -protein dissociation across 5-HT 2A , 5-HT 2B , 5-HT 2C , and 5-HT 1A receptors. The results of the study indicate that IP1 formation offers the most robust and reproducible measure of 5-HT 2 R receptor activation and aligns most closely with human dosing data. Particularly for the 5-HT 2A R, PLC-IP1 activation exhibited a strong correlation with known psychoactive doses, contrasting with assays that relied on Ca²⁺ release. The majority of psychedelics exhibited a signaling bias toward the PLC-IP1 or the PLA 2 -AA pathway at the 5-HT₂ A R. The 5-HT 2A /5-HT 1A activation ratio may provide information about seizure risk and therapeutic potential. Furthermore, the low activation efficacy at the 5-HT₂ B R potentially suggests reduced cardiac risk with intermittent use, and the strong activation of the 5-HT₂ C R is in accordance with the reported low abuse potential of serotonergic psychedelics. This study underscores the importance of pathway-specific profiling in understanding the pharmacology of serotonergic psychedelics. It provides a framework for predicting efficacy and adverse effect potential and lays the groundwork for the rational design of next-generation psychedelics with improved safety and efficacy.