Snapshot of 5-HT2A receptor activation in the mouse brain via IP1 detection

The distinct subjective effects that define psychedelics such as LSD, psilocybin or DOI as drug class are causally linked to activation of the serotonin 2A receptor (5-HT2A R). However, some aspects of 5-HT2A R pharmacology remain elusive, such as what molecular drivers differentiate psychedelic from non-psychedelic 5-HT2A R agonists. We developed an ex vivo platform to obtain snapshots of drug-mediated 5-HT2A R engagement of the canonical G q/11 pathway in native tissue. This non-radioactive methodology captures the pharmacokinetic and pharmacodynamic events leading up to changes in inositol monophosphate (IP1 ) in the mouse brain. The specificity of this method was assessed by comparing IP1 levels in homogenates from the frontal cortex in DOI-treated wild-type and 5-HT2A R-KO animals compared to other brain regions, namely striatum and cerebellum. Furthermore, we encountered that head-twitch response (HTR) counts and IP1 in the frontal cortex were correlated. We observed that IP1 levels in frontal cortex homogenates from mice treated with LSD and lisuride vary in magnitude, consistent with LSD’s 5-HT2A R agonism and psychedelic nature, and lisuride’s lack thereof. MDMA evoked an increase of IP1 signal in the frontal cortex that were not matched by the serotonin precursor 5-HTP or the serotonin reuptake inhibitor fluoxetine. We attribute differences in the readout primarily to the indirect stimulation of 5-HT2A R by MDMA via serotonin release from its presynaptic terminals. This methodology enables capturing a snapshot of IP1 turnover in the mouse brain that can provide mechanistic insights in the study of psychedelics and other serotonergic agents pharmacodynamics.