Snapshot of 5-HT _2A receptor activation in the mouse brain via IP ₁ 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-HT _2A R). However, some aspects of 5-HT _2A R pharmacology remain elusive, such as what molecular drivers differentiate psychedelic from non-psychedelic 5-HT _2A R agonists. We developed an ex vivo platform to obtain snapshots of drug-mediated 5-HT _2A 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 (IP ₁ ) in the mouse brain. The specificity of this method was assessed by comparing IP ₁ levels in homogenates from the frontal cortex in DOI-treated wild-type and 5-HT _2A R-KO animals compared to other brain regions, namely striatum and cerebellum. Furthermore, we encountered that head-twitch response (HTR) counts and IP ₁ in the frontal cortex were correlated. We observed that IP ₁ levels in frontal cortex homogenates from mice treated with LSD and lisuride vary in magnitude, consistent with LSD’s 5-HT _2A R agonism and psychedelic nature, and lisuride’s lack thereof. MDMA evoked an increase of IP ₁ 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-HT _2A R by MDMA via serotonin release from its presynaptic terminals. This methodology enables capturing a snapshot of IP ₁ turnover in the mouse brain that can provide mechanistic insights in the study of psychedelics and other serotonergic agents pharmacodynamics.