Differential Regulation of Inflammatory Responses Following 5‐HT2 Receptor Activation in Pulmonary Tissues
Authors: Flanagan Thomas W. , Billac Gerald , Nichols Charles D. 
Published: May 2022

IntroductionThe concept of functional selectivity and psychedelics is often regarded as the ability of different 5‐HT2A receptor agonist compounds to exert varying degrees of psychoactivity in the central nervous system. However, the distribution and expression of the 5‐HT2A receptor, the primary pharmacological target for psychedelics, in peripheral and immune‐related tissue suggests selective signaling may occur here as well. Using an ovalbumin (OVA)‐induced model of rodent acute asthma, our lab has previously shown that subtle changes to the 2,5‐dimethoxy‐phenethylamine pharmacophore structure can profoundly impact the ability of these compounds to prevent asthmatic exacerbations. Here, through both in vitroand in vivoassays, we interrogate elements of functional selectivity in peripheral tissues following exposure to the 5‐HT2 receptor selective agonists (R)‐2,5‐dimethoxy‐4‐iodoamphetamine [(R)‐DOI] and 2,5‐dimethoxy‐4‐trifluoromethylamphetamine (DOTFM). (R)‐DOI is a full and potent anti‐inflammatory compound, whereas DOTFM has no measurable anti‐inflammatory activity. We find that elements of selective signaling combined with modulation to overall chromatin structure and downstream protein expression provide mechanistic clues regarding how psychedelics may elicit their long‐lasting, therapeutic effects.ObjectiveTo elucidate mechanistic variations between anti‐inflammatory and non‐anti‐inflammatory 5‐HT2 receptor agonists.MethodsWe utilized fluorescence recovery after photobleaching (FRAP) to probe the kinetics of the linker histone H1c transiently transfected into a HEK293 cell line stably expressing the human 5‐HT2A receptor, and calcium mobilization assays in the same cell line to determine differences in signaling between (R)‐DOI and DOTFM. For in vivoassays we measured enhanced pause (PenH) values for BALB/c mice challenged with OVA and treated with either (R)‐DOI or DOTFM, using PenH values as a proxy measure for pulmonary inflammation. Lungs were removed en bloc,processed, and analyzed by qPCR and quantitative proteomics using tandem tag mass spectrometry (TMT).ResultsIn vitro assays indicate modest differences irrespective of phenethylamine compound, whereas whole‐body plethysmography (WBP), qPRC, and proteomic analysis reveal significant differential regulation of proteins relevant for both asthmatic initiation and chromatin compaction dependent upon 5‐HT2A agonist.ConclusionAs 5‐HT2‐mediated expression of asthma‐relevant proteins are differentially regulated by either (R)‐DOI and DOTFM, the downregulation of inflammatory pathways via activation of the 5‐HT2 receptor likely requires an accessible, non‐sterically hindered subunit at the 4‐position of the standard 2,5‐dimethoxy‐ampetamine structure to maximally induce anti‐inflammatory effects. Changes to both linker histone mobility and the expression of proteins relevant to chromatin architecture suggest that modulation to the overall chromatin structure combined with downstream epigenetic regulation is a potential mechanism by which certain 5‐HT2A agonists can exert their therapeutic effects.