Hallucinogenic drugs of abuse are partial agonists at the serotonin 5- HT2A receptor. The molecular mechanisms mediating their hallucinogenic potential are not known. Utilizing the understanding about hallucinogen interactions with the H-HT/2A receptor that we have developed from our previous inter-disciplinary work in an IRPG, we propose to study the specific conformational effects induced by hallucinogenic and non-hallucinogenic serotonergic agonists acting at the 5-HT2A receptor. Agonist binding to a receptor stabilizes the protein in a particular relative distribution of conformations. Each conformational state has a propensity to interact with specific G-proteins, inducing second messenger responses and activation of endogenous genes, and with regulatory proteins, leading to phosphorylation, internalization, and desensitization. These downstream effects will be assayed as reflections on the conformational stages stabilized by a particular agonist. Full and partial 5-HT/2A agonists that differ in their hallucinogenic potential will be studied and the basis for differences in the responses observed will be investigated. Mutations of the H-HT2A receptor that modify relative agonist positioning and efficacy, relative G-protein activation, and stability of the inactive stage have been identified and their effects on the responses elicited in cell lines by different hallucinogens and non- hallucinogens will be determined. In order to assay cellular responses both in cell lines and in neurons in vivo, we will assay intrinsic reporters of cellular signaling (IRC) that manifest rapid alterations in response to signal transduction activation. IRC responses to hallucinogens and non- hallucinogens will be measured in cortical neurons in knock-in mice expressing the human wild-type and mutant receptors in place of the endogenous receptor. These studies will provide insight into the molecular basis for the cellular responses to hallucinogenic chemicals elicited at the 5-HT/2A receptor and may facilitate the development of new therapeutic modalities.
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