"Characterization of the Effects and Mechanism of Action of Psilocybin in a Genetic Model of Autism Spectrum Disorder"

Autism Spectrum Disorder (ASD) is a neurodevelopmental condition characterized by persistent difficulties in social communication and interaction, accompanied by repetitive behaviors and restricted interests. These alterations emerge early in life and significantly affect the quality of life of individuals with ASD and their families. Among the genetic causes of autism, Fragile X Syndrome (FXS) represents the most common monogenic form. Loss of the FMRP protein in FXS profoundly disrupts synaptic plasticity, leading to cognitive and behavioral impairments that overlap with the autistic phenotype, thereby making FXS models valuable tools for investigating disease mechanisms and potential therapeutic interventions. Among the neurobiological systems implicated in ASD and FXS, the serotonergic system plays a central role in regulating neuronal plasticity, cognition, and social behavior. Alterations in serotonin synthesis and signaling, documented in both conditions, suggest that dysregulated serotonergic neurotransmission contributes to specific neurobehavioral dysfunctions. However, to date, no pharmacological treatment is currently available for the core symptoms of ASD or FXS. In recent years, classic psychedelics, particularly psilocybin, have gained scientific interest due to their ability to rapidly induce long-lasting forms of synaptic plasticity, raising the possibility that such compounds may offer innovative therapeutic strategies for disorders characterized by impaired plasticity. The overarching aim of this doctoral thesis was to investigate the effects of psilocybin in the Fmr1- Δ exon 8 rat model, a validated preclinical model of both ASD and FXS, and to identify its potential mechanism of action, with particular focus on serotonergic signaling and neurotrophic BDNF/TrkB pathway. During the first year, I contributed to the behavioral characterization of the Fmr1- Δ exon 8 rat model, assessing recognition memory, social behavior, exploratory activity, and stereotypies. In parallel, I participated in studies focusing on the characterization of the endocannabinoid and dopaminergic systems in this rat model, which helped to refine the neurobehavioral phenotype of Fmr1- Δ exon 8 rats. In the second year, my research focused on elucidating the mechanism of action of psilocybin. After confirming its beneficial effects on short-term recognition memory in Fmr1- Δ exon 8 rats, I employed a pharmacological approach involving pretreatment with antagonists of the 5-HT2A, 5-HT1A, and TrkB receptors. The results demonstrated that psilocybin’s cognitive rescue in Fmr1- Δ exon 8 rats is not mediated by canonical serotonergic receptors, but instead requires activation of the BDNF/TrkB pathway, accompanied by the restoration of plasticity-related markers in the prefrontal cortex. In the third year, I expanded the investigation by evaluating the effects of psilocybin in Fmr1- Δ exon 8 rats across different developmental stages (adolescence and adulthood), including sex-dependent analyses during adolescence, and by assessing long-term effects after treatment. During a six-month research period at the University of Central Florida, under the supervision of Dr. Kiminobu Sugaya, I conducted an exploratory analysis of peripheral biomarkers in Fmr1- Δ exon 8 rats by quantifying serotonin levels in plasma, total extracellular vesicles (EVs), and neuron-derived exosomes isolated via anti-NCAM immunoprecipitation. These preliminary findings suggest peripheral alterations consistent with the FXS phenotype and potential psilocybin-induced modulations of EV-associated serotonin. Overall, the experiments performed during my PhD project contributed to characterize the Fmr1- Δ exon 8 rat model of ASD and demonstrate that psilocybin effectively restores cognitive deficits in this preclinical model through a neurotrophic mechanism dependent on BDNF/TrkB. At the same time, its effects on other behavioral domains vary according to sex and developmental stage. By integrating behavioral, pharmacological, molecular, and peripheral data, this work provides new evidence for the therapeutic potential of classic psychedelics in neurodevelopmental disorders and contributes to establishing the foundation for future translational applications of psilocybin.