Stella, NephiBruchas, Michael REnglish, Anthony2025-08-012025-08-012025-08-012025English_washington_0250E_28174.pdfhttps://hdl.handle.net/1773/53732Thesis (Ph.D.)--University of Washington, 2025Cannabis has been used by humans for thousands of years for its multi-faceted properties as industrial tools, medicinal effects, and recreational value. And in the past several hundred years, international prohibition movements have been in a struggle with scientists investigating its effects on the human body. Despite this, scientists have persisted to isolate the primary psychoactive compound THC and use it as a basis to uncover an entire neurotransmitter system in humans: the endocannabinoid system. This system plays a critical role in modulating behavior, neurological development, and general brain activity. Further research into the endocannabinoid system and the Cannabis plant lags behind other similar fields despite its importance. The research and findings presented here show another step towards expanding our understanding and functional applications with THC and the endocannabinoid system.Cannabinoid compounds such as THC are notoriously difficult to work with due to their high lipophilicity and, in in vivo rodent studies, their high taste avoidance. Therefore, to counter the potent smells and flavors of raw THC, in an effort to uncover its in vivo behavioral effects. To address this gap, we developed a novel gelatin-based method for THC delivery that enhances palatability and voluntary intake in mice. By incorporating THC into a chocolate-flavored nutritional shake (Ensure™) and gelatinizing the mixture, we formulated an E-gel matrix. In mice, we enhanced voluntary consumption of high-dose THC compared to standard gelatin and succeeded in promoting the mice to consume enough to reach highly psychoactive behavioral effects. The use of Cannabis products containing high concentrations of THC is rapidly increasing, with a growing body of evidence links high-THC Cannabis use with increased psychotic and affective symptoms and Cannabis-associated vehicular accidents. The need for tools to investigate the mechanistic insight into how high-potency THC influences the brain is increasingly relevant. The E-gel model thus provides a translationally relevant, voluntary oral intake paradigm for characterizing THC's pharmacokinetics and behavioral impacts in rodents, offering an essential tool for investigating the consequences of high-dose THC exposure. The endogenous cannabinoid receptor CB1 plays a key role in brain development, but the long-term effects of adolescent THC exposure remain poorly understood. To explore its impact on addiction vulnerability, I exposed mice to THC during adolescence and assessed morphine-related behaviors in adulthood, finding no significant alterations in analgesia or drug-seeking. However, female mice showed THC-dependent impairments in memory recall, suggesting a sex-specific effect on learning. Preliminary behavioral analyses using pose estimation also revealed a unique exploratory behavior in THC-exposed animals, warranting further investigation into the neural circuits underlying these effects. Emerging technologies in other fields provide a pathway to overcome technical and political limitations to enable more sophisticated analyses. In recent years, computer vision tools like DeepLabCut and SLEAP have transformed behavioral research by allowing detailed tracking of individual points on animals during experiments, improving our ability to analyze animal behavior with greater precision than the human eye. Here, we utilized the SLEAP pose estimation algorithm within a linear track system that enables high-resolution and high-frame rate visualization of mice side and bottom-up profiles. Then, animal poses were used to calculate key features that to train dose prediction models capable of identifying the dose of THC animals were treated with solely based on their general or specifically locomotor behavior. These models were then utilized, along with the unsupervised identification of nuanced behaviors, to investigate the effects of THC on excitatory/inhibitory balance of cortical neurons in the mPFC. Inhibitory GABAergic neurons and excitatory glutamatergic neurons of the mPFC were observed to be modified in their activity during THC-impaired motor behavior. Manipulation of CB1R expression and activation of specific neurons time locked to movement with a closed-loop optogenetic model revealed the enhanced GABAergic activity correlates with worsened motor behavior, suggesting the E/I balances criticality in modifying THC-impairment. This effect also greatly modified native endocannabinoid signaling as identified by the novel GRABeCB2.0 senor. These findings marked a novel application of behavioral identification utilized to identify a novel THC-dependent modification in E/I balance of the cortical neurons during motor behavior. Together, the research displayed in this thesis combines advanced computational applications and cutting-edge biosensor technologies to address a pressing challenge in cannabinoid research: quantifying the multi-faceted signatures of THC’s effects in vivo.application/pdfen-USnoneBehaviorCannabisComputer VisionCortexMachine LearningTHCPharmacologyBehavioral sciencesNeurosciencesPharmacologyThesis