PROJECT SUMMARY Chronic pain encompasses a wide range of symptoms and pathways, making the condition notoriously difficult to treat. While opioid treatments can be effective, the unwanted side effects and risk of abuse warrant the investigation of safer treatment options. Cannabis sativa has been utilized for millennia for its medicinal properties and has been studied for its therapeutic potential in the treatment of epilepsy, Alzheimer’s disease, appetite loss, and pain. While most people think of the canonical CB1/CB2 receptors when it comes to cannabinoids, other GPCRs and TRP channels can be modulated by a subset of these ligands. In fact, this project focuses on one of the TRP channels that has been coined an “ionotropic cannabinoid receptor,” TRPV1. TRPV1 (also known as the capsaicin receptor) is a homotetrameric, polymodal channel, and is located in the peripheral nervous system and has been implicated in the perception of pain. This project seeks to understand how and where various cannabinoid ligands bind to and activate TRPV1, in order to aid in rational drug design. The mechanism by which TRPV1 relieves pain is paradoxical; upon activation of TRPV1, ions (preferentially calcium) enter the cell and cause a series of calcium-dependent processes. This generates an action potential which is then propagated to the brain, resulting in the sensation of pain. Shortly after activation, TRPV1 is desensitized, rendering it refractory to any further stimulation, resulting in the paradoxical analgesic effect. Because of this, targeting TRPV1, an ion channel that contributes to the detection of painful stimuli, may be an effective approach in treating chronic pain syndromes. By utilizing computational techniques such as modeling and molecular dynamics simulations, interactions of cannabinoid ligands can be observed with TRPV1, which provide atomic level descriptions that are unavailable to experimental studies. Preliminary data suggests that anandamide may bind to TRPV1 in an allosteric site apart from the location of vanilloid binding. To further investigate this result, a grant application to use the supercomputer ANTON2 was written and awarded, and ~10μs of additional data was gathered thus far. Since this work is part of a combined computational/experimental effort that centers on identifying relevant residues for binding and modes of TRPV1 activation/inactivation by cannabinoid ligands, the results from these simulations will be tested via mutagenesis and functional experimental studies by working in the lab of Dr. Mary Abood at Temple University and collaborating with Dr. Eugen Brailoiu for live cell calcium imaging.